Latest Developments

Welcome to our new section of MoragaBiotech.com. We intend to provide our visitors with breaking news about the latest developments being advanced throughout the world in adult stem cell technologies. If you periodically visit this site in the very near future, we should have our public forum up and running.

The goal of the forum is to provide the visitor with the opportunity to ask questions and give their opinions on the results being published by adult stem cell investigators. Hopefully, this forum will also become an invaluable source for scientists to ask anonymous trouble-shooting research questions that can be answered by others, while serving as a world wide, interactive, shared information exchange for adult stem cell research scientists. The forum will be indexed and archived as a public service.

Until we have constructed the Forum section on our website, please submit via e-mail any breaking news we may have missed and you find noteworthy for your fellow investigators to NewsnViews@moragabiotech.com. If appropriate, don't forget to include links to other websites.

We will begin this section by providing a monograph by the Company's scientific founders as a technical primer to adult stem cells. Click here for the monograph file.

News and Views

October 2, 2006---In the September 28th online edition of Stem Cells, Bresagen's scientists and collaborators from Univ. of Georgia and Georgia Institute of Technology, published their findings on the relationship between the extracellular markers, SSEA-3 and SSEA-4, and pluripotency in human embryonic stem cells. These surface epitopes are glycosphingolipids and their expression is tightly regulated during early embryonic development. The investigators used inhibitors to block the biosynthesis of these antigens and confirmed their absence by immunostaining, flow cytometry, and tandem mass spectrometry. Analysis of differentiation in vitro and in teratomas suggested that the SSEA-3 and SSEA-4 were not essential for maintaining pluripotency in embryonic cells. However, the researchers did not rule out that genes encoding these antigens may have a significant role in regulating cellular differentiation.

September 29, 2006---In the September 28th online edition of Stem Cells, scientists from University of North Carolina reported on the negative impacts of using monoclonal antibodies to isolate and enrich mouse hemapoietic stem cells (HSCs) for engraftment. Considering that antibodies are known to alter the physiology of other cell types, the investigators compared a side population (SP) of bone marrow-derived HSCs enriched by Hoechst 33343 dye exclusion to SP cells treated with an antibody cocktail based on the stem cell markers Sca-1+ c-Ki+Lin- (SKL). The researchers found that HSCs treated with the antibody cocktail had decreased expression of the stem cell genes c-Kit, CD34, Tal-1, and Slamf1 compared to the untreated HSCs. Additionally, it was found that the antibody-treated cells had lost their ability to confer long-term hematopoietic engraftment in lethally irradiated mice. Thus, the authors concluded that isolation procedures based on use of antibodies may have negative effects on HSC physiology.

September, 28, 2006---In the September 9th edition of Stem Cells, University of Cambridge's P. E. Hall et al. reported their study on integrins as markers for human neural stem cells. The study consisted of using fluorescence-activated cell sorting (FACS) to enrich for a population of cells which expressed either the α6^hi or β1^hi integrin receptors. The β1^hi FACS-enriched population was reported to contain neural precursor cells and shown to express prominin-1, SOX2, SOX3, nestin, Bmi1, Musashi1, and form neurospheres in vitro. It was further noted that both the β1^hi and prominin-1^hi cells were equally capable of forming neurospheres. The authors concluded that both β1 and α6 integrin subunits provide convenient markers for isolating human neural progenitor stem cells.

September 27, 2006---In the August issue of Stem Cells and Development, E. Price et al. from the University of Missouri reported the isolation of multipotent self-renewing stem cells from the peripheral blood of GFP transgenic pigs. The investigators reported that cells were propagated as nonadherent spheroids with the potential to differentiate into angiogenic, osteogenic, adipogenic and neurogenic phenotypes. The researchers used different inductive agents to induce the cells down various mesodermal and ectodermal lineages. Following neural differentiation, the cells were transplanted into the rat brain. The xenograft survived and migrated throughout the striatum and corpus callosum. The stem cells were propagated for more than 100 doublings without apparently losing their plasticity. The authors concluded that these peripheral blood multipotent adult progenitor cells could provide a superior alternative to embryonic stem cells in regenerative medicine. (Price et al. results confirm our observation of highly plastic stem cells circulating in the peripheral blood of mammals. See our news release on the home page.)

September 26, 2006---Following up on yesterday's post, we would like to provide additional discussion of the research paper by T. Thatava et al. published in the September 21st online edition of Stem Cells. First, it should be noted that the authors concluded that they were able to "reprogram" mouse bone marrow-derived stem cells by culturing the cells in Trichostatin A (TSA). TSA is a hydroxamic acid and potent inhibitor of histone deacetylase. This method of deacetylation of the heterochromatin and subsequent cultivation of the stem cells in the presence of GLP-1 (glucagon-like peptide -1) resulted in the formation of pancreatic-like islet clusters. Second, the investigators provided data demonstrating that the resultant clusters were insulin-producing islet β-cells. With RT-PCR assays and other methods, the authors demonstrated that the cells express an endodermal gene expression profile and used this finding to support the conclusion that the islet-like clusters consist of insulin-producing β-cells. Finally, the investigators provided data supporting their findings that the differentiated cells were glucose responsive by analyzing the culture media for insulin and C-peptide with ELISA and Western blot assays (following glucose challenge), respectively. Although the data provided by the investigators were quite impressive with respect to inducing bone marrow-derived stem cells down the pancreatic lineage, it is unclear how functional the resultant cells were in secreting insulin following exposure to different concentrations of glucose. Since data were not provided comparing the amount of insulin secreted into the culture media by the islet cluster to that of native islets, it is difficult to assess the functionality of these cells. Another question is whether the investigators had actually reprogrammed the stem cells by modifying their chromatin or was it really a matter of activating a quiescent population of resident stem cells in the bone-marrow?

September 25, 2006---In the September 21st online edition of Stem Cells, T. Thatava et al. from the German Res. Center for Biotechnology published their findings of using histone deaceytlase inhibitor, Trichostatin A (TSA), to induce bone marrow-derived stem cells to differentiate into pancreatic islets. In cell cultures with TSA, the investigators were able to form islet-like clusters with cells in the cluster secreting insulin as well as glucagon and somatostatin. Endodermal genes of the pancreatic lineage such as PDX-1, GLUT-2, and PAX 4 were also upregulated in these clusters. Western blot analysis and electron microscopy confirmed the presence of insulin granules within the secretory vesicles of cells comprising the islet-like clusters. The authors concluded that chromatin remodeling can induce bone marrow-derived stem cells into insulin-producing beta cells.

September 22, 2006---J. van Tuyn et al. from Leiden University Medical Center in The Netherlands reported in the September 21st online edition of Stem Cells their study on a subset of epicardial cells which can undergo an epithelial-to-mesenchymal transition (EMT). The investigators found these cells were able to invade the myocardium and differentiate into various mesodermal cell types such as coronary smooth muscles cells and other tissues of the heart. In the study, primary cultures of adult epicardial cells were analyzed for their immunophenotype, transcriptome, transducibility, and differentiation potential. The researchers found that the epicardial cells in vitro spontaneously undergo EMT where the cells closely resemble subepithelial fibroblasts. They further noted that only the epicardium-derived cells (EPDCs) express the cardiomyocyte markers GATA4 and troponin T. EPDCs transfected with the gene encoding either TGF-b or BMP2 obtained the smooth muscle cell's phenotype. The authors concluded that "epicardial cells from human adults recapitulate at least part of the differentiation potential of their embryonic counterparts" and can serve as a model to explore their therapeutic potential. (The study does not address a major concern of whether there was a mixed population of cells found in the subset of the isolated epicardial cells that were cultivated. Another explanation to the reported observations may possibly be that the investigators were co-culturing a more primitive pluripotent stem cell population.)

September 20, 2006---In the September 19th edition of PNAS, A. M. Kocabas et al. published findings from the analysis of the transcriptome of human oocytes. With human oocytes in metaphase II, they measured the total expressed RNA in these oocytes and compared their gene expression profile to the transcriptome of 10 different normal human tissues. The assay consisted of Affymetrix gene chip analysis (Affymetrix Human Genome U133 Plus 2.0 Gene Chip arrays). The researchers found in the oocytes that 5,331 transcripts were significantly up-regulated and 7,074 transcripts significantly down-regulated in comparison to normal tissues. The authors noted that their report provides a "comprehensive expression baseline of genes expressed in in vivo matured human oocytes." They further added that "understanding of the biological role of these genes may expand our knowledge on meiotic cell cycle, fertilization, chromatin remodeling, lineage commitment, pluripotency, tissue regeneration, and morphogenesis." (This paper is the beginning of delineating the proteomics roadmap that will in the future allow one to find those critical proteins that induce a somatic cell's genome to reset itself back to the embryonic pluripotent state during nuclear transfer experiments.)

September 19, 2006---In the Sept. 14th online edition of Stem Cells, Q. He et al. from Queen's University in Belfast, Ireland published an in depth review article on "Multipotent mesenchymal stromal cells in blood." The peripheral blood-derived multi-potent mesenchymal stromal cells (PBMSCs) are defined as cells isolated from adult peripheral blood by primary cultures as a discrete adherent cell population consisting of fibroblast-like cells. Additionally, the authors noted that bone marrow-derived mesenchymal stem cells (BMMSCs) shared many of the characteristics of PBMSCs by their behavior in culture, plasticity, and immunophenotypic surface markers. However, the authors noted that human PBMSCs are negative for the stromal marker, Stro-1, whereas it is a widely used marker to purify BMMSCs. The authors also pointed out that PBMSCs can be enriched in a population of CD133+ cells. Although the PBMSCs are found in very low frequency in circulating blood, they are clonegenic and express the pluripotent embryonic transcription factor Oct4. The authors concluded that knowledge of PBMSCs is still very limited due to the difficulty in isolating these cells. The authors also pointed out that before these cells can be used for cell-based therapies, the technical hurdles such as optimizing culture conditions to grow out a pure population of these rare cells and ascertaining unique surface markers to characterize and study these cells must first be cleared by investigators.

September 18, 2006---The American Chemical Society held their 232nd Annual Meeting last week in San Francisco. There were a number of papers and posters focused on stem cells and regenerative medicine. An interesting presentation (BIOT 46, Thursday, Sept. 14, 3:10 PM) presented by University of Pennsylvania's D. Discher and A. Engler demonstrated that the elasticity/rigidity of the microenvironment affects the signaling pathway that drives stem cells down a particular tissue lineage. The investigators cultured mesenchymal stem cells in various polymer hydrogels with either soft, medium or rigid elasticity. They found that the stem cells grown in a softer environment, which mimic the elasticity of brain tissues, tend to form nerve-like cells. Similarly, cells grown in a medium elastic environment tend to form muscle-like cells, whereas those cells grown in a rigid environment tend to form bone-like cells. Assaying for messenger RNA and protein markers, the researchers found that both the cells' shape (mechanical structure) and the the chemical environment have a significant role in determining a stem cell's fate during differentiation in cell cultures.

September 15, 2006---Scientists from Washington University School of Medicine reported in the August 15th online edition of Blood their study comparing a CXCR4 antagonist, AMD3100, and G-CSF in their ability to mobilize endothelial progenitor cells (EPC) into peripheral blood. When mice were treated with AMD3100 or G-CSF, the study's results demonstrated that the number of circulating angiogenic cells (CAC) in the blood increased by 10.0 and 8.8 fold, respectively. The number of circulating EPC in animals treated with the drugs increased 10 fold with AMD3100 and 21 fold with G-CSF. Interestingly, mobilized EPC had a greater proliferative potential than EPC isolated from untreated blood. Additionally, cryopreservation retained functional activity for CAC, but not for EPC. The authors concluded that "AMD3100 is a potent and rapid mobilizer of angiogenic cells and demonstrates the feasibility of obtaining and storing a large number of CAC by leukapheresis."

September 14, 2006---In the Sept. 7th online edition of Stem Cells, Japanese scientists from Sapporo Medical University reported a novel method for culturing CD34⁺ cells from human cord blood in order to generate platelets on a large scale. The investigators were able to generate over 1.5 x10¹¹ platelets from a unit of cord blood (5 million CD34⁺ cells). The researchers used a complex three-phase culture system in which the CD34⁺ cells were initially co-cultured with human stromal cells that had been transduced with the telomerase gene in a serum-free medium supplemented with stem cell factor (SCF), Flt-3/Flk-2 ligand and thrombopoietin (TPO). The differentiated platelets were found to exhibit a normal morphology (by electron micrographs). The investigators also demonstrated that the platelets were functional by their ability to express P-selectin and gpIIb/IIIa epitopes when exposed to fibrinogen/ADP.

September 13, 2006---Canda's J. Trowbridge et al. from the University of Western Ontario published in the Sept. 12th online edition of PNAS their study on the Hedgehog (Hh) gene and its role in modulating hematopoietic stem cells (HSC). The investigators found that downstream activation of the Hh gene signaling pathway induced primitive HSC cycling and proliferation within the bone marrow's tissue compartment during conditions of both homeostasis and acute tissue regeneration. However, the researchers found that continued activation of the Hh signal pathway was at the expense of HSC function. Long term activation results in suppression of specific cell cycle regulators concomitant with exhaustion of HSCs in the bone marrow niche. However, if an inhibitor of the Hh pathway is administered in vivo, the transcriptional and functional defects in HSCs are rescued in the bone marrow-derived stem cell population.

September 12, 2006---In the September 8th online edition of PNAS, Duke University's S. Oltean et al. reported that alternative gene splicing of the fibroblast growth factor receptor 2 (FGFR2) and expression of its isoform, either IIIb or IIIc, impacts prostate tumor progression in rats. With cells derived from Dunning rat prostate tumors, the epithelial cells expressed the IIIb receptor in contrast to mesenchymal cells which express the IIIc isoform. The investigators developed a RFP reporter gene assay to monitor the transcripts of alternative spliced gene variants. Surprisingly, the researchers found that tumors were highly plastic as the cells transition from a mesenchymal to epithelial phenotype (MET state). It was also noted that the cells actively involved in lung metastases appeared to have undergone this transition to a more epithelium-like state. Histological examination of the lung tissues revealed MET clusters in the lung parenchyma, suggesting that microenvironments such as the stroma and endolthelium induced the transition state. (This elegant study supports the vast amount of data of a cancer stem cell. It is conceivable that gene splicing and silencing is occurring in a small population of dysfunctional stem cells that has migrated and formed clusters in highly vascular tissues such as the lungs.)

September 8, 2006---In a second paper (of three research articles) published in the September 6th online edition of Nature, Univ. of Michigan's Sean J. Morrison and his collaborators also demonstrated that an increased expression of p16^INK4a was linked to senescence in aging stem cells. With neural stem cells from the subventricular zone (SVZ) and olfactory bulb, the investigators found a physiological decline in neural progenitor activity. There was less progenitor proliferation in the SVZ as well as less neurogenesis in the olfactory bulb. The researchers also noted that multipotent progenitor frequency and self-renewal potential decline with age in the mouse forebrain. Aging mice deficient in p16^INK4a showed less of a decline in either progenitor proliferation in the SVZ or neurogenesis in the olfactory bulb. The scientists did note that there appeared to be regional differences in progenitor function, since p16^INK4a expression had less of an effect in the dentate gyrus or enteric nervous system in p16^INK4a deficient mice.

September 7, 2006---In the September 6th online publication of Nature, researchers from the University of North Carolina reported their findings on the cyclin-dependent kinase inhibitor p16^INK4a and its impact on the aging process in adult stem cells. With age, the authors noted that stem cells are not as effective as younger cells in repairing wounds, homing to damaged tissues site, and maintaining the body's tissues. The researchers found that the tumor suppressor gene p16^INK4a is highly expressed in older stem cells which is a protective mechanism for preventing the formation of cancer stem cells. The investigators studied pancreatic islet cells from strains of mice in which the p16^INK4a gene were either deleted or knocked-out. It was observed that islet proliferation persisted in these mice. However, if the mice were genetically altered to overexpress the protein to levels observed in cells from older animals, the islet cells aged prematurely. The authors concluded that if one could attenuate the expression of this tumor suppressor gene, it may be possible to enhance islet regeneration and provide a new treatment regimen for treating diabetic patients.

September 5, 2006---In the August 31st online edition of Stem Cells, John Hopkins' University scientists K. Chaichana et al. published an article noting that the dogma of "no new neuron" is being challenged by the recent discoveries of neural stem cells in the adult brain. These multipotent stem cells are found in the subventricular zone of the lateral ventricles and the dentate gyrus of the hippocampus in adults. The authors also noted that neural stem cells are the source of brain tumors. However, a key component for studying the plasticity of these neural stem cells required neurosphere assays in both normal and cancer brain tissues. The investigators noted that there is no standardized protocol for these neurosphere assays since different investigators use "different growth factors and hormones at different concentrations" in studying neural stem cells with their assays. In the paper, the researchers reviewed the literature for both normal and tumor-derived stem cells studies and compiled the different variables used by other investigators in their neurosphere assays. The wide range of assay methods supported their position on the need to standardize the protocol.

September 1, 2006---In the August 30th online edition of PNAS, scientists from Kyoto University published results on the successful transplantation of spermatogonial cells from rats into immature testis of nude mice. The busulfan-treated mice were used as a xenogeneic surrogate. Rat germ cells were implanted in the semineferous tubules of the mice. With spermatids and spermatozoa from the xenogeneic mice, rat offspring were born following in vitro micro-insemination with rat oocytes. The researchers found that the rat offspring were fertile and had a normal imprinting pattern. The authors noted that "the xenogeneic offspring production by interspecies germ cell transplantation and in vitro micro-insemination will become a powerful tool in animal transgenesis and species conservation."

August 31, 2006---Scientists from the University of Edinburgh's Institute of Stem Cell Research published their findings in the August 25th online issue of Development on the signaling pathways leading to the formation of pancreas and liver during embryonic development of the African clawed frog (Xenopus). The researchers delineated the various molecular pathways during formation of the anterior endoderm. Formation of the anteroposterior axis primarily involves both the canonical Wnt and Nodal-related signaling pathways. The investigators also found that during early development of the anterior endoderm, the homeodomain protein, Hex, acts as a transcription repressor of Tle4 expression, while amplifying Wnt signalinng. It was found that Hex also regulates the expression of Nodal-related genes, Xnr1 and XnrW, but not Xnr5 and Xnr6. It was also demonstrated in mouse embryos that Hex is an important transcriptional factor for embryonic patterning in mammals. The authors concluded that Hex is an important protein for embryonic development since its regulatory function is evolutionarily conserved by acting on the same targets for both amphibians and mammals; particularly, as it amplifies Wnt signaling. The authors also noted that understanding these molecular pathways may lead to making both liver and pancereas from embryonic stem cells.

August 29, 2006---Within the last couple of years, cancer biologists have been focusing on the concept of cancer stem cells as the critical cells implicated in drug resistance and tumor progression. These cells are found in very low frequency in both solid tumors and leukemias. Harvard University's scientists, D. Dingli and F. Michor, published an article in the August 24th online edition of Stem Cells proposing that any successful anti-cancer therapies must also focus on eradicating these cancer stem cells. Similar to normal stem cells maintaining tissue homeostasis, cancer stem cells are important for maintaining tumor growth. The authors noted that standard therapies do not target these cancer stem cells, which they surmise might be the reason for the treatment failure and tumor recurrence in many patients. The authors developed a simple mathematical model for demonstrating the importance of designing new successful therapies that can target and eradicate cancer stem cells in tumors.

August 28, 2006---In the August 25th online issue of Cell, P. Laslo and colleagues from the Univ. of Chicago published the results of their study involving a mathematical model for predicting the genetic circuitry/network that orchestrates the development of immature adult stem cells to differentiate into specialized cell types. The investigators used hemapoietic stem cells which were myeloid progenitor cells as the system for mapping the genetic circuitry that would determine how cells became committed to either macrophages or neutrophils. With hematopoietic stems from PU.1^-/- mice, the researchers studied the effect of how the transcription factors PU.1 and C/EBPα at various concentrations in culture media would trigger the developmental pathway toward the cells becoming either a macrophage or a neutrophils. By increasing the concentration of PU.1, a definitive pattern of gene expression resulted in macrophage differentiation. The investigators also found that the transcription factor modulated "a novel regulatory circuit comprised of counter antagonistic repressors Egr-1/Nab-2 and Gfi-1." These repressors were discovered to function in a redundant process that activated macrophage genes while repressing the genes that drive myeloid progenitors into neutrophils. The authors noted that by understanding the gene circuitry that forges a stem cell fate, they believe their results may provide greater insight into understanding how leukemias and other forms of cancers may arise from these stem cells.

August 25, 2006---Most stem cell biologists accept the principle that the microenvironment has an important role in determining a stem cell's fate. It now appears that both the extracellular and cellular environments have important regulatory function in determining how stem cells become committed to various tissue lineages. For example, there have been reports that solid matrices and nano-pits (see April 18th post), extracellular matrices (ECMs) in the basement membrane of kidneys (see April 26th post), and amniotic matrices for embryonic stem cells (see June 12th post) can regulate the differentiation of stem cells. Conversely, destruction of the ECM and the release of matrix metalloproteinases (MMPs) as a result of tissue injury or disease states can activate and in certain cases have a deleterious effect on stem cells surrounding the injury or disease sites of a particular tissue (see April 17th and April 6th posts). Additionally, A. J. Engler et al. from the Univ. of Pennsylvania reported in today's online edition of Cell a study on elasticity of the extracellular matrix in inducing naive mesenchymal stem cells (MSCs) down various lineages of the mesoderm. The investigators reported that soft matrices induce the MSCs down a neurogenic lineage, stiffer matrices induce the cells down a muscle lineage, and rigid matrices caused the stems to turn into osteogenic phenotypes. The researchers also reported that within the first week of culturing the MSCs, they were able to reprogram the cell to different lineages by the introduction of induction factors into the matrix. However, after several weeks in culture, the cells did not respond to the inductive factors and they were committed to differentiate down a lineage specified by the rigidity of the matrix (It is unclear whether it is really a process of "reprogramming of the cells by the introduction of inductive factors or activation and proliferation of subpopulation of uncommitted cells in the cultures).

August 24, 2006---Following on the theme of yesterday's post on spinal cord injury studies, Y. Ziv et al. published in the August 22nd online edition of PNAS results of their study on the interplay between cells of the immune system and adult neural progenitor cells. The study involved T cells from mice vaccinated with a myelin-derived peptide. With a combination of both T cells and adult neural stem/progenitor cells (aNPCs), the cells were injected into the cerebrospinal fluid of mice with a spinal cord injury. The results of the study confirmed a synergistic effect between the antigen-specific T cells and the microglial cells in modulating neural regeneration at the tissue injury site. The researchers noted that the "nature and intensity of the local T cell and microglial response, expression of brain-derived neurotrophic factor and noggin protein, and appearance of newly formed neurons from the endogenous stem cell pool" reflected the synergistic effect of the implanted cells. The authors concluded that the local immune response plays a major role during the wound-healing response in the recruitment of aNPCs to the lesion site.

August 23, 2006---J. Silver et al. from Case Western, Drexel, and University of Arkansas published a study in the July 16th issue of J. of Neuroscience on restoring mobility in paralyzed rats following spinal cord injury. The research was based on a rat spinal cord injury model, in which the spinal cords were severed at cervical level 3 (C3), a novel approach of creating a peripheral nerve bridge, in which they implanted autologous sciatic nerves into the paralyzed rats. Two and half weeks after the peripheral nerve graft, a small pump was implanted into the spinal column at C5 to deliver a steady dose of chrondoitinase-ABC (ChABC). The study was designed to determine whether the enzyme can induce regeneration and extension of the axons through the scar tissues formed after the wound. ChABC is a known inhibitor of chrondroitin sulfate proteoglycans which prevents the local accumulation of the polymer during the wound-healing response. Also, chrondroitin sulfate proteoglycans act as a barrier and prevent axon extension through a scar following an injury. A nerve bridge was created in which the peripheral nerve engrafted at C3 re-grew through the scar tissues and connected to other axons located at C5. The investigators also "primed the newly re-grown axons for rapid regeneration by clipping their ends." The rats treated with ChABC showed restoration of limb mobility in contrast to the control group (treated with saline) of rats where mobility and the use of their limbs did not improve following peripheral nerve engraftment. (Since stem cells play an important role in the wound-healing response, it would have been of interest to determine their role in spinal nerve regeneration in the experiments reported by the authors.)

August 22, 2006---Swedish scientists from the Karolinska Institute in Stockholm, reported in the August 17th online edition of Stem cells the physiological role of nitric oxide (NO) in determining neural stem cell fate by driving the stem cells toward the astroglial lineage. Since nitric oxide is known to have various important physiological roles such as in regulating vasoconstriction, neurotransmission, and as a powerful mediator of the inflammatory cascade, the authors focused their research on elucidating nitric oxide's effects on regenerating tissues, particularly, in relation to the central nervous system of rats. The scientists found neurogenesis was down-regulated following exposure to pathological levels of NO. Down-regulation of pro-neural genes correlated with decreased expression of Notch-1, neurogenin-2, and βIII-tubulin. Similarly, up-regulation of the JAK/STAT-1 signal transduction pathway upon exposure to NO favored differentiation of rat neural stem cells toward the astroglial lineage. The results from their "delayed-differentiation" experiment suggested that the down-regulatory effects on neurogenesis by NO was irreversible.

August 21, 2006---In the August 15th issue of Cancer Research, S. Bao et al. from Duke University Medical Center published their discovery of stem cell-like glioma cells (SCLGCs) isolated from human brain tumors. These SCLGCs were found to generate tumors from xenografts in SCID mice. The authors also noted that these unique cancer stem cells supported tumor growth by secreting vascular endothelial factor (VEGF), resulting in tumor angiogenesis. Similar to neural stem cells, the investigators found that the SCLGCs express the surface marker CD133 (prominin-1) as well as the ability to form neurospheres, to self-renew, and to differentiate into multiple neural lineages. On the other hand, non-SCLGCs did not secrete elevated levels of VEGF and in xenografts formed very few secondary tumors following implantation into the brains of immuno-compromised mice. The researchers were able to retard tumor growth by using an anti-VEGF monoclonal antibody, bevacizumab, as an inhibitor of angiogenesis. The authors concluded that their studies suggested that a subpopulation of cancer stem cells can contribute to tumor malignancy and are a potential key target for future anti-angiogenic therapies. (The investigators used antibody-conjugated Miltenyi magnetic beads of CD133 to purify a population of cancer stem cells. Similar to previous papers that were reviewed in our posts, the experiments did not demonstrate whether they had a homogenous population of cells which led to their results.)

August 18, 2006---In the August 16th online issue of Development, French scientist, J. Y. Bertrand et al., from the Pasteur Institute reported on the role of fetal spleen stroma in forming myeloid precursor cells. In the fetal spleen, the authors showed that hematopoietic stem cells do not proliferate, but instead differentiate into mature macrophages. They also noted that the B cells in the spleen were probably from the fetal liver and migrated to the fetal spleen. The investigators were able to develop a fetal stromal spleen cell line which secreted high levels of anti-inflammatory cytokines and drove hematopoietic stem cells into mature macrophages. The authors proposed that the F4/800+ spleen macrophages were involved in fetal erythropoiesis and the formation of the splenic architecture during development.

August 17, 2006---N.M. Walton et al. from the University of Florida published in the August 16th online edition of Development results on the isolation and large scale expansion of multipotent astroglial neural progenitors from adult human brain. These adult human neural progenitors (AHNPs) were derived from the forebrain regions (the hippocampus and the subventricular zone). The AHNPs were found to be nestin positive and were grown as an adherent cell population in long-term cultures. In these cell cultures, AHNPs were able to co-express glial and immature neural markers following neurosphere formation (i.e. β-III-tubulin and GFAB). With their expansion culture media containing EGF and bFGF, the investigators were able to expand the cells to over 300 population doublings. From long-term cultures, the cells were found to have a normal karyotype as well as being telomerase positive. The investigators deduced from their long-term cultures that a single progenitor could theoretically form approximately 4x10⁷ cells. Electrophysiochemical patch clamp analysis demonstrated that the differentiated cells reflected a neural phenotype. (Although the study results demonstrated the ability to culture an adherent cell population over the long term, the investigators did not publish data characterizing the stem cells that led to their observations. It is also unclear whether the authors, with their expansion media, can generate these AHNPs as a clonal population from a single cell derived from adult forebrains. It was particularly interesting that the AHNPs were implanted into the brains of mice and histological analysis demonstrated the engrafted cells were able to differentiate into various neuronal lineages.)

August 16, 2006---In the August 15th issue of Genes & Development, I. Kyrmizi et al. published their findings on the main hepatic regulators during liver development and organ regeneration. Their analysis revealed both a "temporal and spatial expression pattern" that increased in complexity of autoregulatory and cross-regulatory circuits during liver development. Within these regulatory circuits, the authors identified a core group of six transcription factors which controlled their own expression levels as well as that of downstream hepatic regulators. The investigators also identified certain factors, which exhibited both promoter and developmental stage-specific regulatory functions, as important features of the network. For example, it was discovered that inactivation of the regulatory factor, HNF-4α, was critical for embryonic development, but not in adult liver where inactivation of the factor resulted in the diminished expression of most hepatic factors. The authors concluded that "the results illustrate the remarkable flexibility of a self-sustaining transcription factor network, built up by complex dominant and redundant regulatory motifs in developing hepatocytes."

August 15, 2006---In the August 3rd online issue of Stem Cells, Japanese scientists from Keio University School of Medicine reported the isolation of multipotent stem cells from mouse cornea. The researchers published their findings of isolating, culturing, and characterizing neural crest-derived cornea precursors (COPs). The COPs expressed the stem cell markers, nestin, Notch1, Musashi-1, ABCG2, and a side-population cell phenotype (using Hoeschst 33342 staining and flow cytometry). Neural cells were shown to express βIII-tubulin, GFAP and neurofilament-M. The stem cells were also Sca-1⁺, CD34⁺, CD45^-, and c-kit^-. The COPs were shown to be multipotent, exhibiting the ability to differentiate into adipocytes, chrondrocytes and neural cells, and were found to be of neural crest origin, expressing the embryonic markers Twist, Snail, Slug, and Sox9. The authors concluded that "taken together, these data indicate the existence of neural crest-derived, multipotent stem cells in the adult cornea."

August 14, 2006---In the August 10th online issue of Stem Cells, French scientists published findings of in vitro differentiation of bone marrow-derived mesenchymal stem cells into functional neuronal cells. The investigators found that these mouse mesenchymal stem cells expressed nestin, a marker for neural progenitor cells. Upon the introduction of neural induction agents into the culture media, the mesenchymal stem cells morphed into distinct neuronal shape and expressed the neuronal markers NF-L and class III beta-tubulin. Additionally, the scientists demonstrated that cytosolic calcium concentration increased following stimulation by specific neuronal activators. The authors concluded that a clonal population of multipotent stem cells were found in the bone marrow that could be differentiated into neuronal lineages. (It should be noted that these types of studies have been reported by others in papers published over 5 years ago. For example, Romero-Ramos et al. from UCLA published their study in the Journal of Neuroscience Research in 2002 of muscle-derived rat stem cells that were Thy-1/CD90 positive and converted them into neurons. Click here to download the paper)

August 11, 2006---In yesterday's online issue of Cell, Japanese scientists published their study on in vitro induction of mouse fibroblast cultures into pluripotent stem cells. They used both adult and mouse embryonic stem cells in their induction process and obtained similar results. These induced pluripotent stem cells (iPS) reflected the morphology and markers of embryonic stem cells. When the investigators implanted the iPS into a blastocyst they were found to participate in mouse embryonic development. The inductive agents to obtain the results in the study were Oct3/4, Sox2, c-myc, and K1f4 under embryonic culture conditions. It was further noted that the Nanog transcription factor was not needed for generating these iPS. The authors concluded pluripotent stem cells can be generated directly from fibroblast cultures with the addition of a few defined factors. (It should be noted that the iPS from both mouse embryonic fibroblasts and the tail tip of adult mice were derived using embryonic cells and transgenic animals transduced with the bgeo [fusion β-galactosidase and neomycin resistant genes] in order to select G418 antibiotic resistant colonies in vitro. Transfection was conducted with a retroviral vector near the Fbx15 locus in order to select out pluripotent cells with cultures containing high concentration of the antibiotic. The investigators also pointed out that the iPS from adult transgenic tissues were able to form tumors in SCID mice. There is also the question whether the use of a retroviral vector in the study may have transformed the iPS to cancer causing stem cells. Additionally, it is not clear whether the transcription factors affected a subpopulation of stem cells since the iPS were found in a very low frequency in the cell cultures.)

August 10, 2006---Along the lines of our recent posts regarding cancer stem cells, J.E. Ippolito et al. from Washington University School of Medicine published in the August 8th online issue of PNAS a comprehensive study of the linkage between cellular communications, energy utilization, and profile ration of metastatic neuroendocrine cancers. Using a combination of magic angle (54.7 degrees) spinning nuclear magnetic resonance spectroscopy (MAS-NMR) and gene chip microarrays, the authors investigated the metabolic features that support metastasis in human neuroendocrine (NE) cancers and prostate NE cancer (PNEC) cell lines derived from transgenic mice. The investigators reported that increased proliferation of malignant NE cells was correlated with stimulation of the GABA, glycine, and glutamate receptors. The subsequent intracellular substrates were metabolized and various metabolic pathways were assayed with various inhibitors. It was very interesting that the researchers were able to accurately predict in silico biotransformations of the metabolome in these neoplastic cells by detecting the gene transcripts with GeneChip profiling. The paper demonstrated the importance of cell-cell communication via neurotransmitter and subsequent receptor-ligand interactions which leads to metabolism of specific substrates and which "support the aggressive behavior of human NE cancers." (It would be interesting to study freshly isolated primordial adult stem cells with the authors' assay systems and determine how well their in silico predictions correlate to the data obtained from their studies.)

August 9, 2006---Scientists from University of Illinois published in the August 7th online issue of Developmental Cell the results of a study delineating the mechanism involved in tissue regeneration of freshwater planarians. The flatworms have an extraordinary ability to regenerate their tissues, particularly after they have been cut up into several pieces. Similar to salamanders, the ability to regenerate their tissues is attributed to stem cells (neoblasts) that form a blastema at the wound site. These neoblasts are maintained throughout the life of planariums. The investigators discovered that the Bruno-like family of RNA binding proteins (Bruli) is critical for regulating neoblasts during formation of the blastema and differentiation into native tissues. With RNAi, the researchers found that Bruli knockdowns resulted in depletion and elimination of neoblasts. They reported that neoblasts lacking Bruli were still able to respond to the wound stimuli and generate progenies that could form the blastema and regenerate the damaged tissues. However, since the neoblasts lacking Bruli were not able to self-renew, the authors concluded that Bruli was critical for self-renewal and maintaining the stem cell niche in planarians.

August 8, 2006---In an August 4th press release, researchers from UCLA's Institute for Stem Cell Biology reported that neural stem cells produced from federally approved embryonic stem cell lines were inferior to neural stem cells derived from human fetal tissues. The researchers noted that the metabolic marker, CPT 1A, was under-expressed in the neural progenitors from embryonic stem cell lines. Camitine palmitoyl transferase type-1A deficiency (CPT 1A) can lead to the inability to break down fats (fatty acid oxidation) and which eventually leads to hypoglycemia. The investigators also noted that neural stem cells from fetal tissue did not demonstrate any abnormal DNA modifications with respect to cytosine methylation pattern when compared with cancer cells. The "gene silencing" patterns from fetal neural progenitors were found to be normal using microarray analysis. The researchers intend to use other established human embryonic stem cell lines to determine if their findings were an anomaly with respect to neural progenitor cells derived from other established and federally approved embryonic cell lines. The study was published in the recent online issue of Human Molecular Genetics. (It also difficult to analyze a population of cells from long-term cultures, especially when they are compared to cells freshly isolated from human tissues. Cell lines are notorious for producing artifacts. This was particularly noted in our May 16th post in which Gerald Dermer, Ph.D. made the case against current cancer research being conducted on transformed cell lines in his book "The Immortal Cell: Why Cancer Research Fails.")

August 7, 2006---In the August 3rd online issue of Stem Cells, S. Chiba from the University of Tokyo Hospital published a review article on the Notch signaling pathway. The author noted that the Notch gene is well conserved throughout evolution from nematodes to humans. In humans, the Notch homologue was first identified in a subset of T lymphoblastic leukemias and denoted as a TAN-1 (translocation-associated Notch homologue 1 or Notch 1) gene. It was noted that the Notch signaling pathway was critical for guiding neural progenitors to differentiate into glial cells. Notch is also involved in regulating apoptosis in mammals, but the author noted that it is not clear whether it is pro- or anti-apoptotic in developing neural stem cells. Notch also has an important role in organogenesis including the kidney, liver, pancreas, and heart. The author emphasized that Notch signaling is involved in cell fate determination during embryogenesis by: 1) affecting differentiation from primordial cells to tissue-specific stem cells during early- to mid-stage embryo; 2) inhibiting tissue- or organ-specific stem cells or progenitors from further differentiation; and, 3) blocking the "default pathway" and promoting the "alternative pathway" during organ formation in mid- to late-stage embryo development. In adult stem cells, Notch has a very important role for maintaining the hemapoietic stem cell niche in the bone marrow as well guiding differentiation of the progenitors down various tissue lineages. In summary, the author concluded that "ex vivo stem cell expansion is fundamental to the success of stem cell-based regenerative medicine and that Notch signaling most likely has a role in the expansion of the stem cells." It was also noted by the author that since Notch signaling on progenitor cell survival has been demonstrated, the tumorigenic aspects must be considered.

August 4, 2006---As a follow-up to our July 20th post on multipotent stem cells and their relationship to a side population of cells in adult tissues, K. Iohara et al. from Kyushu University and the National Institute for Longevity Sciences in Japan reported in the July 27 online issue of Stem Cells the isolation of a side population (SP) multipotent stem cells from porcine dental pulp tissue. With flow cytometry and exclusion of the DNA binding dye, Hoechst 33342, the investigators found a SP of stem cells that enriched the dental pulp. These SP cells expressed the Polycomb group transcription repressor, Bmi1, which suggested their ability to self-renew without senescence. The researchers were able to convert 30% of the SP to the chondrogenic lineage in which they expressed type II collagen and aggrecan. Adipogenic conversion of SP cells was confirmed by in vitro induction and expression of PPARγ and αP2. The scientists also found that they were able to convert 90% of the SP to the neural lineages in which the cells expressed neurofilament and neuromodulin. With the addition of bone morphogenetic protein 2 (BMP2) into 3-dimensional pellet cultures, the SP cells expressed dentin sialophosphoprotien (Dspp) and enamelysin. The authors concluded that SP cells from dental pulp could be a useful cell therapy approach for dentin regeneration.

August 3, 2006---In the August 1st online issue of PNAS, M. Knockaert et al. from Rockefeller University reported results involving the signaling pathways for bone morphogenetic proteins (BMPs) during embryonic development. With Xenopus embryos as a model for their study, the investigators examined the Smad transcription factor and how it is activated and recycled between the nucleus and cytoplasm of a cell. Smad transcription factors are the signal transducers for TGF-β and the BMP family of cytokines and morphogens. Ligand binding with these cytokine receptors results in C-terminal serine phosphorylation of Smad, which in turn drives the transcription factor into the nucleus. Dephosphorylation of the Smad in the nucleus allows the transcription factor to recycle back to the cytoplasm which the authors describe as allowing "agonist sensing by the cell." The researchers identified SCP (small C-terminal domain phosphatase) as the critical mediators of Smad activity. In Xenopus, inhibition of BMP signaling by SCP2/Os4 results in neuralization of the embryo. With human cells, the authors noted that RNAi-mediated depletion of SCP1 and SCP2 enhances Smad1 activity and the duration of the BMP responsive genes which eventually leads to the induction of the secondary dorsal axis of the embryo. (In our May 31st post, we summarize the role of another member of the TGF-β superfamily, Lefty, as one of the determinants for whether a stem cell maintains its "stemness" state or exit it by differentiating into progenitor/precursor cells. It was also noted that Smad2/3 regulated Lefty while activated Smads maintained pluripotency in a stem cell. Now through Knockaert et al., we find modulation of Smad activity in the nucleus is through the presence of SCP. In turn, Lefty1 and Lefty2 blocks Nodal signaling...a morphogen described by the authors in yesterday's post with respect to their presence in melanomas.)

August 2, 2006---In continuing our recent theme on the relationship between cancer and stem cells, J.M. Topczewka et al. from Northwestern University Feinberg School of Medicine and Loyola University Medical Center reported in the July 30th online issue of Nature Medicine the discovery of an embryonic morphogen, Nodal, produced by human melanoma cells. The researchers used developing zebrafish as a model and biosensor for tumor-derived signaling pathways in humans and their involvement in developmental pathways of embryonic zebrafish. They found that Nodal contributed to the ectopic formation of the embryonic axis. Additionally, Nodal is present in 60% of the human metastatic tumors, but not in normal skin. It was particularly interesting to note that inhibition of the Nodal signaling in melanoma cells resulted in a reduction of invasiveness, colony formation and tumorigenicity in these malignant cells. Further, the researchers reported that Nodal inhibition promoted reversion of the melanoma cells toward a melanocyte phenotype. The authors concluded that their data suggest that Nodal provides a "new molecular target for regulating tumor progression," particularly, in melanomas. (The only question one would raise is more of semantics, but of critical importance. By inhibiting Nodal signaling in the melanomas, is it really a reversion of the neoplastic cells to melanocytes? That would be the case if one assumes that melanomas arose through a process of "dedifferentiation" of melanocytes. However, there is a growing body of scientific data supporting the concept that many cancers such as melanomas, leukemias, and ovarian and breast cancers are the result of dysfunctional or "deranged" cancer stem cells that are blocked, through genomic perturbation, from maturing into terminally differentiated somatic cells. Thus, one may also interpret the data that Nodal inhibition unblocked or allowed the cancer stem cells to continue on their path towards differentiating into a melanocyte.)

August 1, 2006---As a follow-up to our July 28th post, in the same August 8th issue of Stem Cells, L. M. Reid and her colleagues from the University of North Carolina published results on the characterization of pluripotent hepatic progenitors. The investigators defined hepatic stem cells as precursors to hepatoblasts that remain in the liver throughout the life of an organism. The gene expression profiling of the hepatic stem cells demonstrated that the cells expressed high levels of cytokeratin-19 (CK19), neuronal cell adhesion molecule (NCAM), epithelial cell adhesion molecule (EpCAM), and claudin-3 (CLDN-3). It was also noted that these cells expressed low levels of albumin and they did not express alpha-fetoprotein (AFP) and adult liver-specific proteins. In contrast, the hepatoblasts, the predominant cell in fetal and neonatal livers, declined in their numbers with age and in normal adult livers they represented less than 0.1% of the total cell population. The hepatoblasts also expressed high levels of AFP concomitant with low levels of CK19 and a loss in gene expression of both NCAM, and CLDN-13. The authors further noted that mature hepatocytes lose the ability to express the hepatic stem cell and progenitor markers while expressing high levels of albumin, cytochrome P4503A4, connexins, phosphophenopyruvate carboxykinase and transferrin. (In the past, Reid et al. had published their results on the ability of these hepatic stem cells to differentiate in vitro into cells of tissues from ectodermal and mesodermal origins.)

July 31, 2006---Investigators from M.D. Anderson Cancer Center in Texas and their collaborator from France published a study in the July 28th issue of PNAS on the Wilms tumor suppressor gene, Wt1, modulating Sox9 expression in the maintenance of tubular architecture in the developing testis. The authors noted that mutation of the transcription factor and Wt1 result in genitourinary anomalies and gonadal dysgenesis. However, the role of Wt1 for testis development after the initial steps of sex determination during embryonic development is not well defined. In a mouse strain carrying the conditional knockout allele, the investigators reported disruption of developing seminiferous tubules and subsequent loss ot Sertoli and germ cells, which is reflected in the mutant testes by severe hypoplasia. It is further noted that in the conditional mutants, the testis-determining gene, Sox9 was turned off on embryonic day 14.5 following ablation of Wt1 function. (This is another example where genes associated with cancers are also found to be intimately involved in various pathways of differentiation and embryonic development.)

July 28, 2006---The liver is one of the few organs in adults that is able to regenerate itself following injury or a hepatectomy. Accordingly, there has been extensive research into the cell populations that may give rise to the regenerative capabilities of the liver. In the August 8th issue of Stem Cells , M.A. Walkup and D.A. Gerber from the University of North Carolina School of Medicine provide a short, but comprehensive overview of hepatic stem cell biology in their article entitled, "Hepatic Stem Cells: In Search of." The authors noted that one area of extensive study is the identification of hepatic stem and progenitor cell subpopulations. These hepatic progenitors were described as "either bipotent or multipotent with capability of undergoing multiple rounds of replication." It was noted that in adult murine livers a population of oval cells express the hepatic markers albumin, alpha-fetoprotein (AFP), and cytokeratin 19 (CK-19, a bilary epithelial cell marker). These small cells are found in the terminal bile ducts with a high nuclear to cytoplasm morphology. In a liver injury model, these cells appear to be activated, which suggests that oval cells are at least bipotent hepatic progenitors. (It was particularly interesting to learn that the antigenic markers for hepatocarcinoma are also expressed on oval cells...OV-6, OC-2, and OC-3. This supports our thesis that many cancer markers are really stem markers that are not downregulated due to the stem cell's inability to terminally differentiate. Further, the authors noted that activation of the oval cell compartment occurs prior to hepatocellular carcinoma development. It was stated that "one of the links between hepatocellular carcinoma and hepatic progenitor cells is the ductular reaction that occurs with chronic hepatitis.) The authors concluded in order to use hepatic progenitors in a clinical setting, there must be a strategy for expanding these cells to a "critical mass" while controlling/maintaining their regenerative capacity.

July 27, 2006---In the July 20th issue of Neuron, A. Alvarez-Buylla and colleagues from UCSF and Spain published their findings on a subpopulation of neural progenitor stem cells (B cells) that are able to form gliomas. These B cells were found in the subventricular zone (SVZ) of adult brains which along with the dentate gyrus are areas enriched with neural stem cells that are able to produce neurons and oligodendrocytes. Of particular note was the identification of PDGFRα (platelet-derived growth factor receptor-alpha) expressed on the surfaces of these B cells. These particular B cells expressed the glial cell protein, GFAP, as well as exhibiting morphological features of astrocytes. The authors also noted that the requirement of PDGF and the expression of its receptor in B cells for oligodendrogenesis but not neurogenesis were demonstrated in conditional ablation of the PDGFRα on B cells. Infusion of PDGF resulted in "large hyperplasias with some features of gliomas." The investigators hypothesized that PDGFRα signaling may regulate the balance between oligodendrocyte and neuron production. They further proposed that excessive activation by PDGF in the SVZ may result in tumor formation.

July 26, 2006---In the July 20th online issue of PNAS, investigators from Duke University reported that an enzyme, aldehyde dehydrogenase (ALDH), was a key regulator in the differentiation of hemapoietic stem cells (HSCs). It was noted that in the liver the ALDH enzyme metabolizes vitamin A to retinoic acid. Through receptor-ligand interactions, retinoic acids induce differentiation, tissue patterning, and embryonic development. Additionally, all-trans-retinoic acid (ATRA) has been used therapeutically as an inductive agent to treat patients with promyelocytic leukemia. The authors noted that ALDH is enriched within HSCs and is a marker for hematopoietic progenitors. An inhibitor of ALDH is diethylaminobenzaldehyde (DEAB). When DEAB is added to short-term cultures of HSCs, the cells have significantly reduced metabolic activity in ALDH along with a four-fold increase in the expansion of primitive HSC (CD34⁺CD38^-lin^-) progenitors. The researchers also demonstrated that a higher percentage of these long-term re-populating cells were able to engraft the bone marrow of NOD/SCID mice. The scientists further noted that inhibition of ALDH by DEAB also decreased retinoic acid activity and up-regulated HOXB4 expression in HSCs. HOXB4 encodes for a homeo box protein that targets self-renewal concomitant with in vitro expansion of HSCs. The author concluded that inhibition of ALDH and retinoic acid activity can induce in vitro expansion of human HSCs, as well as expanding a subpopulation of long-term re-populating progenitor cells.

July 25, 2006---Taiwanese scientists published in the July 20th online issue of Stem Cells results on the in vitro differentiation of multi-potent cells from human placenta into hepatocytes. These placenta-derived multi-potent cells (PDMCs) were cultivated in a hepatic differentiation culture medium. During cultivation in the specialized medium, the cells differentiated and exhibited morphologies of "polygonal epithelial-like cells." The authors reported that the differentiated cells also expressed albumin and cytokeratin 18, markers that are specific to hepatocytes. Typical of liver cells, the investigators demonstrated that the differentiated cells could uptake lipoprotein and store glycogen. When the drug rifampicin was added to the culture media, gene expression of CYP3A4 was upregulated, a response similar to that observed in human live cells. From the results, the authors concluded that PDMC is another source of stem cells that could be used to generate tissue lineages of all three germ layers, similar to embryonic stem cells, and could be used specifically for "progenitor cells that are capable of differentiating toward hepatocyte-like cells in vitro."

July 24, 2006---D-Q. Tang et al. from the University of Florida reported in the August 2006 issue of J. Laboratory Investigation the role of Pax4 in "transdifferentiation" of rat liver cell lines which were transfected with a Pdx1-VP16 gene. Pdx1 is a master transcription factor regulator in the development of the pancreas. Pdx1-VP16 is a fusion protein consisting of Pdx1 and the C-terminal activation doman (80 amino acids) of the Herpes simplex viral protein VP-16. The fusion protein confers a hyperactive form of Pdx1. In the cell line (WB-1) expressing Pax4, transfection of the Pdx1-VP16 gene resulted in expression of late pancreatic β-cell differentiation transcription factors Pax6, Isl-1, and MafA. The gene expression profile of these transduced cells resembled that of an insulinoma. On the other hand, the cell line (WB-2) which does not express Pax4 were found to be incapable of differentiating into β-cell precurors and insulin producing cells. In in vivo experiments with streptozotocin-induced diabetic mice implanted with the WB-1 tranduced cells, the investigators were able to produce insuling-secreting cells that were capable of inducing normoglycemia in the diabetic mice. (It is not clear whether WB-1 cell line had "transdifferentiated" as a result of the transduction of the Pdx1 gene. Does the cell line represent a population of bipotent stem cells?)

July 21, 2006---Australian scientists from the Walter and Eliza Hall Institute published in the July 19th edition of the J. National Cancer Institute the results of their study for identifying a stem cell population in mouse mammary tissues that was similar to a population of basal cells found in human breast tumors. With flow cytometry and differential expression of CD24 and a6b1-integrin complex, the investigators were able to enrich for a population of stem cells from normal mouse mammary tissue. These cells had the triple negative phenotype for estrogen receptor α (ERα), progesterone receptor (PR), and the erbB2 (Her2), which in humans are prognostic markers for breast cancers. Similar to basal cell breast cancers, these stem cells expressed the epidermal growth factor receptor (EGFR)/ErbB1. The colony-forming progenies derived from these stem cells were identified as CD24⁺ cells. The basal cell progenies had low levels of either CD29 or CD49 expressed on their surface, while other progenies co-expressed markers with luminal features. The normal stem cells were also positive for p63, which is a hallmark of human basal tumors. The authors deduced that these findings in mice support the thesis of a cancer stem cell or tumor-initiating cell (gate-keeper event) evolving from normal stem cells. It was further noted that the cancer stem cell can give rise either to a population of basal cells with the triple negative hormone receptor phenotype or a luminal cell population which is positive for the steroid hormone receptors particularly if the cells were positive for the estrogen receptor.

July 20, 2006---In following up on the July 18th post, the research paper by P.K. Donahoe and her group from the Harvard Medical School was published in the July 18th early online edition of PNAS. In our post, we noted that Harvard scientists had identified by flow cytometry a unique population of somatic stem cells that they described as a side population of cancer stem/progenitor cells. Using differential efflux of the DNA-binding dye, Hoeschst 33342, in rodent ovarian cancer cell lines, a side population (SP) of cells was identified as breast cancer-resistance protein 1-expressing (BCRP1), verapamil-sensitive cells. The SP cells also expressed the typical OVCAR-3 and OVCAR-8 surface markers normally found on the surface of transformed cell lines. In vivo experiments demonstrated that the SP cells formed measurable tumors 7 weeks after injection into the dorsal footpad of nude mice. In contrast, the non-side population cells (NSP) formed tumors 10 weeks post injection. After enriching for SP cells by flow cytometry and growing them in cultures, the investigators found a NSP population of cells. They deduced that the NSPs were formed through asymmetric cell division of the SP cells. The authors also noted that Mullerian Inhibiting Substance (MIS) inhibited the proliferation of both SP and NSP cells, whereas the lipophilic chemotherapeutic agent, doxorubicin, more significantly inhibited the NSP cells. The experimental results led the researchers to conclude that "in the future, individualized therapy must incorporate analysis of the stem cell-like subpopulation of ovarian cancer cells when designing therapeutic strategies for ovarian cancer patients."

July 19, 2006---Yesterday in the U.S. Senate, the senators approved by an overwhelming majority of 63 -37 votes a bill for the federal government to fund human embryonic stem cell research. However, the Senate lacked the two-thirds majority for the measure to become law if president Bush vetos the bill, something which he has vowed to do. In yesterday's Wall Street Journal, it was reported that even if the bill was to pass, another problem facing stem cell researchers is the broad patents held by the Wisconsin Alumni Research Foundation (WARF). University of Wisconsin researcher, James Thomson, who first isolated and propagated stem cells from human embryos in 1998, assigned to WARF his the rights to the inventions. WARF has stated publicly in the past that they were going to aggressively pursue a policy of charging commercial users a licensing fee ranging from $75,000 to $250,000 with subsequent annual payments and royalties. On the other hand, WARF will allow academic laboratories to use human embryonic stem cells to conduct their research without a charge. However, the state of California is being pulled into a potential patent dispute with WARF after the California Institute for Regenerative Medicine (CIRM) recently announced that they proposed to take 25% of the revenues in excess of $500,000 that universities received from patented discoveries funded by Prop. 71. WARF noted last spring that they should participate in CIRM's revenues since they were moving into commercial use of WARF's patents. Joining the fray is a consumer watchdog group, the Foundation for Taxpayer and Consumer Rights of Santa Monica which is asking the U.S. PTO to overturn the 3 patents held by WARF. As part of this appeal process, a declaration by Jeanne Loring, a stem cell researcher at the Burnham Institute with her own embryonic stem cell lines licensed to CyThera, Inc (acquired by Novocell in Irvine), reasoned that the patents shouldn't have been issued to Thomson because earlier discoveries with animal embryonic stem cells made the human discoveries an obvious step.

July 18, 2006---A. Krivstov et al. at the Dana-Farber Cancer Institute and Children's Hospital in Boston reported in the July 16th online issue of Nature the isolation of hematopoietic progenitor cells and conversion of them into leukemic cancer stem cells in mice. Mouse granulocyte-macrophage progenitors (GMPs) were isolated from the bone marrow that were IL-7R^-, Lin^-, Sca-1^-, c-Kit⁺, CD34⁺, and FcgRII/RIII^hi. The investigators transfected the GMPs with a mixed lineage leukemia (MLL) gene that encoded for the MLL-AF9 fusion protein. The transfected cells were subsequently injected into sublethally irradiated syngeneic recipient mice. The researchers noted that within 80 days the animals receiving the transfected GMPs developed oligoclonal acute myelogenous leukemia (AML) and animals receiving at least 500 or more of the leukemic cells died within 55 days. With L-GMPs grown in cytokine supplemented methylcellulose, the cells were found to have higher blast colony-forming activity, which the authors attributed to the presence of leukemia stem cells (LSCs). It was also noted that as little as four cells injected into a secondary recipient could initiate AML. However, if the L-GMPs were cultivated in liquid cultures containing IL-3, the cells began to differentiate (converted to a c-Kit^- immunophenotype) while losing their capacity to induce AML in a recipient. With Affymetrix chip analysis to measure gene expression, the L-GMPs had a gene expression profile that resembled a less committed progenitor, where the genes were expressing a signature (group of HOX genes and Mef2c) that is normally associated with self-renewal. Moreover, L-GMPs grown in liquid cultures lost the self-renewal-associated gene signature. The investigators also found overlapping self-renewal gene expression signatures in LSCs derived from humans with AML. The authors concluded that "these data suggest that ectopic re-activation of genes associated with self-renewal is a central feature of leukemic transformation of progenitor cells."

July 17, 2006---In a Reuters news release on July 11th, P.K. Donahoe et al. from Mass. General and Harvard Medical School reported they had isolated primitive cells from both rodent and human tissues that resemble stem cells. They found these unique cells acted as master cells found in ovarian tumors of mice and various tumors of humans. Like cancer cells, these cells had the capacity for unlimited self-renewal and proliferation. The investigators noted that when these "side-population of cells were injected in mice they formed tumors. They also noted that Mullerian Inhibiting Substance (MIS) slowed the tumor's growth. MIS is naturally secreted by Sertoli cells in the the testes and causes regression of the Mullerian ducts during embryonic development. In previous reports, Donahoe et al. noted that ovarian cancer cells expressed a receptor for MIS. It was speculated that better characterization of these side-population cells could result in new anti-cancer agents as well as MIS itself being used as a possible drug for treating ovarian cancer. (In the press release, it was noted that the paper was published in the most recent issue of PNAS. However, we were not able to find the paper in the either the current issue or early online edition of PNAS. We will in a separate post describe in more detail these unique side-population of cancer stem cells described by the authors. It should also be noted that Clarke and Becker wrote an article in July issue of Scientific American describing these rare and hard to find cancer stem cells in tumors.)

July 14, 2006---In the last few years, stem cell biologists have had a strong interest in understanding the Wnt family of genes and the role its proteins play in various signaling pathways that control proliferation and differentiation of stem cells. The Wnt proteins have also been implicated in tissue homeostasis in adults and Wnt mutations are thought to be involved in various forms of cancers. In this regard, German scientists discovered that genetic aberration in the Wnt/β-catenin signaling pathways can induce chromosomal instability. The researchers noted that mutations in the APC tumor suppressor gene in colon cancer up-regulated Wnt/β-catenin activity. The authors published their results in the July 11th issue of PNAS, noting that activation of Wnt/β-catenin led to transcription of various target genes including the conductin/AXIN2 genes. The scientists discovered that up-regulation and over expression of conductin causes chromosomal instability. In cancer cells, it was demonstrated that conductin localizes along the mitotic spindle during cell division, binding to and compromising the activity of the spindle checkpoint enzyme, polo-like kinase I. In knock-down experiments of conductin with RNA-i in colon carcinoma cells, the checkpoint is enforced and abrogates chromosomal instability.

July 13, 2006---J. Mehta et al. from Northwestern University School of Medicine published results of their clinical study in the July issue of Bone Marrow Transplantation. In the study, they compared younger and older donors of bone marrow-derived hematopoietic stem cells in allogeneic transplants into patients with hematologic malignancies. The study consisted of 63 patients ranging in age from 27-66 years (median 52) who were allografted with donors aged 24-64 years (median). Prior to the allograft, the patients were administered an immunosuppresive regimen of cyclosporine or tacrolimus, and mycophenolate mofetil to prevent graft-versus-host disease (GVHD). In the two year cumulative study, the investigators found the incidences of relapse and treatment related mortality were 55% and 24%, respectively, and the 2-year probabilities of overall survival and disease-free survival were 36% and 21%, respectively. Poor performance status were correlated to donors of age greater than 45 years. The authors stated, "this finding raises the question of whether a young 10-allele-matched unrelated donor is superior to an older matched sibling donor in patients where the clinical situation permits a choice between such donors." (Another question is whether age and cell dosage of a putative and more primordial subpopulation of stem cells are important variables that warrant further investigation. Efficacy may be a function of the number of these cells engrafted into the recipient and may be contributing to the study's results.)

July 12, 2006---As a company focusing on advancing adult stem cell technologies for regenerative medicine, we avoid the political debate concerning the moral and religious issues of embryonic stem cell research. However, Michael Kinsley as the Op-ed columnist for the Washington Post, wrote in his July 7th column that there is an irrational and moral dilemma that proponents of a ban on embryonic stem cell research face while either ignoring or supporting fertility clinics. He notes that if one assumes that embryonic stem cell research is a taking of a human life, than one needs to address the issue of in vitro fertilization where IVF clinics intentionally produce extra embryos as a standard practice for their clients. These extras embryos are then frozen and eventually destroyed. Kinsley argues that "in short, if embryos are human beings with full human rights, fertility clinics are death camps -- with a side order of cold-blooded eugenics. No one who truly believes in the humanity of embryos could possibly think otherwise." Kinsley, a political conservative with Parkinson's disease, supports embryonic stem cell research for his own health issues. However, like many supporters of embryonic stem cell research, they often make misguided statements about the potency of adult stem cells in order to support their position on embryonic stem cells. Kinsley states: "No other potential therapy -- including adult stem cells -- is nearly as promising for my ailment and others. Evaluate that as you wish." Irrespective of one's position on embryonic stem cells and its potential to cure human ailments, we believe one only has to read the below posts on adult stem cells in order for you to draw your own conclusion regarding their potential as cell-based therapies; and, using Kinsley's words, "evaluate that as you wish."

July 11, 2006---Scientists from the Unviersity of Edinburgh reported in this month's issue of Am. J. Physiology-Gastrointestinal and Liver Physiology that they had identified a primitive liver precursor cell from human fetuses. The research group confirmed that similar to rodents these fetal hepatic progenitor cells were Thy-1⁺ cells. Cells derived from the midtrimester of human fetal liver were assayed with a panel of antibodies to Thy-1, cytokeratins 18 and 19, vimentin, CD34, CD45, and fibrinogen. With flourescence-activated cell sorting (FACS), the investigators were able to purify and sort out two subpopulations of primitive liver cells in which one group expressed a Thy-1⁺ marker and another had a Thy-1⁺CD34⁺ phenotype. They noted that the Thy-1⁺ cells also expressed cytokeratin 18 and cytokeratin 19. The scientists also demonstrated by RT-PCR that the "Thy-1⁺CD34⁺ cells expressed mRNA for alpha-fetoprotein, transferrin, and the hepatocyte nuclear factor-4α." They concluded that their "data suggest that Thy-1 is a marker of liver stem cells in human fetal liver." (It should be noted that Young et al. in 1994 had discovered and patented a muscle-derived pluripotent mesenchymal adult stem with the Thy-1 and CD34 phenotype. Additionally, in the below June 28th post, researchers from the Univ. of Washington reported identifying a human fetal multipotent progenitor cell which among other surface markers were also found to be CD90⁺ and CD34⁺.)

July 10, 2006---University of Iowa scientists, S. Bonde and N. Zavazava, published in the June 22, 2006 online issue of Stem Cells their results on the immunogenicity and the ability of mouse embryonic stem cells (ESCs) to be engrafted into allogeneic adult recipients. The investigators reported that the mouse-derived ESCs did not stimulate T cells in vitro. The ability for ESCs to escape the host's immune response appears to be mediated through a process that requires cell-cell interaction. They further demonstrated that the mouse ESCs were resistant to lysis by the host's natural killer cells despite high expression of RAE-1 ligand to NK cell's receptor NKG2D. In sublethally irradiated recipients, the ESCs were able to integrate with the lymphoid tissues. Differentiated donor cells were found in the thymus, spleen, and liver. With immunohistochemical analysis, the scientists established multilineage mixed chimeras. They also believed that inhibition of the host's responses to the engrafted cells occurred through the deletion of alloreactive T cell by inducing T cell apoptosis. (There are reports which also demonstrate that bone marrow-derived and hematopoietic adult stem cells escape the immune surviellance of its host. Other investigators have proposed mechanisms in which primitive stem cells may either serve as tolerogens or possibly that the naive stem cells that migrate to the host's lymphoid tissues, such as the thymus, could differentiate into regulatory T cells while trafficking through those tissues.)

July 7, 2006---In our July 5th post, the study of Fazel et al. was summarized with respect to bone marrow-derived c-kit+ stem cells and their role in neovascularization following a heart attack. In that same issue of the J. Clinical Investigations, K. Chien from Harvard University published his commentary on the work and reviewed some of the relevant clinical studies of using bone marrow-derived cells as a therapeutic approach for regenerating cardiomyocytes. Chien noted that the data from the Canadian research group suggest that bone marrow precursor cells apparently are involved mechanistically in the paracrine angiogenic pathways for neoangiogenesis rather than direct differentiation into either cardiomyocytes or vascular cells. Since very little difference in long-term effects are observed in patients between the control group and those receiving bone marrow-derived cells after a heart attack, Chien questions whether the observations by Fazel and others might be more closely related to the "biology of wound healing rather than regenerative medicine." He speculated that the short-term effects of the infiltrating or transplanted cells may be based more on the "triggering of an inflammatory response with secondary release of cytokines and nonspecific angiogenic cues." In previous experience with bone marrow-derived stem cells, Chien speculated that similarities in the placebo-controlled group and the treated group over the long term may be attributed to inflammatory pathways leading to cardiac scar formation. (We believed Chien is correct in his assumption. Furthermore, cell dosage of pluripotent or multipotent cells and the timing for administration of the stem cells is also critical for observing long-term benefit with stem cell therapy to treat heart attacks.)

July 6, 2006---D.Levy and G. Inghirami from NYU School of Medicine and University of Turin, respectively, wrote a brief commentary on STAT3 as a multifaceted oncogene in the July 5th issue of PNAS. STAT proteins are a family of transcription factors of signal transducers and activators of transcription, thus, the acronym. Activation of the protein occur through a site specific tyrosine phosphorylation which results in a conformational rearragnement and dimerization. The authors noted that STAT proteins were initially identified as intermediaries in the interferon signaling pathway. As additional members of this family were discovered, these proteins were found as important mediators of growth and proliferation in cells. The association between STAT proteins and cancer was made when it was discovered that STAT3 was a substrate for the tyrosine kinase oncoprotein, v-Src. Cancer biologists identified STAT3 as an oncogene when cancer cells were found to have high levels of STAT3. The authors further noted that "a considerable amount of experimental and clinical observations have now confirmed such a role for STAT3, and a remarkable degree of diversity has been uncovered for the molecular mechanisms at the basis of STAT3 action." (However, we are now finding that some oncogenes are also expressed in normal adult stem cells. Are these oncogenes really stem cell genes that are not downregulated in activated stem cells that are blocked in their ability to differentiate?)

July 5, 2006---In a press release, Canadian scientists from the McEwen Centre for Regenerative Medicine at Toronto General Hospital reported the role of an unique subset of bone marrow-derived adult stem cells in repairing heart tissues after a heart attack. They found that c-kit+ stem cells respond to a myocardial infarct by mobilizing to the ischemic tissues and producing angiogenic cytokines, which stimulates the growth of new blood vessels at the damaged tissue site. The researchers noted that neoangiogenesis is the first major step "in preventing the vicious downward spiral of heart failure." The investigators use bone marrow cells from transgenic mice with the GFP (green fluorescent protein) marker for tracking the cells when they were implanted into a host deficient in c-kit+ stem cells. The data demonstrated that mobilization of the c-kit+ cells occurs through binding of the c-kit receptor with stem cell factor. Mice that were deficient in c-kit+ cells could neither home to the damaged tissue site nor repair the damaged heart. The researchers found that heart function in these mice were dramatically impaired 42 days after a heart attack. The authors noted that the results suggest that an infusion of bone marrow cells after a heart attack may benefit and prevent a patient from progressing to heart failure. Alternatively, they proposed that cells genetically engineered to secrete stem cell factor could be injected into the damaged tissue to enhance mobilization of the bone marrow cells to the heart tissue. The paper was published in The Journal of Thoracic and Cardiovascular Surgery.

June 30, 2006---In the June 25th online issue of Nature, Ron McKay and his collaborators at NIH reported the role of the Notch signaling pathway, which they confirm regulates neural differentiation and survival of both murine somatic and embryonic stem cells in vivo and in vitro. Notch encodes for a transmembrane receptor that is cleaved to release an intracellular fragment, Nicd. The cleaved fragment is directly involved in transcriptional control of target genes hairy and enhancer of split 3 (Hes3) as well as Sonic hedgehog (Shh). The intracellular cleavage product of Notch also regulates the serine/threonine kinase, Akt, and STAT3. Notch ligands such as Delta-like 4 (Dll4) and Jagged 1 (Jag1), when introduced into cell culture, rapidly reduce cell death. It was noted that the survival effect of Notch was blocked by DAPT, a γ-secretase inhibitor that prevented phosphorylation of Ser 727 on STAT3. The researchers used osmotic pumps to administer artificial cerebrospinal fluid (ACSF), FGF2, Dll4, or combination of FGF2 plus Dll4. The data showed that cells stimulated to divide by D114 survive for long periods in the parenchyma of the normal brain in an immature state. In a rat stroke model, in which the scientists occluded the cerebral artery, and delivered FGF2 and Dll4, significant improvements in motor skills were observed compared to the group receiving either ACSF, FGF, or D114 alone. The investigators also demonstrated in vitro that the Notch receptor had an important role in low-oxygen conditions, suggesting that the Notch ligands may "stimulate regenerative responses to the oxygen deprivation that follows ischemia in vivo."

June 29, 2006---Researchers from the Oklahoma Medical Research Foundation (Oklahoma City, OK) in collaboration with investigators from Osaka University reported in the June issue of Immunity the expression of toll-like receptors (TLR) on hematopoietic stem cells (HSCs). Toll-like receptors are known for their role in the recognition of microbial and viral antigens, which leads to innate immune responses. The research group discovered that TLR along with the Myd88 coreceptor drove the HSCs to differentiate into myeloid progenitor lineages as well as inducing lymphoid progenitors to become dendritic cells of the immune system. The investigators demonstrated that lipopolysaccharide (LPS), a B-cell mitogen, interacted directly with the TLR4/MD-2 receptor complex of both HSCs and downstream progenitor cells in the bone marrow. It was also reported by the authors that the epitope, CD14, enhanced the recognition of LPS by both stem and progenitor cells. The scientists proposed that further research may provide a means for replenishing and fortifying the innate immune system during infection.

June 28, 2006---Researchers from the Univ. of Washington reported in yesterday's issue of PNAS isolation of multipotent progenitor cells from human fetal liver. They reported the isolation of a human fetal liver multipotent progenitor cell (hFLMPC) population which is "capable of differentiating into liver and mesenchymal cell lineages." With feeder layers, the investigators were able to maintain the cells in an undifferentiated state for up to 100 population doublings. The average doubling time for the cells was 46 hours. The authors reported that the immunophenotype of hFLMPC is CD34⁺, CD90⁺, c-kit⁺, c-met⁺, EPCAM⁺, SSEA-4⁺, CK18⁺, CK19⁺, albumin^-, a-fetoprotein^-, CD44h⁺, and vimentin⁺. When the cells were cultured in special media, the researchers found that the hFLMPC differentiated into fat, bone cartilage, endothelial, hepatocytes and bile duct cells. Because the cells differentiated into bile duct and hepatic cells, the investigators deduced that the data suggest the hFLMPCs were a mesenchymal-epithelial transition cell that probably was of mesendodermal lineage. They also proposed that the hFLMPC could be a valuable tool for investigating human liver development, liver injury, and hepatic repopulation.

June 27, 2006---W. Huttner and his colleagues from the Max Planck Institute in Germany have been investigating with mouse embryonic neuroepithelial (NE) cells how stem and progenitor cells regulate either symmetric or asymmetric cell division. In the June 23rd online issue of PNAS, the researchers published their results on the role of the Aspm (abnormal spindle-like microcephaly-associated) protien in maintaining symmetrical proliferation of NE cells. The investigators found that Aspm concentrated on the mitotic spindle poles of NE cells during cell division. The protein is apparently down-regulated when cell switches from a prolifertive state to undergo neurogenic division. Down regulating Aspm was achieved through RNA interference. The results also suggested that the protein is crucial for "maintaining a cleavage plane orientation that allows symmetric, proliferative divisions of NE cells during brain development." It should also be noted that Huttner presented his research on the polarized organization of neural stem cells and its significance for symmetric versus asymmetric cell division at the Dec. 6, 2003 ASH annual meeting in San Diego. In his presentation, Huttner presented data focusing on the membrane protein, prominin (CD133) and its location in defining the apical region of the cell during mitosis. He also noted that prominin is associated with a novel, cholesterol-based lipid rafts. During cell division in NE cells, the cleavage plane relative to the apico-basal axis determined whether a cell maintained symmetrical cell division.

June 23, 2006---In yesterday's online issue of Stem Cells , University of Nebraska investigators, M. Mimeault and S.K. Batra, published their review of the recent advances in stem cells for tissue repair and cancer therapy. The paper reviews the most recent advances made in embryonic (ESCs), fetal, (FSCs), and adult stem cell research. The authors reviewed in depth "the localization of the adult stem cells and their niches as well as their implication in the tissue repair after injuries and during cancer progression." They also emphasized "the interactions between certain developmental signaling factors, such as hormones, epidermal growth factor (EGF), hedgehog, Wnt/{beta}-catenin and Notch." The authors further noted that "these factors and their pathways are involved in the stringent regulation of the self-renewal and/or differentiation of adult stem cells." The article also illustrated "novel strategies for the treatment of both diverse degenerating disorders, by cell replacement, and some metastatic cancer types, by molecular targeting multiple tumorigenic signaling elements in cancer progenitor cells."

June 22, 2006---Determining whether adult stem cells can circulate in the peripheral blood is a goal shared by many researchers in the field of adult stem cell biology. Other than CD34⁺ hematopoietic stem cells (and most recently our BLSCs), most investigators have had a difficult time either isolating or identifying the adult stem cell of interest in the blood. The inability to isolate stem cells in the blood has been attributed to the assumption that only a small number of stem cells are found in circulation. However, French scientists have discovered that multipotent mesechymal stem cells (MSCs) that can give rise to adipocytes, osteoblasts, chondrocytes, and vascular smooth muscle cells do circulate in the blood. They also found that these stem cells can be mobilized into the peripheral blood by hypoxia. The investigators published their results online in the June 15th issue of Stem Cells. In experiments with rats, there was approximately a 15-fold increase in the number of MSCs circulating in the blood and these MSCs were the same as those found in the bone marrow with respect to immunophenotype and functionality. Since the number of circulating hematopoietic stem cells did not dramatically increase, the investigators concluded that mobilizing MSCs into peripheral blood under hypoxic conditions was specific for MSCs.

June 21, 2006---In yesterday's press release, scientists from John Hopkins University reported that they were able to restore movement to paralyzed rats. The study encompassed the implantation of embryonic stem cells concomitant with a cocktail of drugs (3 drugs) and proteins (2 proteins) to repair damaged nerves in rats who had their spinal cord severed. In their experiments, the investigators reported that 11 out of the 15 paralzyed rats receiving the embryonic stem cell therapy "showed significant movement and the ability to control their hind limbs 24 weeks after the stem cells were implanted." The researchers also noted that if one of the components of the cell-based regimen was removed, i.e. either cells or one of the components in the cocktail, none of the rats within that group demonstrated improvement in their ability to stand or walk. The study and the results are to be pulblished in the Annals of Neurology. (It should be noted that in the past there have been periodic reports by other invesitgators who were able to restore hind limb mobility using adult stem cells from the bone marrow in a rat spinal cord injury animal model. It would be of great interest if the study's result could be repeated with adult stem cells.)

June 20, 2006---Stem cell biologists in recent years have focused their research on the regulatory mechanism(s) of stem cell homeostasis in various tissue compartments of the body. It has not been clearly delineated how the body regulates and maintains the constant turnover of cells in the tissues and organs of the body, in which old cells are replaced by new cells. It is well known that stem cells give rise to the new cells that replenish the effete cells of the body; however, there is a fine balance where too many new cells can give rise to excessive tissue growth and tumors. On the other hand, too few cells can lead to both tissue and organ degeneration. Swedish scientists from the Karolinska Institute in Stockholm reported in the June 15th issue of Cell that a key protein in the intestine, EphB, is one of the key regulators of proliferation and an endogenous mitogen for intestinal progenitor cells. Since more than 10 billion cells are generated every day in the human intestine, the researchers used the mouse intestine as a model for studying the role of EphB receptor signaling in regulating proliferation in the intestine. The scientists concluded from their data that EphB receptors are "key coordinators of migration and proliferation in the intestinal stem cell niche."

June 19, 2006---Scientists from the U.K. reported in last week's Nature online that the embryonic protein, Nanog, conferred pluripotency after cell fusion. Austin Smith and his colleagues at Cambridge University in the U.K found that by fusing embryonic stem (ES) cells with adult neural stem cells the pluripotent gene in the somatic cell's genome was activated. The researchers further demonstrated that Nanog from the embryonic cells resulted in a two-hundred fold increase in the number of recovered cell hybrids. The investigators also found that embryonic cell fusion with other somatic cells such as thymocytes increased the number of cell hybrids that could be recovered from the culture dish, but not to the same level as they observed with progenitor stem cells. In their experiments, cell hybrids were formed by using PEG and expression of pluripotency was measured by inserting the Oct4 gene along with the green flourescent protein transgene whose expression is under the control of the Oct4 transgene. In control experiments, the cell hybrids formed by fusing ES x ES cells resulted in two-fold increase in hybrid yield. The authors conclude that overexpression of Nanog significantly enhanced the ability to generate cell hybrids with adult progenitor cells and that "Nanog can instate pluripotency with up to 100% efficiency on the differentiation status of the somatic cells."

June 16, 2006---Researchers from the Salk Institute in La Jolla, California reported in the June 15th issue of Neuron another role for the growth factor, FGF (fibroblast growth factor), in which the protein serves as a chemoattractant for developing neurons that express the transcription factor LIM. The scientists noted that this transcription factor confers upon developing neurons navigational properties to innervate specific tissues during development. They also noted that the dermomyotome is a transient structure that gives rise to axial muscle and apparently produces chemoattractants for the medial-class spinal motor neurons (MMCm). The experimental results suggest that the dermomyotome secretes FGF in vitro, while the MMCm expresses the FGF receptor (FGFR1). Receptor expression in the MMCm is induced by the transcription factor LIM (Lhx3). The authors concluded that the "results point to a role for FGF signaling in axon guidance and further unravel how downstream effectors of LIM codes direct wiring of the developing nervous system." (It appears that researchers are discovering that FGF is an important molecule, similar to TGF-β in activating stem cells and modulating repair of damaged tissues. The press release from the Salk Institute noted that "the multi-tasking members of the FGF growth factor family regulate blood vessel formation, wound repair, lung maturation, and development of skeletal muscle, blood and bone marrow cells. The Salk study adds on more jobs to an already long list.")

June 15, 2006---In the June 14th issue of Journal of Neuroscience, Canadian scientists reported that skin progenitor cells from both mice and humans could be converted into Schwann cells. Schwann cells are a variety of neuroglia that mainly provide myelin insulation to axons in the peripheral nervous system of vertebrates. The investigators noted that Schwann cells derived in vitro from skin progenitor cells (SKPs), derived from the dermis of neonates, were able to produce myelin proteins when they were co-cultured and in close contact with sensory neurons. The researchers surmised that the dermis may serve as another source of neural progenitor stem cells since the dermis is formed from the neural crest during embryogenesis. They also speculated that if skin progenitor cells were isolated from the dermis they may respond to neural crest cues such as neuregulins when grown in tissue cultures. The results demonstrated that the skin progenitor cells differentiated into Schwann cells when grown in media containing forskolin and neuregulin-1β. When the differentiated Schwann cells from SKP were co-cultured with dorsal root ganglion neurons from mutant mice (shiverer mice) that lack basic myelin proteins, the Schwann cells became myelinated. Myelination occurred 3 weeks after cultivation with those cells that were in close proximity or in direct contact with axons. The investigators also demonstrated that naive SKP or differentiated SKP-derived Schwann cells from both mice and humans were able to able form myelinated axons when they were implanted into the brain or sciatic nerve of shiverer mice. The scientists proposed that the dermis may be a source of neural progenitor cells for treating various nervous system injuries and dysmyelinating disorders.

June 14, 2006---In yesterday's press release, the University of Buffalo announced that one of its stem cell scientists, Te-Chung Lee, Ph.D., received a $1.98 million research grant to investigate the feasbility of using bone marrow-derived stem cells to treat a serious heart condition called as hibernating myocardium. This condition is manifested by a loss of heart function as a result of reduced blood flow in certain regions of the heart over an extended period of time in which the stenotic arteries slow down the metabolism of cardiomyocytes to a "hibernating or resting state." Lee and his colleagues intend to use a swine model to determine whether bone marrow-derived mesenchymal stem cells (MSCs) can change the down-regulated myocardial adaptive response and "improve cardiac function of the hibernating myocardium." The characteristic, potential, and homing properties of the MSCs, as well as the potency of MSCs from older donors, will be studied in the first phases of the investigation. In the press release, Lee noted that these bone marrow-derived stem cells do not appear to generate a strong immune response in the host after transplantation. He intends to conduct experiments to rigorously explore immune response characteristics. If the lack of an immune response is established conclusively, future therapeutic applications could be based on off-the-shelf allogeneic stem cell products.

June 13, 2006---Scientists from the University of Florida reported in the most recent issue of Investigative Opthamalogy & Visual Science results on the generation of retinal pigment epithelium (RPE) from bone marrow-derived stem cells. Stem cells from the bone marrow of transgenic mice, which had been transfected with the gene encoding the green flourescent protein (gfp), were transplanted into lethally irradiated inbred strain of C57Bl/6 recipient mice. By breaching the Bruch's membrane of the eye in order to induce neovascularization and pre-treating with an injection of sodium iodate to damage the RPE, the investigators evaluated the recruitment of the bone marrow-derived stem cells to the damaged tissue site. The results of the experiments showed that bone marrow-derived stem cells can home to the damaged tissues and repair the RPE. Since the cells in the RPE contained gfp and the Y chromosome of the donor's cells, the researchers ruled out cell fusion and concluded that the bone marrow-derived cells were differentiating into RPE. Retinal pigment epithelium plays a vital role in forming the outer layer of the retina and supporting the neurons that receive light. Damage to the RPE is associated with diseases such as age-related macular degeneration (AMD). Over 1.7 million people in the U.S. are diagnosed with AMD. The scientists speculated that use of bone marrow-derived stem cell therapy to repair the damaged retinal tissues may provide an alternative to surgery.

June 12, 2006---In the June 9th online issue of PNAS, Japanese scientists published results that demonstrate the inductive activity of the amniotic membrane matrix on human embryonic stem cells. The researchers found that when embryonic stem cells were cultured in a serum-free medium containing the amniotic membrane matrices, the cells differentiated into various neuronal lineages of the central nervous system such as motor neurons, dopaminergic neurons, and retinal pigment epithelium. The investigators noted that similar results were obtained if the embryonic stem cells were co-cultured with mouse stromal cells. The scientists propose that such readily available human tissue provides "a suitable and versatile system for generating neural tissues for clinical application." (It should be noted that stem cell biologists are now realizing that extracellular matrices have important roles for growing and inducing both embryonic and adult stem cells to various tissue lineages. These matrices provide an unique microenvironment for maintaining the stem cells both in vitro and in vivo.)

June 9, 2006---In a press release from the University of Copenhagen, scientists discovered a family of enzymes that prevented cancer cells from "aging" like normal cells. The researchers found that the Jumonji-enzymes were upregulated and their levels in cancer cells were higher than in normal cells. During the normal aging process of cells, the investigators noted that the nucleus of the cell has more heterochromatin which indicates less active genes. With more densely packed DNA structures in the nucleus of older cells, these aged cells will eventually stop dividing. The Danish scientists discovered that the Jumonji-enzymes loosen the DNA structures. They found that the genes in the densely packed DNA were inactive, whereas in the loosened DNA structures an increase in gene activity could result in uncontrolled cell growth. They concluded that abnormal levels of these enzymes could result in cancer. The investigators speculated that these enzymes could also become potential drug targets to combat cancer. (If one accepts the paradigm that most cancers results from deranged stem cells, it would be of great interest to determine what role(s) these enzymes may have in maintaining adult stem cellness.)

June 8, 2006---In a press release yesterday, D. Bohmann and H. Jasper from the University of Rochester Medical Center, reported that upregulating the Jun kinase (JNK) signal transduction pathway in fruit flies increased lifespans by 40% (85 days instead of 60 days). The JNK pathway is particularly interesting since it has been characterized for its role in modulating the balance between cell growth and death during development, inflammation, and cancer. The researchers discovered that the JNK signaling pathway is activated during oxidative stress, an environmental condition involving the generation of oxygen free radicals that is accelerated duing the aging process. The authors noted that under oxidative stress, JNK targets the same protein as the insulin receptor found on the surfaces of cells. The researchers suggested that "JNK detects stress in the environment and not only prepares the body to deal with insults from the environment, but also limits the amount of insulin uptake, which prepares the body for stress by slowing its metabolism and limiting its energy output. It might also offer a new way to control insulin production." They also noted that the insulin receptor may also have a role in the aging process since they observed that restricting caloric intake results in increased lifespans from worms to flies, and perhaps humans. (Other investigators reported that JNK plays a critical role in dorsal closure of the fruit fly by controlling cell fate and cell sheet morphogenesis of the embryo. Thus, the JNK pathway may also have a critical role in adult stem cells by not only determining the cell's fate but also by targeting cancer stem cells and perhaps expanding the life span of humans.)

June 7, 2006---With new research tools for stem cell biology, scientists are now exploring the relationship between the loss of vigor in the adult stem cell compartment of an organism and the aging process. For example, it is well established that the wound healing response to repair damaged tissues is of longer duration with age. However, little is known about the intracellular nuclear proteins and their role in the aging process. In this regard, investigators from Northwestern University, NIH, and Sweden published their findings online in the June 6th issue of PNAS of the epigenetic effects of mutant lamina A (LAΔ50) in the premature aging disease, Hutchinson-Gilford Progeria Syndrome (HGPS). In HGPS, the mutant LAΔ50 protein appears to be the cause of the disease as a result of single base nucleotide substitution of the lamina A gene in chromosome 1. The mutation leads to an abnormally shaped nucleus and a loss of heterochromatin in a cell. The investigators demonstrated that the accumulation of LAΔ50 resulted in changes involving epigenetic controls for the facultative and constitutive heterochromatin in HGPS. The data suggest that LAΔ50 appears to alter the histone methylation sites regulating heterochromatin. In facultative heterochromatin, methylation of histone H3 (H3K27me3) and H3K20me3 are lost on the inactive X chromosome in cells from females with the disease and there is decreased methylation of histone H4 (H4K9me3) for the constitutive heterochromatin. The authors pointed out that this loss in histone methylation in HGPS cells increases as the cells age in culture. (It would be of interest to compare the histone methylation patterns for facultative and constitutive heterochromatin in stem cells from HGPS to methylation patterns in stem cells derived from normal individuals. The proposed experiment may provide one with greater insights into the aging process).

June 6, 2006----Scientists from Harvard Medical School reported in the June 5th online issue of PNAS that the tumor suppressor gene, Rb, is "dispensable for self-renewal and multilineage differentiation of hematopoietic stem cells. Data from other investigators suggest that regulators of the cell cycle from the G1-S phase transition determine a progenitor cell's fate for self-renewal. Since the retinoblastoma gene, Rb, has a central role in regulating G1-S phase of the cell cycle, the investigators conducted experiments to examine the hematopoietic stem cell (HSC) population (lin-c^-Kit⁺Sca-1⁺) in the bone marrow of mice which had been bred for conditional somatic deletion of the Rb allele. The researchers discovered that in the absence of Rb, the HSC contributed normally to both myeloid and lymphoid lineages in both primary and secondary recipients. Also, the investigators found that there was no transformation in the recipients receiving Rb negative HSCs. Thus, the authors concluded that Rb did not have an intrinsic role in the self-renewal of HSCs, but they did not rule out the possiblity that the gene may have an extrinsic role in maintaining either the microenvironment or the stem cell niche for hematopoiesis.

June 5, 2006---For stem cell biologists, the Notch genes are of interest for their role in various signaling pathways that determine a stem cell's fate. In the last couple of years, there has been an explosion in research papers on cancer stem cells in which scientists find cells within solid tumors that can differentiate into other tissue phenotyes. In the June 2nd issue of PNAS, scientists from Columbia and Harvard universities reported on the role of Notch1 in mammary tumorigenesis in transgenic mice. Within a mammary tumor virus containing LTR, the virus activated the intracellular domain of the Notch1 receptor, N1(IC), which activated Myc genes giving rise to the invasive form of adenocarcinoma. The authors established that Myc genes are direct transcriptional targets of N1(IC) and the data suggest that it acted upon the downstream Cbf1 transcriptional effector.

June 2, 2006---The American Society of Clinical Oncologists (ASCO) is holding its annual meeting in Atlanta from today until the 6th. J.A. Barrett et al. reported their findings on stem cell transplantation (SCT) to treat malignancy. After more than 35 years of clinical experience with SCT, outcomes have significantly improved over time as a result of greater experience with the therapy and improved supportive care for opportunistic infections following myeloablation. The authors noted the indications for SCT has widened as a result of volunteer donors, umbilical cord blood banks, and advances in HLA typing of mismatched tissues. However, morbidity and mortality with allogeneic SCT are still high, creating a barrier to greater therapeutic successes. The authors noted that new therapies such as imatinib (monoclonal antibody to CD22 for treating chronic myelogenous leukemia) as emerging therapies brings into the question the role of allogeneic SCT in the management of malignant diseases. The paper reviews significant advances and the changing indications for using allogeneic SCT in treating certain forms of cancers. (It should also be noted that better characterization of the stem cell population and cell dose needed for a particular therapeutic regimen may lead to more positive clinical outcomes and potential expansion of the use of allogeneic SCT in the future.)

June 1, 2006---At last week's American Urological Association annual meeting in Atlanta, scientists from Wake Forest University reported in a press release that they had artificially reconstructed penises in rabbits. The penises were engineered using penile tissues (most likely containing stem cells) and a collagen matrix. The artifical penises were found to be functional and maintained erectile pressures equal to normal rabbit penises. The researchers noted that their findings would provide a new treatment for males with certain erectile dysfunction such as corporal fibrosis. This condition results from the spongy tissue in the penis which is converted to scar tissue resulting from either an infection associated with a prosthetic implant or from a lack of oxygen associated with various cardiovascular diseases such as diabetes.

May 31, 2006---Scientists from Stony Brook and Rockefeller Universities in New York reviewed in Stem Cells the role of the gene, Lefty, which is expressed both in undifferentiated stem cells as well as a stem cells beginning to differentiate. The authors support the notion that Lefty is one of those unique genes which they determine is expressed in cells at the crossroad between either maintaining its state of "stemness" or differentiating into committed progenitor cells. The authors noted that Lefty is a member of the TGF-beta super family and a marker for embryonic stem cells. In mouse embryonic stem cells, LIF/Stat3 are important for maintaining the stemness; however, within 48 hrs. after cytokines are withdrawn, Lefty expression is upregulated. Similarly, Lefty expression is increased upon retoinic acid introduction (a differentiation inductive agent). In humans, Lefty is regulated by the transcription factors Oct3/4 and signaling by WNT and Smad2/3 pathways. The authors concluded that not only is Lefty important for maintaining the stem cell state of self-renewal, but it is also important for re-routing the cells as they exit such a state to a differentiation pathway. (It would be interesting to assay for Lefty expression in primitive adult stem cells).

May 30, 2006---P. Anversa and colleagues from New York Medical College published in the online issue of PNAS data identifying a cardiac stem cell (CSC) pool within human heart tissues. They examined the CSC pool with respect to telomerase and pool size following a response to tissue injury such as an acute myocardial infarct and chronic end-stage postinfarction cardiomyopathy. The results demonstrated that the number of CSC in the cardiac pool increased 28% following acute ischemia, 14% in chronic infarcts and 1.5% in controls. The authors concluded that a cardiac stem cell pool resides in heart tissues. The investigators noted that it was believed that myocardial regeneration was restricted to mycardium and adjacent to the infarct. However, the data presented suggest that CSCs reside throughout the heart and within an infarct. The ability of the CSCs to contribute and regenerate cardiomyocytes in response to ischemic injury may be a function of the size and competency of the CSC pool as well as the duration of the insult on the damaged tissue(s). (Our scientific founders have long demonstrated that pluripotent adult stem cells exist in most tissue compartments of the body. The results have been published in the above monograph by Young et al. demonstrating that these stem cells exists in all mammals. One should note that Aniversa did not characterize the cells in the CSC pool but used histological analysis of the biopsied tissues following an infarct.)

May 26, 2006---In the May 24th online issue of PNAS, scientists from Spain and the NIH reported that hypermethylation of CpG islands silences the WRN gene associated with Werner syndrome. Werner syndrome is an inherited disorder link to premature aging, genomic instability, and increased cancer risk. The disease is caused by mutations of the WRN gene, resulting in a loss of proper helicase and exonuclease activity. The researchers demonstrated that the epigenetic inactivation of WRN resulted in WRN-associated exonuclease activity and chromosomal instability as well as apoptosis induced by topoisomerase inhibitors. These detrimental cellular events were reversed either by demethylating agents of DNA or by insertion of active WRN into the cell. The investigators also presented data suggesting WRN-associated tumor suppressor activity. For example, in screening a large collection of primary tumors, the researhers demonstrated that hypermethylation of the WRN CpG islands were common in cancers of epithelial and mesenchymal origins. (It would also be of interest to determine whether epigenetic inactivation of WRN may have a regulatory role in adult stem cells with respect to self-renewal and differentiation.)

May 25, 2006---Researchers from New York's Memorial Sloan-Kettering Cancer Center and the Rockefeller University reported online in the May 21st issue of Nature that mono-, bi-, and trimethylated states of specific histone lysine residues of the chromatin have specific biological function ranging from heterochromatin formation, transcriptional regulation, and X-chromosome inactivation. Although the correlation between specific methylated sites and the biological read-out for its function is poorly understood, the authors found that read-out of histone H3 trimethylated at lysine 4 (H3K4me3) is through preferential binding of the PHD finger of the BPTF which is a subunit of the nucleosome remodeling factor (NURF). H3K4me3 is associated with transcription start sites of active genes. Using crystallography and NMR analysis of the protein structure, the researchers determined the molecular basis for the epigenetic events regulating gene expression. These proteins were critically important in maintaining the activity of the developmental Hox genes. (The data provide additional evidence supporting the concept of histone coding for segregating transcriptionally active from inactive chromatin domains in eukaryotic genomes. It would be very interesting to investigate the difference in specific regions of the chromatin of a somatic cell nucleus before and after it is implanted into an ex-nucleated oocyte. Histone coding may also be important for regulating the potential of adult stem cells as well.)

May 23, 2006---In the May 5th issue of Stem Cells, Dutch scientists Kamminga and de Haan reviewed the use of hematopoietic stem cells (HSC) as a model for correlating the aging process and stem cells. The paper addresses the roles of both extrinsic and intrinsic factors that contribute to the stem cell aging process, noting that the progenitor stem cell's cycling activity and the HSC pool size had a correlation with aging. The authors compared the difference in the stem cells in two inbred strains of mice with different life spans. In aged C57BL/6 (B6) mice, the cycle rate was slow with a small percentage of the HSCs in S-phase of the cell cycle, but the pool size of HSC was relatively stable during the aging process. In contrast, DBA/2 (D2) mice with a shorter life span than the B6 mice have more stem cells undergoing cell division. The D2 mice had a relatively larger pool of HSCs, but the pool size decreased with age. The authors suggested that these observations lead one to conclude that the stem cell compartment is pre-determined and thus, largely intrinsically regulated. Other factors contributing to the aging process and regulation of the various stem cell compartments within mammals are telomere shortening, DNA damage, and protein damage. The authors also noted that the capacity to maintain tissue lineage commitment following inductive signals could be attributed to epigenetic events involving cellular memory. Modification of the heterochromatin by regulatory proteins such as PcG complexes as well as heterochromatin re-localization may impart cellular memory and ultimately regulate the aging process. The authors concluded that the aging process begins in utero following the first division of the fertilized egg, which they define as pre-program events. Therefore, the gradual loss of stem cell potential is the underlying mechanism for the aging process.

May 22, 2006---At this year's annual meeting of the American Urologist Association held in Atlanta from May 20-25, Austrian scientists reported in their poster sessions the therapeutic use of muscle-derived stem cells from pigs and humans to treat incontinence. The clinical study, which began in September 2002, consisted of 186 patients, including 122 women and 63 men ranging in age from 36 years to 85 years old. After one year following treatment, 153 patients did not need to wear absorption pads. The therapy involved a biopsy of muscle tissue from a patient's upper arm and the use of nucleated cells (adult stem cells) grown for seven weeks in a collagen matrix. After cell cultivation, fibroblasts in the tissue cultures were injected back into the same patient's urethral submucosa. The cultivated myoblasts with additional collagen were also injected into the rhabdosphincter muscle that surrounds the urethra. The cells were delivered using a transurethal ultrasound probe. Postoperative improvement in thickness of the uretha (39%) and rhabosphincter muscle (52%), as well contractility of the rhabdosphincter muscle (265%) were measured by ultrasound. The authors concluded that the therapy was safe with no side-effects and represented a state-of-art use of autologous adult stem cells to treat urinary incontinence. The cost for the therapy was $16,000 per treatment.

May 19, 2006---Rudolf Jaenisch et al. from MIT and Univ. of Edinburgh reported in yesterday's online PNAS some of the underlying mechanisms resulting in the inability/inefficiencies to reprogram the donor genome to form embryonic stem cells after somatic cell nuclear transfer (SCNT). The authors demonstrated that the nucleus of neural stem cells produced higher number of embryonic stem cells from the inner cell mass of a blastocyst than terminally differentiated neuronal donor cells following nuclear transfer. The research group also presented data suggesting that hypomethylation of the donor's genome impart greater cloning efficiency. The authors concluded that differentiation and the epigenetic state of the donor's nucleus influence reprogramming efficiencies in forming embyronic stem cell clones.

May 18, 2006---A Scottish research team from the University of Edinburgh reported the discovery of a transcription factor, PouV, in the African claw frogs that is equivalent in function to the mammalian pluripotent trancsription factor, Oct4. It is well documented that Oct4 is a protein expressed in both embryonic and primordial adult stem cells. Oct4 is also known to regulate "stem cellness" by allowing cells to become units of self-renewal. In Development, the Edinburgh research team published their work where embryonic cells lacking Oct4 lost their unlimited proliferative potential and began to differentiate. However, if these same Oct4^- embryonic cells were transfected with PouV transgene, the transcription factor restored the embryonic cells' ability to self-renew. The researchers proposed that amphibians such as frogs and salamanders are other animal model systems that should be explored to uncover the underlying mechanisms and signaling pathways for regulating stem cells to either proliferate or differentiate. These animal models may provide greater insight into how stem cells are able to differentiate into a multitude of tissue types during both embryogenesis and tissue regeneration in adults. This complex process appears to be well conserved very early during the evolution of animals to humans.

May 17, 2006---Scientists from the Scripps Research Institute reported in the May 12th issue of J. Biological Chemistry a signaling pathway for regulating proliferation and differentiation of pancreatic progenitor cells. They noted that inhibitors of certain DNA transcription factors in stem cells behave as a switch which regulate both proliferation and differentiation of stem cells. The authors noted these transcription factors which drive differentiation are basic helix-loop-helix proteins and their inhibitors are Id proteins. With mouse pancreatic progenitors, the researchers found that BMP4 (bone morphogenetic proteins) signficantly reduced pancreatic epithelial ductal cell expansion while expanding the pancreatic progenitor cell population. The proposed regulatory mechanism is that BMP4 stimulation promotes the binding of Id2 (an inhibitor of bHLH and differentiation) to the NeuroD (bHLH) transcription factor. NeuroD binding is required for differentiation of progenitors to pancreatic islet cells. The authors propose a novel signaling pathway in which the presence of differentiation inhibitors (Id proteins) shift the balance in a population of progenitor cells to proliferate. The authors also suggest that understanding these regulatory pathways in stem cells can advance the technology for expanding progenitor cells in vitro for tissue regeneration; particularly, in the case of expanding pancreatic islets beta-cell mass for treating Type I diabetes. (There are other biological systems which support this concept. For example, it is well known in cellular immunology that there is a two-step signaling process in proliferating and differentiating antigen specific T cells from a reservoir of naive quiescent T cells residing within the thymus.)

May 16, 2006---Researchers from the National Cancer Institute reported in the May 15 issue of Cancer Cell an in-depth study on the use of human glioblastoma stem cell lines as a research tool for studying brain cancers. The authors noted that freshly isolated glioblastoma cells grown in culture media containing serum result in the propagation of a heterogeneous population of cells with morphologies that are reminiscent of fibroblasts and epithelial cells. However, if the glioma cells were grown in serum-free culture media supplemented with bFGF and EGF, the cells were clonogenic and had the potential for multilineage differentiation. The scientists described these cells as tumor stem cells (TSCs). In contrast, glioma cells grown in serum lost its tumorigenic potential. It was further noted that primary glioblastoma isolates grown in culture "bear remarkable similarity to neural stem cells." These similarities were reflected in the studies in which the scientists demonstrated that TSCs grown in serum-free defined media had the ability to form neurospheres in vitro, the potential for unlimited self-renewal, the ability to terminally differentiate into glial and neuronal lineages, and gene expression profiles that were similar to normal neural stem cells. The authors proposed that studies on the natural history of cancer and its etiology should in the future consist of tissue assay systems where the TSCs or primary cancer tissue isolates are studied in culture conditions that do not contain serum. (It was proposed over a decade ago that cancer biologists were on the wrong track by using transformed cell lines to study various forms of cancers. Gerald B. Dermer, Ph.D. in his 1994 book, "The Immortal Cell: Why Cancer Research Fails" describes the failure of cancer research to recognize that using transformed cell lines to conduct cancer research was a path doomed for failure. Dermer noted that the transformed cells were artifacts and thus, the data being published were not meaningful with respect to cancer biology. Similarly, it should be noted that the above paper supports the concept that serum is a wound-healing inductive agent that leads to stem cell cultures giving rise to a heterogenous population of lineage committed cells.)

May 15, 2006---In yeterday's online of edition of Nature Neuroscience, researchers at Children's Hospital in Boston reported the discovery of a new growth factor that stimulated the growth of nerve fibers in the brain and spinal cord. This new factor, oncomodulin, induced in vitro a 5-7 fold increase in regeneration of axons from retinal ganglion cells. This protein is secreted by macrophages as an extremely acidic calcium-binding protein. Oncomodulin is found in both fetal placenta and neoplastic tissues. In their previous studies, the researchers fortuitously discovered that injury to the optic nerve activated axon growth. The activation process was initiated by macrophage infiltration to the tissue injury site as a result of inflammation. The nerve fibers respond to oncomodulin when intracellular levels of cyclic-AMP are increased. (It should be noted that these findings support the concept that in wound and tissue repair the inflammatory response results in debridement of the damaged tissues by phagocytic cells such as macrophages. Another interpretation of the data may be that infiltration of phagocytic cells leads to activation of resident undifferentiated stem cells to regenerate axons.)

May 12, 2006---In this month's issue of Nature Reviews Neuroscience, Italian scientists G. Martino and S. Pluchino review the therapeutic potential of neural precursor stem cells. The paper provides an excellent overview of the advances that have been made with adult neural progenitor/precursor cells (aNPCs). The authors noted that in chronic neurodegenerative diseases, whereby there are multifocal lesions in the brain, chronic inflammation leads to recruitment of aNPCs to the damaged tissue sites. However, the wound-healing response usually leads to scar formation and permanent damage of the tissues in the CNS. Furthermore, the authors cited in vivo results in which aNPCs implanted at the diseased sites were not able to regenerate the damaged tissues back to a fully functional state. It was noted, however, that the implanted aNPCs demonstrated a neuroprotective effect, but the implanted cells were incapable of terminally differentiating into the appropriate neural phenotypes when the tissues were examined histologically. Therefore, the authors suggested that therapeutic efficacy reported by others may be attributed to bystander mechanism(s) in which the implanted cells form an atypical ecotopic (perivascular) niche by providing neurotrophic and immunomodulation support to the neural stem cells within the germinal centers of the subventricular zone in the lateral ventricles and the subgranular zone in the hippocampal dentate gyrus of the brain during the tissue remodeling process. The authors noted that immunomodulation by the transplanted aNPCs may occur from the cells secreting soluble molecules such as certain cytokines, chemokines, and various surface receptors that may downregulate inflammatory T cells and macrophages at the targeted tissues. (Another hypothesis is that the aNPCs may not be the correct cells for remodeling the damaged tissues. We have observed that in order to obtain good tissue integration, one needs to implant a more primitive lineage of uncommitted stem cells within target tissues such as brain, heart, or skeletal muscle.)

May 11, 2006---In this month's issue of Nature Medicine, scientists from Cornell Medical College in New York reported their findings which delineate another mechanism for neovascularization/revascularization in chronically damaged or diseased tissues. The researcher reported that chemokine-mediated release of SDF-1 (stoma-derived factor 1) from platelets recruited CXCR4⁺ VEGFR1⁺ bone marrow-derived hematopoietic stem cells to home to the damaged tissue site and form new blood vessels. The authors noted that VEGF (vascular endolthelial growth factor), which has long been know to induce angiogenesis, had a short term effect on neovascularization of ischemic tissues associated with chronic diseases such as diabetes. During inflammation, however, chemokines such as GM-CSF, TPO (thrombopoietin), EPO (erythropoietin), and sKitL (soluble Kit ligand) induce blood platelets to secrete SDF-1 at the site of tissue inflammation and recruit bone marrow stem cells to form new blood vessels. By ligating the femoral artery in the hindlimb of mice, the ischemia resulted in an increase in plasma levels of the above chemokines listed along with a subsequent increase in SDF-1 levels. Angiogenesis resulted from CXCR4⁺ VEGFR⁺ mobilized from the bone marrow to the ischemic tissues. Furthermore, ligation of the hindlimb in TPO and sKitL knock-out mice had a profound affect in suppressing neo-angiogenesis. The authors proposed that regulating SDF-1 levels in damaged tissues may be a new therapeutic approach for managing diabetics with poor blood circulation to the patients' limbs and prevent amputation of those limbs.

May 10, 2006---In yesterday's press release, Oregon Health & Science University in Portland announced that their scientists had published in PNAS results supporting their thesis about the role of cell fusion occurring not only in normal intestinal epithelium, where cells are proliferating at a high rate, but also as a possible mechanism for tumorigenesis in intestinal adenomas where cells are hyperproliferating. The paper was published in the April 10th issue of PNAS which was discussed in our April 11th post below (with a link for downloading the paper). In our April 11th post, we did not discuss the possibility of cell fusion giving rise to a predisposition for cancers. In that regard, the paper describes experiments where bone marrow-derived cells double-labelled with EGFP and LacZ (ROSA26 mice) were injected into male recipients. Cell fusion was measured by the presence of the recipient's Y chromosome and the double label in cells lining the intestinal epithelium. The authors demonstrated that cell fusion occurred in the recipient's tissues where there was a propensity for a high turnover of cells in normal tissues as well as in mice bred for intestinal adenomas (Min mice). The researchers ruled out phagocytosis by the donor's macrophages since immunohistochemical assays did not reflect monoclonal ant-CD45 staining in cells with the double label and the Y chromosome marker. However, the authors noted in the paper that 60% of the Y chromosome were present in the donor derived cells (We find this result problematic. This finding does not conclusively rule out phagocytosis by donor-derived macrophages or other phagocytic cells as an alternative explanation for the published results. In our view, the data may be more meaningful if the authors were able to conduct Cre/lox recombination assays to support their hypothesis of cell fusion by bone marrow-derived cells trafficking through tissues where cells are hyperproliferating.)

May 9, 2006---Chinese scientists from the Shanghai Institute of Genetics reported in the May 8th online edition of PNAS, the engraftment of human, cord blood-derived, hematopoietic stem cells into fetal goats (45-55 days of gestation). The researchers had isolated and transfected a primitive population of CD34⁺Lin^- from the cord blood of males with GFP. The postnatal goat tissue were examined for GFP⁺ cells and found the cells had differentiated and integrated into both hematopoietic and nonhematopoietic organs including blood, bone marrow, spleen, muscle, kidney, liver and heart of the recipient goat. Two years after birth, the investigators found by microarray analysis that the livers and blood of the goat expressed human genes. The researchers ruled out the possibility of cell fusion by DNA content analysis. The authors proposed their human/goat xenotransplant model could be used for studying stem cell kinetics for engraftment, gene expression and plasticity "under noninjured conditions." (Although the paper describes an elegant set of experiments, there is still the question of purity of the CD34⁺Lin^- hematopoietic stem cells used in the study as well as their ability to self-renew in vitro.)

May 8, 2006---In today's online publication in Nature Immunology, scientists (Ken Shortman, Donald Metcalf et al.) from the Walter and Eliza Hall Institute in Australia published their findings of an unique subpopulation of progenitor dendritic cells (DCs) from the spleens of mice. These precursor cells of the immune system were pre-committed to form both CD8⁺ and CD8^- conventional dendritic cells (cDCs) of hematopoietic stem cells giving rise to various subpopulations. With flow cytometry, the authors found the cells were CD11c^int, CD45RA^lo, CD43^int, SIRP-α^int, CD4^-, CD8^-. and MHC-II negative. The researchers noted that the pre-DCs comprised 0.05% of the splenocytes and they were not monocytes. In adoptive transfer experiments, monocytes were unable to generate cDCs and maintain a reservoir of cDCs in the spleens of the recipients. The authors noted that a distinct population of spleen DCs were formed during inflamation concomitant with the presence of GM-CSF (granulocyte-macrophage colony stimulating factor). By manipulating the various inductive pathways to form various subtypes of DCs, the scientists speculated on the feasibility of using various subpopulations DCs to mitigate autoimmune diseases.

May 5, 2006---In the April 24th issue of Stem Cells, Priddle et al. from the Univ. of Nottingham (Nottingham, U.K.) provide an overview of exploring various approaches in overcoming the immune barrier for allogeneic stem cell therapies, specifically focusing on embryonic stem cells. The authors noted that in experiments where one could establishe hematopoietic mixed chimeras, the chimerics appeared immune tolerant to the alloantigens of the donor cells and more receptive to an allograft. These observations were confirmed in vitro with mixed lymphocyte reactions (MLRs). Priddle proposed that inducing tolerance in the recipient with donor hematopoietic stem cells is an approach that should be further explored prior to implanting pluripotent stem cells. (This approach of using hematopoietic stem cells as tolerogens could also be explored with adult stem cells, particularly in regenerating tissues damaged by an autoimmune response which can give rise to diseases such as type I diabetes and Lupus.)

Added note: It has been brought to our attention that in the May 3rd post regarding Ratajczak's paper in this month's Leukemia, the work was first presented last December at ASH in Atlanta and subsequently published online in the Feb. 23rd publication of Leukemia.

May 4, 2006---The University of Pittsburgh announced in yesterday's press relaease that they have received $3.7 million from the Defensed Advanced Research Projects Agency (DARPA) a grant to conduct research on limb regeneration. DARPA has funded a number of institutions for conducting research on the underlying mechanism for regenerating limbs in salamanders and human fetuses. In salamanders, the process for regenerating limbs is the formation of the blastema, which is a ridge of stem cells formed at the outer margins of the wound site. This wound-healing and limb regeneration capacity exist in human fetuses but lost after birth. For a more detailed exposition on salamander limb regeneration, download the monograph above (the section begins on page 5). An unfortunate consequence of war is the loss of limbs. In Iraq, approximately 6% of our soldiers (16,000) have either lost their limbs during combat or had them amputated as a result of a wound.

May 3, 2006---Ratajczak et al. from the Univ. of Louisville published in this month's issue of Leukemia a very small embryonic-like pluripotent stem cell in murine bone marrow. The investigators reported a very small Sca-1⁺ lin^- CD45^- cell in the bone marrow of mice. These very small embryonic-like (VSEL) cells represented 0.02% of the nucleated cells aspirated from the marrow with sizes ranging from 2-4 microns in diameter. The authors reported that the VSEL cells have a gene expression profile similar to embryonic cells. With real-time RT-PCR, the researchers demonstrated the VSEL cells expressed the surface marker SSEA-1 along with the transcription factors Nanog, Rex-1, and Oct-4. However, the inability to propagate these micro cells in vitro required co-culturing experiments to demonstrate pluripotency of the cells. In co-culture experiments, the researchers were able to induce gene expression profiles representing the phenotypes of cells from the three germ layer lineages. The scientists also noted that the cells expressed the chemokine receptor CXCR-4, which provided the cells the capacity to aggregrate around and on bone marrow-derived fibroblasts as well as home to damaged tissue sites following injection of the cells into the tail vein. The research group also reported a higher percentage of VSEL cells were in the bone marrow of young mice and the numbers decreased with age. (This paper is of great interest since it supports the observations of our proprietary BLSCs described in our Technology Section. There has been skepticism by the general scientific community whether such small cells existed in postnatal tissues; despite others periodically describing a small "side-population of cells found in the bone marrow. It is noteworthy that the authors were unable to grow their cells. Some of this difficulty may be attributed to using low concentrations of serum as one of the growth constituents in their culture media. The paper also supports our view that reports of transdifferentiation in bone-marrow-derived nucleated cells may simply be a case of "overlapping" populations of non-hematopoietic populations of bone marrow-derived stem cells not identified by other investigators.)

May 2, 2006---In yesterday's press release, Israeli scientist, Blondheim et al., reported that bone marrow-derived mesenchymal stem cells are predisposed to become neurons under appropriate induction conditions. In their in vitro studies published in last month's Journal of Stem Cells and Development, the group reported an adherent cell population that were positive for the surface marker CD29, and after 14 days of cultivation the adherent cell population became predominantly positive for the surface markers CD29, CD44, CD90, and CD105. With RT-PCR analysis of the adherent cell population, the investigators discovered the cells expressed 12 neural genes and 8 genes related to dopaminergic neurons. The scientists noted that since these bone marrow-derived mesenchymal stem cells had a predisposition to form neurons, the cells could be used in an autologous cell-based therapeutic approach in the future to treat neurodegenerative diseases. (Although the data appeared interesting, the authors failed to cite the work noted in our April 28th post of the paper in the J. Neuroscience Research by Romero-Ramos, Chesselet et al. published in 2002. See April 28th post below for access to the paper. Furthermore, the above group does not describe either: 1. the ability to expand the adherent cell population concomitant with describing the number of cell culture passages while still maintaining neuronal gene expression; or 2. the purity of the population used to conduct their gene expression profile analysis. One can not rule out from the published data the possibility that the investigators are cultivating a mix population of bone marrow-derived cells consisting of both multipotent/pluripotent stem cells and mesenchymal progenitors. It is conceivable that the cell of interest perhaps may not express all of the surface markers described in the paper. The mesenchymal progenitors reported to be nestin positive and have the surface marker CD105 seem problematic in supporting their thesis that they have discovered an unique set of mesenchymal progenitors which have a predisposition for various neuronal phenotypes.)

May 1, 2006---In the April 28th online issue of PNAS, researchers from Mass. General, Harvard University Medical School, and Tufts University published their results in determining the role of the retinoblastoma gene, pRB, in cochlear and vestibular hair cell development and related hearing loss. In a conditional knockout mouse model with the pRB gene deleted in hair cells of the inner ear, the progeny mice survived up to 6 months, but lost their hearing and balance. From the published data, the pRB tumor suppressor gene appears to also regulate function and maturation in specific postnatal cochlear and vestibular hair cells. As a tumor suppressor gene, the Rb protein is found to regulate cellular proliferation which induces embryonic mammalian hair cells to exit the cell cycle, but does not induce differentiation. The authors reported that the Rb protein appears to have different regulatory roles with respect to maturation of postnatal auditory hair cells and proliferation of the supporting cells in the inner ear. The investigators propose that regulating transient non-expression of pRb may be an useful therapeutic approach for treating hearing loss in a population of auditory hair cells that do not spontaneouly regenerate. (It is not surprising to learn that many of the these "so-call oncogenes and tumor suppressor genes are also having stem cell regulatory function with respect to the pool of progenitor cells exiting the cell-cycle, and induction and differentiation of the stem cells into different tissue phenotypes).

April 28, 2006---In yesterday's press release, scientists from the University of Rochester in New York announced that they have advanced the stem cell technology needed to regenerate neural tissues damaged as a result of a spinal cord injury. The investigators noted that in their rat models regeneration of the damaged neural tissues required the implantation of glial-derived astrocytes (GDA) at the damaged tissue site. The researchers reported that rats which did not receive pre-induced GDA along with embryonic stem cells were not able to regenerate their nerves and had difficulty walking after the spinal cord injury 4 weeks post-implantation. The scientists speculated that GDA and its presence in damaged tissue is necessary to prevent scar tissue formation, rescuing motor pathway neurons, and remodeling the damaged tissue back to a functional state. (This raises the question about the need for using embryonic stem cells in their experimental protocol. It has been frequently reported that adult stem cells from various tissues can also restore motor function in various spinal cord injury animal models. Click here for a paper reporting the use of muscle-derived stem cells from rats differentiating into neurons in cell cultures.)

April 27, 2006---In last week's press release, a Scottsdale, Arizona-based company announced they were moving their operations to Costa Rica in order to treat patients who had suffered a stroke and cancer patients using umbilical cord blood-derived stem cells. What is not clear from the press release is the treat regimen with respect to dose, number of treatment cycles, cost of the treatment, or even preclinical data supporting their therapeutic approach for implanting the cells into patients. Are these just opportunistic companies who are taking advantage of the rising wave in stem cell research by moving their operations offshore? Are they potentially preying on desperate people who are willing and can afford to pay a large sum of money for unproven therapy? There are rumors of companies using bone marrow-derived stem cells and treating patients for various chronic diseases in different foreign countries for $300,000 per implantation of allogeneic stem cells. We may never know the results of the therapies being provided by these companies, except possibly for some anecdotal "before and after" marketing videos of patients who were successfully treated. How will these companies impact companies who are investing a large sum of money for conducting preclinical and clinical trials in the U.S. in order to obtain FDA approval for commercializing their cell-based therapies? These types of questions and topics will be raised and discussed in the future in our Forum section.

April 26, 2006---Scientists from Beth Israel Deaconess Medical Center and Harvard Medical School announced in a press release that in an animal model of Alport Syndrome, they were able to repair the damaged kidney with transplanted allogeneic bone marrow cells. The experiments involved knock-out mice deficient in type IV collagen. Alport Syndrome is an inherited disease in which there is a deficiency of type IV collagen in the basement membranes of the glomeruli of the kidneys, which leads to kidney failures. Patients with the disease must undergo kidney dialysis for their whole life. The experimental results were published in the online April 24th issue of PNAS demonstrating that bone-marrow-derived allogeneic adult stem cells can repair the defect by producing the various type IV collagens making up the extracellular matrix of the kidney's basement membrane. (We have seen many of these reported experimental results in which stem cells from either bone marrow, fat, or umbilical cord blood have an effect on repairing various types of damaged tissues. However, the stem cell population does not appear to be fully characterized by the investigators. We call it plug-in-play type of science.)

April 25, 2006---Japanese scientists reported in a paper published in yesterday's online edition of PNAS that they the soluble carbohydrate-protein compound, Galectin-1, promotes the proliferation and differentiation of neural progenitor stem cells. The neural progenitor stem cells that responded to this growth factor are found in the subventricular (SVZ) and hippocampal detente gyrus regions of the forebrain. The researchers reported that Galectin-1, in vitro, induced a mouse cell line, OP9CM, to form neurospheres prior to the stem cells differentiating into neurons and glial cells. In vivo experiments demonstrated that infusion of Galectin-1 significantly increase proliferation of progenitor stem cells in the mouse forebrains. It was also noted by the authors that other groups had reported Galectin-1 supporting the growth of hematopoietic stem cells in mammals.

April 24, 2006---At a EuroStell conference in Italy, scientists from the Netherlands announced today in a press release that they were able to grow sperm-producing stem cells in mouse testes. The research group also reported that glial-derived neurotrophic factor (GNDF) was also involved in spermatogenesis. High levels of GDNF were found in neonates which decreased during adulthood. They also noted that fibroblast growth factor was needed for growing the stem cells. With their breakthrough in the growth of spermatogonial stem cells, the investigators speculated that they may now have an approach for restoring male fertility following cancer therapy. At the same conference, scientists from the University of Iceland reported that they are developing methods for growing stem cells derived from tissue biopsies of breast cancer patients. They believe that over twenty percent of the breast cancers involve diseased stem cells. With three-dimensional matrices for growing cells, the researchers are investigating cell-cell interactions and cell signaling pathways. The group is particularly interested in the tyrosine kinase receptor signal pathways and the downstream signaling events in a both normal and cancer cells.

April 21, 2006---In yesterday's press release, Rush University Medical Center in Chicago announced they would be one of 15 sites participating in a Phase I clinical study to treat heart attack patients with bone marrow-derived meschymal stem cells (MSCs). The study will be a double-blind, placebo-controlled clinical trial in which allogeneic MSCs will be delivered intravenously to the patient. The stem cells being tested are Osiris Therapeutics' mesenchymal progenitor cells having the surface markers CD29, CD44, CD105, and CD166. The company has claimed that these cells are immune-privileged stem cells that are not rejected by a recipient's immune system. It is also claimed that these MSCs can migrate to the site of inflammation or tissue damage following their infusion into the patient's blood.

April 20, 2006---Scientists from MIT published a paper online in yesterday's Nature as well as in Cell describing the role of the Polycomb Group of protein (PcG) complexes which are known to be transcriptional repressors that maintain "stem cellness" and the ability to self-renew. With mouse embryonic stem cells, the investigators demonstrated that PcG appears to regulate both embryonic-associated stem cell transcription factors and genes involved in maintaining pluripotency as well as differentiation of embryonic cells. The researchers demonstrated that one of the Polycomb repressive complexes, PRC2, regulated many embryonic-associated transcription factors including Oct 4, Nanog, and SOX2. The involvement of PcG complexes in adult stem cells is still unknown due to the lack of a purified population of adult pluripotent stem cells; however, in a 2004 review article by Valk-Lingbeek et al. in Cell, the authors noted that PcG serving as epigenetic chromatin modifiers were involved in cancer development as well as in the maintenance of both embryonic and adult stem cells.

April 19, 2006---Scientists at Ohio State University reported that either overexpression or dysfunctional microRNA (miRNAs) can give rise to various forms of cancers. Carlo Croce and his group reported online in PNAS that they had observed miRNA155 being linked to various forms of cancers. His group demonstrated that overexpression of the message gave rise to leukemias and lymphomas and, in breast cancer patients, its overexpression in the malignant tissues indicated a poor prognosis. In transgenic animals, the researchers discovered that miRNA blocked differentiation of B cells in the hypertrophic spleens of mice which suggested that miRNA block differentiation of hematopoietic stem cells. They noted that while the mechanisms where certain groups of miRNAs act as oncogenes while other act as tumor suppressor genes have not been delineated, they suggested that a new class of artificially designed molecules (antagomirs) that block the miRNA155 expression could be used as a new therapeutic approach to treat patients with specific forms of leukemias or lymphomas.

April 18, 2006---In the April issue of the New Scientist, scientists from the Univ. of Glasgow reported that they were able to direct bone marrow-derived stem cells to differentiate into a specific tissue lineage when the cells were grown on a solid substrate that had been etched with different nano-patterns. Using electron beam lithography, the same technology used to manufacture microchips, the researchers found that they could coax the stem cells to become muscle, bone, cartilage, or fat when the cells were grown on a substrate containing different type of etched patterns. The nano-patterns consisted of nano-pits produced in various types of solid polymeric surfaces. The ratio of order to non-ordered nano-patterns also affected stem cell differentiation as well as the cells' rate of growth.

April 17, 2006---In today's online issue of The Scientist, there is an interesting article by Kenney et al. from the Lawrence Berkeley National Laboratory describing the interplay between neoplastic cells, the "disordered" microenvironment which gives rise to a tumor, and the wound-healing process (see our April 6th posting regarding the relationship between wound-healing and stem cells giving rise to cancers). The authors make the case that tumor progression (Ed. Note: It would be interesting to investigate whether mutations of tumor suppressor genes such as EGFR, p53, and PTEN of a particular subpopulation of resident adult stem cells are more apt to give rise to the cancer stem cells) is an interplay between the neoplastic cells and local microenvironment, particularly, the tumor stroma that is coevolving with the tumor. The authors further make the analogy that tumor formation is not a lone cellular renegade, but "akin to a desperate addict needing to get its fix from the dealers in the neighborhood." The investigators also emphasized the role of the extracellular matrix in regulating cellular responses and their receptors as potential therapeutic targets as anti-cancer therapeutics.

April 14, 2006---Japanese scientists report in the April 18th issue of J. Am. College of Cardiology the discovery of a placenta growth factor, P1GF, that apparently triggers new blood vessel formation after a heart attack (acute myocardial infarct/AMI). Placenta growth factor was found to have wound-healing activity by activating hemapoietic stem (flt-3 positive) cells and inducing angiogensis. In a 98-patient, placebo-controlled study, the plasma levels of P1GF are significantly elevated following an AMI. The investigators concluded that overexpression of P1GF by endothelial cells is a compensatory mechanism to repair tissue damage after a heart attack. They also noted that VEGF (vascular endothelial growth factor), another angiogenic factor, could induce formation of new blood vessels, but that the newly formed blood vessels function poorly. The researchers propose that P1GF (as a stem cell growth factor?) could be another therapeutic approach for limiting tissue damage and restoring cardiac function following an AMI.

April 13, 2006---In a press release yesterday, scientists at Northwestern University announced that their findings may suggest a paradigm shift in the pathogenesis of heart valves. It was previously thought that damage to heart valves was associated with wear and tear. However, the Northwestern investigators noted that high cholesterol in the blood triggers chronic inflammation around the various heart valves which causes cells in those tissues to "reprogram," taking on a bone phenotype in the aortic valve and cartilage phenotype in the mitral valve. Their results will be published in the April 18th issue of the J. American College of Cardiology. The researchers further noted that conventional wisdom associated the aging process to calcification of these valves. (It would be interesting to investigate whether there is actually a reprogramming of these valve-derived cells or whether resident stem cells are being activated and induced to become bone or cartilage as result of inflammatory cytokines and high levels of cholesterol circulating in the blood. Further, in the Nov. 5, 2004 issue of Science, there is speculation about the relationship between chronic inflammation and cancer. Stem cells may be the common link between various disease processes and wound-healing. During chronic inflammation, various forms of disease may arise as a result of mutated or diseased stem cells. See our perspective on Stem Cells and Cancer.)

April 12, 2006---The Los Angeles Times reported yesterday that $12.1 million in training grants were awarded to 16 California institutions to conduct stem cell research in the fields of umbilical cord blood, embryonic and adult stem cells. The grants will be used to establish training courses and support graduate, post-doctoral and clinical fellows. The grants were funded by the issuance of a $14 million Bond Anticipation Notes (BAN), approved last week by The California Stem Cell Research and Cures Finance Committee, to six philanthropic groups in California who purchased the notes. The notes are redeemable if the state issues the $3 billion General Obligation bonds approved by voters in 2004 through the passage of Prop. 71. However, the California Institute of Regenerative Medicine (CIRM) has been in a legal imbroglio with various groups challenging the constitutional aspects of the voter-approved initiative. If the courts rule in favor of the plaintiff, particularly in the appeal process, the BAN holders will not be able to redeem the notes and their investments would convert to a donation. Additionally, UCLA announced in a press release that they were awarded $1.23 million in grants, the largest amount awarded by the CIRM, to fund 16 trainees and scientists associated with the University's recently formed Institute of Stem Cell Biology and Medicine (ISCBM).

April 11, 2006---Scientists from Univ. of Oregon reported in the April 10th online issue of PNAS that bone marrow-derived cells, when transplanted into recipient mice, fused with normal and neoplastic intestinal epithelial cells of the host. By using either unfractionated bone marrow-derived cells or hemapoietic stem cells (HSCs), the investigators concluded that the female donors' HSCs were fusing with the recipients' cells in the small intestine as early as two weeks and as late as 14 months after transplantation. The male recipient had been lethally irradiated 4 days prior to transplantation. The researchers speculated that cell fusion between HSCs and the local stem cell niche of the small intestine was one of the mechanisms for regenerating intestinal epithelium that was damaged or diseased. Cell fusion was not observed in non-irradiated mice. Cell fusion was assayed by staining cells expressing both the donor's double marker (EGFP and β-gal) and the recipient's Y chromosome.

April 10, 2006---Researchers at the NIH announced in a press release that an adult stem cell factor (SCF) promotes angiogenesis when it is overexpressed in patients with brain tumors as well as following brain injury. The investigators noted that the growth of newly formed blood vessels, which supports the survival and expansion of the brain tumors, results in a poor prognosis for cancer patients, particularly, those who are diagnosed with glioblastomas. Since SCF is also expressed in patients with brain trauma, the researchers speculated that standard invasive procedures such as a surgical biopsy or surgical resection of the brain tumor may actually induce a proangiogenic response which may help promote survival and growth of the cancer. The results were reported in this month's issue of Cancer Cell.

April 7, 2006---Scientists at Univ. of California, San Diego and Stanford University reported in a press release yesterday that a mutation in hematopoietic stem cells (HSCs) give rise to the blood disorder polycythemia vera. The investigators further noted in today's PNAS that their data supported the notion of targeting mutated or dysfunctional adult stem cells may provide greater insight into how cancer may arise from these stem cells (see "Stem Cells and Cancer in our Technologies Section). In last year's May 3rd issue of Science, Jamieson et al. reported that a mutation in JAK2 (a non-receptor tyrosine kinase), in which one amino acid substitution from valine to phenylalanine at position 617 in the signal transducing protein of HSCs, results in bypassing activation of the erythropoietin receptor which eventually leads to excessive production of red and white blood cells. Anne Leonard noted yesterday in her Stem Cell Research Progress Blog that S. Morrison and his research group at the University of Michigan, are also exploring various therapeutic approaches to target "cancer stem cells" that give rise to leukemia. In collaboration with Hong Wu from UCLA, the investigators have found that targeting the tumor suppressor gene, Pten (phosphatase and tensin homolog), with the anti-cancer drug, rapamycin, selectively kills the cancer-causing hematopoietic stem cells.

April 6, 2006---In last month's issue of Stem Cells, Mannello et al. from the Stem Cell Institute at the Univ. of Bologna (Bologna, Italy) reported on the role of matrix metalloproteinases (MMP/matrixins) during a wound-healing/tissue regeneration response. Their findings suggest a regulatory role for MMPs for proliferation and differentiation of mesenchymal stem cells. (It is also known that activation of quiescent adult stem cells within a given tissue is critically important for tissue maintenance or repair. At last month's symposium of Los Angeles Wound-Healing Initiative at the Saban Inst. in the Children's Hospital of Los Angeles, W. Garner and his group from the USC School of Medicine reported their findings which suggest a critical role for MMP-9 in the wound-healing process. They found that overexpression of MMP-9 resulted in chronic wounds and inhibited migration of keratinocytes to the wound site. Using a burn wound model, Garner's group also reported that MMP-9 activity within a wound is regulated by alpha-ACT and stromal factor TIMP-1.)

April 5, 2006---Researchers at Duke University announced in a press release the use of growth factors that enhance differentiation of human adipose-derived adult stem cells (hADAS) into bone and cartilage. The scientists reported that when their fat-derived (from liposuction) stem cells were grown in a cocktail consisting of BMP-6, the cells not only differentiated into chondrocytes, but they also observed a 2 log increase in the expression of the extracellular matrix, aggrecan, and a 3-fold increase in type II collagen production. These two macromolecules are important components for remodeling the articular cartilage and providing the tensile strength needed for normal tissue and joint function.

April 4, 2006---Israeli scientists from the Hebrew University in Jerusalem reported repairing Achilles tendons in mice. The investigators used progenitor mesenchymal stem cell lines, C3H10T1/2, which were transfected with genes encoding for Smad8 and BMP-2. Activation of Smad8 by BMP2 enhance differentiation and tenocytic repair of the damaged Achilles tendon. The data was reported in this month's issue of J. Clinical Investigation.

April 4, 2006---Researchers from Wake Forest and Harvard Universities announced that they have grown urinary bladders in tissue cultures. Using a small piece of bladder tissue from each patient, the bladder-derived cells were incubated for a week and then seeded onto scaffolds of biodegradeable polymers and collagen. After another seven weeks of incubation, the "neo-bladders were reimplanted back into the patients. These newly grown bladders were implanted autologously into seven children with spinal bifida and malfunctioning bladders. Improved urinary controls were found in all seven patients receiving the implants with a mean follow-up of 4 years. The results are reported in this weeks issue of Lancet (premium content). This new finding is being highlighted by the mainstream media as an alternative to embryonic stem cell research. (One wonders whether the first one week incubation step described above was necessary for cellular expansion of adult stem cells from the biopsied tissue prior to seeding on a scaffold.)

April 3, 2006---A private biotech company, Prime Gen, based in Irvine, California also announced they have found highly plastic cells from the testes, similar to the findings by a German research group (see March 24 posting below). The US company researchers claim they have "reprogrammed" their cells from the testes and differentiated them into nerve, heart, and bone.

March 31, 2006---British scientists discover new cancer-associated gene involved in metastasis. They reported the S100P gene as a new "metastagene" which encodes for one of the S100 family of proteins. Other proteins reported to be metastatic are S100A4, osteopontin, and AGR2. Of particular interest is that S100P is expressed in cells within many normal tissues such as placenta, spleen, colon, prostate, lung and heart. (Intriguing finding which may suggest that gene expression may also occur in normal adult stem cells.)

March 30, 2006---Canadian research group reports in the March 29th issue of J. Neuroscience functional recovery in spinal-injured rats, when implanted with brain-derived neural progenitor cells. In a spinal cord injury rat model, neural precursor cells from transgenic animals labeled with yellow fluorescent protein were implanted into the damage site 2 to 8 weeks post trauma. Engraftment of labeled cells resulting in remyelination along with restoration of limb function (improvement in the BBB score) was enhanced by co-administration of growth factors, minocycline (anti-inflammatory agent), and cyclosporine A.

March 30, 2006---For those scientists who have been slaving all day in front of their tissue culture hoods and then, finding out that their cultures are either turning yellow due to bacterial contamination or the cells are starting to look funky after 4-5 passages (you're probably cultivating progenitor cells), don't fret, help is on the way. Take a break and visit National Lampoon's Website. They are doing a series of spoofs on stem cells. It should brighten your day after you visit their site.

March 24, 2006---An UPI press release noted that German scientists reported in Nature (to be published online this week) that they had identified embryonic-like stem cells in the testicles of adult mice. These cells were shown to be multipotent in their ability to differentiate into tissue phenotypes of all three germ layers of the body. Without seeing the data, it appears the German scientists' results may support our data, where we demonstrated BLSCs residing in rat testes. The below photomicrograph displays Moraga's BLSCs (Very dark brown staining cells) lining the semineferous tubules of the testicle.

Because they were also found around the semineferous tubules, the German scientists labeled their cells as “spermatogonial” (can’t wait to see the data and cells with more updates and commentary to follow).