Stem cells to replace or regenerate the diabetic pancreas: Huge potential & existing hurdles.

Various stem cell sources are being explored to treat diabetes since the proof-of-concept for cell therapy was laid down by transplanting cadaveric islets as a part of Edmonton protocol in 2000. Human embryonic stem (hES) cells derived pancreatic progenitors have got US-FDA approval to be used in clinical trials to treat type 1 diabetes mellitus (T1DM). However, these progenitors more closely resemble their foetal counterparts and thus whether they will provide long-term regeneration of adult human pancreas remains to be demonstrated. In addition to lifestyle changes and administration of insulin sensitizers, regeneration of islets from endogenous pancreatic stem cells may benefit T2DM patients. The true identity of pancreatic stem cells, whether these exist or not, whether regeneration involves reduplication of existing islets or ductal epithelial cells transdifferentiate, remains a highly controversial area. We have recently demonstrated that a novel population of very small embryonic-like stem cells (VSELs) is involved during regeneration of adult mouse pancreas after partial-pancreatectomy. VSELs (pluripotent stem cells in adult organs) should be appreciated as an alternative for regenerative medicine as these are autologous (thus immune rejection issues do not exist) with no associated risk of teratoma formation. T2DM is a result of VSELs dysfunction with age and uncontrolled proliferation of VSELs possibly results in pancreatic cancer. Extensive brainstorming and financial support are required to exploit the potential of endogenous VSELs to regenerate the pancreas in a patient with diabetes.

Stem Cells Cloning and Therapeutic Potential.

Stem cells have huge potential to transform current manner in which medicine is practiced. Rather than treating diseased cells with medicines and antibiotics, stem cells can just replace the diseased cells with healthy cells. But it will take time before this research gets translated to the clinic. At present, various types of stem cells like human embryonic stem (hES) cells, induced pluripotent stem (iPS) cells, fetal stem cells, adult tissue-specific stem cells (HSCs, MSCs, etc.), very small embryonic-like stem cells (VSELs) and related technologies like therapeutic cloning are subject to extensive research. Clinicians appear to be in a hurry to apply the stem cells to their patients and there is a huge industry banking stem cells for future autologus use. However, the scientifc community is still not sure which is the best stem cell candidate for regenerative medicine. The chapter provides an update on various fronts and also discusses whether there exists a need to bank stem cells for future use. The author is puzzled by realizing as to what needs to be repaired/ regenerated-the stem cells or their microenvironment ‘niche’!

Stem Cells in Drug Screening for Neurodegenerative Disease

Because the average human life span has recently increased, the number of patients who are diagnosed with neurodegenerative diseases has escalated. Recent advances in stem cell research have given us access to unlimited numbers of multi-potent or pluripotent cells for screening for new drugs for neurodegenerative diseases. Neural stem cells (NSCs) are a good model with which to screen effective drugs that increase neurogenesis. Recent technologies for human embryonic stem cells (ESCs) or induced pluripotent stem cells (iPSCs) can provide human cells that harbour specific neurodegenerative disease. This article discusses the use of NSCs, ESCs and iPSCs for neurodegenerative drug screening and toxicity evaluation. In addition, we introduce drugs or natural products that are recently identified to affect the stem cell fate to generate neurons or glia.

Neuronal Differentiation of a Human Induced Pluripotent Stem Cell Line (FS-1) Derived from Newborn Foreskin Fibroblasts

Isolation of induced pluripotent stem cells (iPSCs) from fully differentiated somatic cells has revolutionized existing concepts of cell differentiation and stem cells. Importantly, iPSCs generated from somatic cells of patients can be used to model different types of human diseases. They may also serve as autologous cell sources that can be used in transplantation therapy. In this study, we investigated the neuronal properties of an iPSC line that is derived from human neonatal foreskin fibroblasts (FS-1). We initially examined the morphology and marker expression of FS-1 cells at undifferentiated stage. We then spontaneously differentiated FS-1 cells in suspension culture and examined the expression of markers representing three germ layers. We finally differentiated FS-1 cells into neuronal lineages by co-culturing them with PA6 stromal cells, and found that, under the conditions we used, they have a tendency to differentiate into more forebrain-type neurons, suggesting that FS-1 iPSC-derived neural cells will be useful to be used in cell therapy of stroke or Huntington's disease, among others. Taken together, FS-1 cells derived from human neonatal fibroblasts exhibit very similar properties with human ES cells, and can provide useful sources for cell therapy and various other applications.

Argonaute-2-null embryonic stem cells are retarded in self-renewal and differentiation.

RNA interference (RNAi) pathways regulate self-renewal and differentiation of embryonic stem (ES) cells. Argonaute 2 (Ago2) is a vital component of RNA-induced silencing complex (RISC) and the only Ago protein with slicer activity. We generated Ago2-deficient ES cells by conditional gene targeting. Ago2-deficient ES cells are defective in the small-RNA-mediated gene silencing and are significantly compromised in biogenesis of mature microRNA. The self-renewal rate of Ago2-deficient ES cells is affected due to failure of silencing of Cdkn1a by EScell- specific microRNAs (miRNA) in the absence of Ago2. Interestingly, unlike Dicer- and Dgcr8-deficient ES cells, they differentiate to all three germ layers both in vivo and in vitro. However, early differentiation of Ago2-deficient ES cells is delayed by 2–4 days as indicated by persistence of higher levels of self-renewal/ pluripotency markers during differentiation. Further, appearance of morphological and differentiation markers is also delayed during the differentiation. In this study we show that Ago2 is essential for normal self-renewal and differentiation. Also, our data suggest that self-renewal and differentiation of ES cells are regulated by both siRNA and miRNA pathways.

Influence of dermal analog composed with MSCs upon the differentiation potency of ES cell-derived epidermal stem cells / 基础医学与临床

Objective To explore the differentiation potency of ES cell-derived epidermal stem cells compounded by dermal analogs reconstructed with bone MSCs in hypodermis.Methods The dermal analogs were reconstructed with rat bone MSCs and compound gel-gelatin sponge.E14-ES cells,labeled with Hoechst 33342,were cocultured with human amnion.Four days later,epidermal stem cell clones were formed.The skin analogs,reconstructed with ES cell-derived epidermal stem cells and dermal analogs,were transplanted into 129 mice hypodermis.The differentiation tissue of skin analogs was sampled at 2,4,6,8 weeks.The sections were observed with HE staining,immunohistochemical and di-labeled immunofluorescence methods to test the expression of ?1 integrin,CK15,CK19,CEA,CK18.Results The sections were showed tubular or follicular like structures formed with simple or stratified epithelium at 2 and 4 weeks.Keratinized stratified squamous epithelium,sweat glands-like,sebaceous glands-like and hair follicles-like structures were observed at 6 week and 8 week after transplantation.The cells labeled by Hoechst 33342,formed tubular or follicular like structures,expressed?1 integrin,CK15,CK19,CEA and CK18positive respectively at 2 and 4 weeks.The sweat glands-like structure expressed CEA and CK18 positive respectively at 6 and 8 weeks.There were more sebaceous glands-like structures.Conclusion ES cell-derived epidermal stem cells compounded by dermal analogs reconstructed with bone MSCs can differentiate into keratinized stratified squamous epithelium,sweat glands-like,sebaceous glands-like and hair follicles-like structures in hypodermis.

Establishment of Embryonic Stem Cell Line from Isolated Blastomeres from Mouse Preimplantation Embryos / 대한불임학회지

OBJECTIVE: The aim of this study was to investigate whether embryonic stem (ES) cells can be established from isolated blastomeres of mouse embryos. METHODS: Blastomeres were separated from mouse (C57Bl/6J) 2- or 4-cell embryos. Isolated blastomeres or whole 4-cell embryos were co-cultured with mitosis-arrested STO feeder cells in DMEM supplemented with recombinant murine leukemia inhibitory factor and ES-qualified fetal bovine serum. After the tentative ES cell lines were maintained from isolated blastomeres or whole embryos, some of them were frozen and the others were sub-cultured continually. Characteristics of tentative ES cell lines as were evaluated for specific gene expressions with immunocytochemistry and RT-PCR. RESULTS: One ES cell line (3.0%) was established from isolated blastomere of 2-cell embryo and one cell line (4.0%) from isolated two blastomeres of 4-cell embryo. And five cell lines (16.7%) were established from whole 4-cell embryos. Both cell lines from isolated blastomere and whole embryo expressed mouse ES cells specific markers such as SSEA-1, Oct-4 and alkaline phosphatase. Marker genes of three germ layers were expressed from embryoid bodies of both cell lines. CONCLUSION: This study suggests that mouse ES cells could be established from isolated blastomeres, although the efficiency is lower than whole embryos. This animal model could be applied to establishment of autologous human ES cells from biopsied blastomeres of preimplantation embryos in human IVF-ET program.

Isolation and Culture of Human Embryonic Stem-like Cells from Abnormal Blastocysts / 대한불임학회지

No abstract available.

STUDY ON THE Brn-3a COMMITTED INDUCTION OF EMBRYONIC STEM CELLS INTO NEURONS / 解剖学报

Objective To investigate if the transcription factor Brn 3a can induce committedly ES cells into neurons. Methods pJ5 Brn 3a recombinant expression factors containing objective gene Brn 3a were introduced into mouse ES D3 cells by liposome mediated transfection. The Brn 3a transfected ES cells were cultured to undergo differentiation into neurons. Finally immunohistochemical technique was implied to detect the expression of Brn 3a before and after transfection and some specific marked antigens of mature neural cell and glia cell were detected after the ES cells differentiation into neuron like cells. Results 1 Before transfection ES cells do not express Brn 3a and the Brn 3a immunoreaction is negative. After transfection of Brn 3a gene 24?h the ES cells could be detected to have Brn 3a and the immunoreaction is positive.2 When this Brn 3a transfected ES cells were cultured whithin 1 week over 70% ES cells could differentiate into neuron like cells bearing obvious neurites. These neuron like cells express NF H+L 、 NSE、SY and do not express GFAP.Conclusion Brn 3a transcription factor can successfully induce ES cells to differentiate into neuron like cells. [

PRIMARY STUDY ON THE DIFFERENTIATION POTENTIAL OF ESCELL DERIVED EPIDERMAL LIKE STEM CELLS / 解剖学报

Objective To investigate the differentiation potenitial of ES cell derived epidermal like stem cells,to lay a base for the study of their differentiation mechanism,as well as seek new source to provide seed cells for skin engineering. Methods Mouse ES cells labeled or unlabeled by Hoechst 33342 were cocultrued with human amnion for 4 days.The epidermal like stem cell clones formed on the surface of amnion were digested with trypsin and transplanted into hypodermis of nude mice for 10,20 and 45 days,then the differentiation pattern of the donor cells were observed and estimated with morphological and immunohistochemical method. Results The grafts may differentiate into tubular or follicular like structures lined with simple or stratified epithelial like cells which expressed ? 1 integin,CK19,CK15,PK involucrin and CEA respectively after 10 to 30 days of transplantation.Keratinized stratified squamous epithelium,sebaceous gland like,sweat gland like and hair follicle like structures were observed after 45 days ofter transplantation.Conclusion ES cell derived epidermal like cells might have differentation potential to diffreentiate into keratinized stratified squamous epithelium,sebaceous like,sweat gland like and hair follicle like structures.