Project description:The generation of insulin-producing pancreatic cells from stem cells in vitro would provide an unprecedented cell source for drug discovery and cell transplantation therapy in diabetes. However, insulin-producing cells previously generated from human pluripotent stem cells (hPSC) lack many functional characteristics of bona fide β cells. Here we report a scalable differentiation protocol that can generate hundreds of millions of glucose-responsive β cells from hPSC in vitro. These stem cell derived cells (SC) express markers found in mature β cells, flux Ca2+ in response to glucose, package insulin into secretory granules and secrete quantities of insulin comparable to adult β cells in response to multiple sequential glucose challenges in vitro. Furthermore, these cells secrete human insulin into the serum of mice shortly after transplantation in a glucose-regulated manner, and transplantation of these cells ameliorates hyperglycemia in diabetic mice. Differentiated cells were sorted and processed for RNA isolation using the MARIS protocol published previously (PMID: 24516164.) Human embryonic stem cell (hESC) line HUES8 was differentiated into SC-beta cells. Two biological replicates were analyzed. Those data were normalized together with and compared to existing, previously published data from Hrvatin et al. ( (PMID: 24516164) from human islet -derived insulin+ cells, undifferentiated HUES8 hES cells, and insulin+ cells derived from HUES8 cells according to previously published protocols.
Project description:The expression profile of miRNAs in MSCs and differentiated Insulin-producing cells was examined using RNA-seq technology. The expression of 411 miRNAs was observed, which we classified into three groups according to expression levels in differentiated Insulin-producing cells relative to that in MSCs: group I contained 107 miRNAs with an increased level of expression, group II contained 250 miRNAs that showed no change in expression and group III contained 54 miRNAs with decreased levels of expression.
Project description:Induced pluripotent stem cells can be differentiated in vitro into insulin-producing β cells. In this study we unveil the transcriptional dynamics of the differentiation and explain the heterogeneity of the final cell product, in comparison with β cells from organ donors.
Project description:The remarkable differentiation capacity of pluripotent stem cells into any adult cell types have enabled researchers to model human embryonic development and disease process in dishes, as well as deriving specialized cells for replacing damaged tissues. Type 1 diabetes is a degenerative disease characterized by autoimmune destruction of the insulin-producing beta islet cells in the pancreas. Recent advances have led to the establishment of different methods to direct differentiation of human or mouse pluripotent stem cells toward beta cell lineages. However, existing strategies have not yet succeeded in generating fully functional beta cells in vitro. Thus, it remains a major challenge to identify novel regulators of beta cell differentiation and maturation, and the islet-specific genetic and epigenetic regulatory networks are logical targets. To obtain a comprehensive view of the microRNA expression pattern during in vitro directed differentiation of hPSC into pancreatic beta islet cells, we collected 16 samples of 6 stages of differentiated derivatives, 2 samples of human fetal pancreas and 5 samples of purified human beta islet cells for analysis. With these samples, we performed genome-wide microRNA expression profiling using the Illumina Human v2 MicroRNA Expression BeadChips (1,146 assays).
Project description:This is a mathematical model describing the hematopoietic lineages with leukemia lineages, as controlled by end-product negative feedback inhibition. Variables include hematopoietic stem cells, progenitor cells, terminally differentiated HSCs, leukemia stem cells, and terminally differentiated leukemia stem cells.
Project description:The rare SLC30A8 mutation encoding a truncating p.Arg138* variant (R138X) in zinc transporter 8 (ZnT8) is associated with a 65% reduced risk for type 2 diabetes. To determine whether ZnT8 is required for beta cell development and function, we derived human pluripotent stem cells carrying the R138X mutation and differentiated them into insulin-producing cells. We found that human pluripotent stem cells with homozygous or heterozygous R138X mutation and the null (KO) mutation have normal efficiency of differentiation towards insulin-producing cells, but these cells show diffuse granules that lack crystalline zinc-containing insulin granules. Insulin secretion is not compromised in vitro by KO or R138X mutations in human embryonic stem cell-derived beta cells (sc-beta cells). Likewise, the ability of sc-beta cells to secrete insulin and maintain glucose homeostasis after transplantation into mice was comparable across different genotypes. Interestingly, sc-beta cells with the ZnT8 KO mutation showed increased cytoplasmic zinc, and cells with either KO or R138X mutation were resistant to apoptosis when extracellular zinc was limiting. These findings are consistent with a protective role of zinc in cell death and with the protective role of zinc in T2D.
Project description:A Drosophila microRNA (dme-miR-14) is involved in regulating the levels of insulin-like peptides (ilps) from the neuronal insulin-producing cells (IPCs). This is crucial for regulation of fat content in adult flies. Finding the target of this microRNA is crucial for understanding the regulation of ilp gene expression. We used microarrays to identify genes that are upregulated specifically in the neuronal IPCs in the microRNA mutants.
Project description:We determined the global microRNA expression profiles of primary human gallbladder cells and genetically reprogrammed human gallbladder cells and compared with pancreatic beta cells to ascertain the degree of cellular transdifferentatiation of insulin-producing human gallbladder cells to become beta-like cells. First, we cultured patient-derived gallbladder cells and then we transduced these with beta cell transcription factors to reprogram gallbladder cells to become beta-like cells. We used a pan-islet surface monoclonal antibody to enrich for insulin-producing reprogrammed human gallbladder cells using FACS.