Production Of Insulin-producing Cells Research Articles

Production of insulin producing cells from cord blood mesenchymal stem cells and their potential in cell therapy

Introduction: Mesenchymal stem cells (MSCs) were described as adherent cells with a fibroblast-like appearance, have a great capacity for self-renewal while maintaining their multipotency and differentiation into multiple tissues in vivo and in vitro. Methods: MSCs were isolated from cord blood of Sudanese donors using Ficoll-Hypaque gradient density protocol, and differentiate into β- like cells using 3-step protocol. STZ induced diabetic rats were injected intraperitoneally with the differentiated islet β- like cells and blood glucose levels were monitored for seven days. Results: The adherent cell appeared round and sphere after one-week of incubation, and the fibroblast-like colony was strongly attached after three weeks of seeding. The phenotyping of cells showed positivity for CD13, and negativity for CD34, CD45 and HLADR. MSCs were induced into islet-like cells using a 3-step (15-days) protocol. The differentiated cells showed positive diathizone stain and positive imuno-reactivity to anti-human insulin antibody. Secretion of insulin by insulin-producing cells showed positive result with >3.4 u/ml scale reading in high glucose concentration medium. After one-week of transplantation the level of blood glucose was reduced from 410 to 225 mg/dl in the experimental rat. Conclusion: Human UCB-MSCs can be differentiated into insulin-secreting cells invitro, and are able to produce and secrete insulin in response to high glucose concentration in vivo and in vitro.
 Keywords: Cord blood, Mesenchymal stem cell, islets β-like cells

MafA Overexpression: A New Efficient Protocol for In Vitro Differentiation of Adipose-Derived Mesenchymal Stem Cells into Functional Insulin-Producing Cells.

Objective We proposed a novel differentiation method for the efficient differentiation of adipose-derived mesenchymal stem cells (ADMSCs) into functional insulin-producing cells (IPCs) based on MafA overexpression. Materials and Methods In this experimental study, a eukaryotic expression vector containing MafA [MafA/pcDNA3.1(+)] was constructed and purified. ADMSCs were differentiated into IPCs. ADMSCs were assigned in two groups including control (C), and the MafA overexpressed (MafA+) groups. The ADMSCs were transfected by MafA/pcDNA 3.1(+) at day 10 of the differentiation. Differentiated cells were analyzed for the expression of multiple β cell specific genes (Nkx2.2, Ngn3, Isl-1, Pdx1, MafA, Nkx6.1, and Insulin) using real-time polymerase chain reaction (PCR). The insulin secretion potency of the differentiated cells in response to glucose exposure was also determined using an enzyme-linked immunosorbent assay (ELISA) method and Dithizone (DTZ) staining. The IPCs from the control manipulated group, and un-differentiated ADMSCs group were transplanted to streptozotocin (STZ)-diabetic rats. Rats were monitored for blood glucose and insulin concentration. Results The results revealed that ADMSCs were successfully differentiated into IPCs through the 14 day differentiation protocol. The expression of β-cell specific genes in MafA+ IPCs was higher than in control cells. Glucose-induced insulin secretion after the exposure of IPCs to glucose was higher in MafA+ group than the control group. The STZ- diabetic rats showed an ability to secrete insulin and apparent hyperglycemic condition adjustment after transplantation of the control IPCs. The mean insulin concentration of diabetic rats that were transplanted by manipulated IPCs was significantly higher than ADMSCs-transplanted rats; however, no effect was observed in the concentration of blood glucose. ConclusionThe overexpression of MafA can be used as a novel promising approach for the efficient production of IPCs from ADMSCs in vitro. However, the future therapeutic use of the MafA+ IPCs in diabetic animals needs further investigations.

Encapsulated Islet Transplantation: Where Do We Stand?

Transplantation of pancreatic islets encapsulated within immuno-protective microcapsules is a strategy that has the potential to overcome graft rejection without the need for toxic immunosuppressive medication. However, despite promising preclinical studies, clinical trials using encapsulated islets have lacked long-term efficacy, and although generally considered clinically safe, have not been encouraging overall. One of the major factors limiting the long-term function of encapsulated islets is the host's immunological reaction to the transplanted graft which is often manifested as pericapsular fibrotic overgrowth (PFO). PFO forms a barrier on the capsule surface that prevents the ingress of oxygen and nutrients leading to islet cell starvation, hypoxia and death. The mechanism of PFO formation is still not elucidated fully and studies using a pig model have tried to understand the host immune response to empty alginate microcapsules. In this review, the varied strategies to overcome or reduce PFO are discussed, including alginate purification, altering microcapsule geometry, modifying alginate chemical composition, co-encapsulation with immunomodulatory cells, administration of pharmacological agents, and alternative transplantation sites. Nanoencapsulation technologies, such as conformal and layer-by-layer coating technologies, as well as nanofiber, thin-film nanoporous devices, and silicone based NanoGland devices are also addressed. Finally, this review outlines recent progress in imaging technologies to track encapsulated cells, as well as promising perspectives concerning the production of insulin-producing cells from stem cells for encapsulation.

Dachshund homologues play a conserved role in islet cell development

All metazoans use insulin to control energy metabolism, but they secrete it from different cells: neurons in the central nervous system in invertebrates and endocrine cells in the gut or pancreas in vertebrates. Despite their origins in different germ layers, all of these insulin-producing cells share common functional features and gene expression patterns. In this study, we tested the role in insulin-producing cells of the vertebrate homologues of Dachshund, a transcriptional regulator that marks the earliest committed progenitors of the neural insulin-producing cells in Drosophila. Both zebrafish and mice expressed a single dominant Dachshund homologue in the pancreatic endocrine lineage, and in both species loss of this homologue reduced the numbers of all islet cell types including the insulin-producing β-cells. In mice, Dach1 gene deletion left the pancreatic progenitor cells unaltered, but blocked the perinatal burst of proliferation of differentiated β-cells that normally generates most of the β-cell mass. In β-cells, Dach1 bound to the promoter of the cell cycle inhibitor p27Kip1, which constrains β-cell proliferation. Taken together, these data demonstrate a conserved role for Dachshund homologues in the production of insulin-producing cells.