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A Model for Human Islet Transplantation to Immunodeficient Streptozotocin-Induced Diabetic Mice.

ABSTRACT: Streptozotocin (STZ) is a cytotoxic glucose analogue that causes beta cell death and is widely used to induce experimental diabetes in rodents. The sensitivity of beta cells to STZ is species-specific and human beta cells are resistant to STZ. In experimental islet transplantation to rodents, STZ-diabetes must be induced before transplantation to avoid destruction of grafted islets by STZ. In human islet transplantation, injection of STZ before transplantation is inconvenient and costly, since human islet availability depends on organ donation and frail STZ-diabetic mice must be kept for unpredictable lapses of time until a human islet preparation is available. Based on the high resistance of human beta cells to STZ, we have tested a new model for STZ-diabetes induction in which STZ is injected after human islet transplantation. Human and mouse islets were transplanted under the kidney capsule of athymic nude mice, and 10-14 days after transplantation mice were intraperitoneally injected with five consecutive daily doses of STZ or vehicle. Beta-cell death increased and beta-cell mass was reduced in mouse islet grafts after STZ injection. In contrast, in human islet grafts beta cell death and mass did not change after STZ injection. Mice transplanted with rodent islets developed hyperglycemia after STZ-injection. Mice transplanted with human islets remained normoglycemic and developed hyperglycemia when the graft was harvested. STZ had no detectable toxic effects on beta cell death, mass and function of human transplanted islets. We provide a new, more convenient and cost-saving model for human islet transplantation to STZ-diabetic recipients in which STZ is injected after islet transplantation.

SUBMITTER: Estil Les E

PROVIDER: S-EPMC6299193 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature

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A Model for Human Islet Transplantation to Immunodeficient Streptozotocin-Induced Diabetic Mice.

Estil Les Elisabet E Téllez Noèlia N Nacher Montserrat M Montanya Eduard E

Cell transplantation 20181001 11

Streptozotocin (STZ) is a cytotoxic glucose analogue that causes beta cell death and is widely used to induce experimental diabetes in rodents. The sensitivity of beta cells to STZ is species-specific and human beta cells are resistant to STZ. In experimental islet transplantation to rodents, STZ-diabetes must be induced before transplantation to avoid destruction of grafted islets by STZ. In human islet transplantation, injection of STZ before transplantation is inconvenient and costly, since h ...[more]

PMID: 30269527

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Project description:BACKGROUND:Dental pulp stem cells (DPSCs) have high proliferation and multi-differentiation capabilities that maintain their functionality after cryopreservation. In our previous study, we demonstrated that cryopreserved rat DPSCs improved diabetic polyneuropathy and that the efficacy of cryopreserved rat DPSCs was equivalent to that of freshly isolated rat DPSCs. The present study was conducted to evaluate whether transplantation of cryopreserved human DPSCs (hDPSCs) is also effective for the treatment of diabetic polyneuropathy. METHODS:hDPSCs were isolated from human impacted third molars being extracted for orthodontic reasons. Eight weeks after the induction of diabetes in nude mice, hDPSCs (1?×?105/limb) were unilaterally transplanted into the hindlimb skeletal muscle, and vehicle (saline) was injected into the opposite side as a control. The effects of hDPSCs were analyzed at 4?weeks after transplantation. RESULTS:hDPSC transplantation significantly ameliorated reduced sensory perception thresholds, delayed nerve conduction velocity, and decreased the blood flow to the sciatic nerve in diabetic mice 4?weeks post-transplantation. Cultured hDPSCs secreted the vascular endothelial growth factor (VEGF) and nerve growth factor (NGF) proteins. A subset of the transplanted hDPSCs was localized around the muscle bundles and expressed the human VEGF and NGF genes at the transplanted site. The capillary/muscle bundle ratio was significantly increased on the hDPSC-transplanted side of the gastrocnemius muscles in diabetic mice. Neutralizing antibodies against VEGF and NGF negated the effects of hDPSC transplantation on the nerve conduction velocity in diabetic mice, suggesting that VEGF and NGF may play roles in the effects of hDPSC transplantation on diabetic polyneuropathy. CONCLUSIONS:These results suggest that stem cell transplantation with hDPSCs may be efficacious in treating diabetic polyneuropathy via the angiogenic and neurotrophic mechanisms of hDPSC-secreted factors.

Project description:BackgroundDiabetic nephropathy (DN) is a severe complication of diabetes mellitus (DM). Pancreas or islet transplantation has been reported to prevent the development of DN lesions and ameliorate or reverse existing glomerular lesions in animal models. Shortage of pancreas donor is a severe problem. Islets derived from stem cells may offer a potential solution to this problem.ObjectiveTo evaluate the effect of stem cell-derived islet transplantation on DN in a rat model of streptozotocin-induced DM.MethodsPancreatic progenitor cells were isolated from aborted fetuses of 8 weeks of gestation. And islets were prepared by suspension culture after a differentiation of progenitor cells in medium containing glucagon-like peptide-1 (Glp-1) and nicotinamide. Then islets were transplanted into the liver of diabetic rats via portal vein. Blood glucose, urinary volume, 24 h urinary protein and urinary albumin were measured once biweekly for 16 weeks. Graft survival was evaluated by monitoring human C-peptide level in rat sera and by immunohistochemical staining for human mitochondrial antigen and human C-peptide in liver tissue. The effect of progenitor-derived islets on filtration membrane was examined by electron microscopy and real-time polymerase chain reaction (PCR). Immunohistochemical staining, real-time PCR and western blot were employed for detecting fibronectin, protein kinase C beta (PKCβ), protein kinase A (PKA), inducible nitric oxide synthase (iNOS) and superoxide dismutase (SOD).ResultsIslet-like clusters derived from 8th gestational-week human fetal pancreatic progenitors survived in rat liver. And elevated serum level of human C-peptide was detected. Blood glucose, 24 h urinary protein and urinary albumin were lower in progenitor cell group than those in DN or insulin treatment group. Glomerular basement membrane thickness and fibronectin accumulation decreased significantly while podocytes improved morphologically in progenitor cell group. Furthermore, receptor of advanced glycation end products and PKCβ became down-regulated whereas PKA up-regulated by progenitor cell-derived islets. And iNOS rose while SOD declined.ConclusionsDN may be reversed by transplantation of human fetal pancreatic progenitor cell-derived islets. And fetal pancreatic progenitor cells offer potential resources for cell replacement therapy.

Project description:In clinical and experimental human pancreatic islet transplantations, establishing pretransplant assessments that accurately predict transplantation outcomes is crucial. Conventional in vitro viability assessment that relies on manual counting of viable islets is a routine pretransplant assessment. However, this method does not correlate with transplantation outcomes; to improve the method, we recently introduced a semi-automated method using imaging software to objectively determine area-based viability. The goal of the present study was to correlate semi-automated viability assessment with posttransplantation outcomes of human islet transplantations in diabetic immunodeficient mice, the gold standard for in vivo functional assessment of isolated human islets. We collected data from 61 human islet isolations and 188 subsequent in vivo mouse transplantations. We assessed islet viability by fluorescein diacetate and propidium iodide staining using both the conventional and semi-automated method. Transplantations of 1,200 islet equivalents under the kidney capsule were performed in streptozotocin-induced diabetic immunodeficient mice. Among the pretransplant variables, including donor factors and post-isolation assessments, viability measured using the semi-automated method demonstrated a strong influence on in vivo islet transplantation outcomes in multivariate analysis. We calculated an optimized cutoff value (96.1%) for viability measured using the semi-automated method and showed a significant difference in diabetes reversal rate for islets with viability above this cutoff (77% reversal) vs. below this cutoff (49% reversal). We performed a detailed analysis to show that both the objective measurement and the improved area-based scoring system, which distinguished between small and large islets, were key features of the semi-automated method that allowed for precise evaluation of viability. Taken together, our results suggest that semi-automated viability assessment offers a promising alternative pretransplant assessment over conventional manual assessment to predict human islet transplantation outcomes.

Project description:BACKGROUND:Adipose-derived mesenchymal stem cells (ASCs) therapy is emerging as a novel therapeutic option for the treatment of a variety of diseases including diabetes and diabetic wound healing. Multiple studies indicate that ASCs could promote wound healing and reverse diabetes. However, whether ASCs from diabetic donors retain their therapeutic functions and the mechanisms of how ASCs contribute to wound healing remain largely unknown. In this study, we explored the cutaneous wound healing ability of ASCs collected from C57BL/6 mice that had been rendered diabetic with streptozotocin (STZ). METHODS:ASCs were harvested from adipose tissues of type 1 diabetic (T1D) or normal C57BL/6 mice. Cell phenotypes were evaluated by flow cytometry analysis, and cell differentiation into adipocytes, chondrocytes, and osteocytes was compared. Secretions of transforming growth factor ? (TGF-?1), basic fibroblast growth factor (bFGF), and vascular endothelial growth factor (VEGF) by ASCs were assessed by ELISA. Migration and proliferation of fibroblasts co-cultured with T1D ASCs or control ASCs were also compared. The therapeutic effects of T1D and control ASCs in promoting wound closure were measured in vivo in a T1D wound mouse model. Granulation tissues were collected and stained with H&E at 14th day. CD34 and collagen I were detected by immunohistochemistry. Expressions of IL-6, ?-SMA, CD31, collagen I, and collagen III were quantified by real-time PCR. GFP-expressing ASCs were used to trace in vivo cell differentiation. RESULTS:T1D ASCs and control ASCs showed similar expression of cell surface markers (CD29, CD34, CD105) and proliferation pattern. They can both differentiate into different cell types. T1D ASCs secreted similar amounts of VEGF and bFGF, but less TGF-? compared with control ASCs. Like control ASCs, T1D ASCs promoted the proliferation and migration of skin fibroblast cells. When injected in cutaneous wound of T1D mice, T1D ASCs increased wound closure and hair follicle regeneration at a comparable extent as ASCs. Mice receiving T1D ASCs or ASCs exhibited significantly higher expressions of collagen I, collagen III, and CD31 and reduced expression of IL-6 in wound tissues. Immunohistochemistry staining showed increased angiogenesis in mice receiving ASCs as was evident by increased CD34+ cells and collagen I staining. GFP+ ASCs injection showed that ASCs differentiated into fibroblasts and endothelial cells in vivo. CONCLUSIONS:Our results suggest that T1D ASCs could accelerate cutaneous wound healing. Mechanisms may include increasing fibroblast growth and migration, skin angiogenesis, and differentiation into fibroblasts and endothelial cells. This study provides evidence that diabetic ASCs may be used as a therapeutic option in cutaneous wound healing in diabetic recipients.

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A Model for Human Islet Transplantation to Immunodeficient Streptozotocin-Induced Diabetic Mice.

Publications

A Model for Human Islet Transplantation to Immunodeficient Streptozotocin-Induced Diabetic Mice.

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