The aim of this study was to provide evidence for further in vivo maturation of insulin-producing cells (IPCs) derived from human bone marrow-derived mesenchymal stem cells (HBM-MSCs). exact mechanism(s) involved requires further investigation. 1. Introduction In the year 2000, it was estimated that 150 million people were affected buy Ibotenic Acid by DM, and this number is usually expected to double in Sirt4 2025 . For type 1 diabetes, maintenance of appropriate glycemic control using exogenous insulin is usually possible but imposes a burden on patients. Transplantation of an intact pancreas as well as isolated pancreatic islets is usually ideal alternative. However, the shortage of cadaveric organs and the need for immunosuppression are limiting factors . Type 2 diabetes can be treated initially using oral medications, but eventually 27% of patients become insulin-dependent. Of these, less than half achieve the recommended HB A1C level . Recent progress in the field of regenerative therapies provides the potential for the generation of surrogate values were corrected using Bonferroni adjustments. A value of <0.05 was considered significant. The mean values were used as a measure of variance. The median values were utilized only if there were extreme observations. 3. Results 3.1. Characterization of the Cultured HBM-MSCs At the end of expansion, the cultured cells became spindle-shaped, fibroblast-like cells that arranged in monolayers. Flow cytometry revealed that these cells expressed high levels of CD73, CD90, and CD105 but negligible levels of CD14, CD34, and CD45 (Supplementary Table 1 in Supplementary Material, available online at http://dx.doi.org/10.1155/2015/575837). These cells could be differentiated to form adipocytes, chondrocytes, and osteocytes when the appropriate growth factors were added (Supplementary Physique 1). Accordingly, evidence for their multilineage potential was confirmed. 3.2. Functional Evaluation of Differentiated HBM-MSCs At the end of differentiation, flow cytometric analysis indicated that the percentage of generated IPCs was meager, ranging between 0.12% and 3.4%. The presence of insulin granules within the cytoplasm of the IPCs was detected via immunocytochemistry. Immunostaining for c-peptide was also positive in the IPCs. Coexpression of insulin and c-peptide within the same cells was detected via electronic merging (Physique 1). GCG staining was detected in some of the examined samples. However, positive staining for SST was not detected. Physique 1 Immunofluorescence staining of differentiated HBM-MSCs ((a) selected field). (a) Positive staining for intracytoplasmic insulin granules (green) with counterstaining for DAPI (blue). (w) Positive staining for c-peptide (red) with counterstaining for DAPI ... 3.3. Outcomes of the In Vivo Transplantation Experiments Out of the 29 transplanted animals, 5 mice did not tolerate the surgical procedure. The blood glucose levels of the surviving animals became normalized within a few days after transplantation (4 1.6 days). Thereafter, the animals remained euglycemic throughout the observation period. The serum levels of human insulin and human c-peptide were measurable one week after transplantation, and these values also remained unchanged throughout the observation period. Serum levels of mouse insulin became negligible after induction of diabetes (Table 1). The results of the oral glucose tolerance test were normal. The corresponding c-peptide level measurements indicated parallel changes, providing evidence that the transplanted cells were glucose-responsive and insulin-secreting (Physique 2, Supplementary Table 2). Physique 2 The oral glucose tolerance test performed 4 weeks after transplantation. (a) The blood glucose levels 2, 4, and 12 weeks after transplantation displayed normal patterns. (w) The corresponding serum human c-peptide values exhibited a comparable pattern. ... Table buy Ibotenic Acid 1 Mean blood glucose and serum human insulin, c-peptide, and serum mouse insulin levels in mice transplanted with HBM-MSCs (human bone marrow-derived mesenchymal stem cells). Immunohistochemistry of the HBM-MSC-bearing kidneys buy Ibotenic Acid revealed that the percentage of IPCs increased gradually, peaking at 4 weeks after transplantation (~18%) without any substantial change thereafter (Physique 3, Supplementary Table 3). Again, the coexpression of insulin and c-peptide within the cytoplasm of these cells was confirmed. Positive staining for GCG and SST was detected in some cells that did not display.