walnut male flower extract potentiate bone marrow mesanchymal stem cells into the beta cells, invivo and invitro

Introduction: Recently stem cells differentiation and transplantation has attracted many researchers. It has been shown that rat injured pancreatic extracts (RIPE) can promote the differentiation of rat bone marrow mesenchymal stem cells (BMMSCs) into βcells, therefore, in this study we aimed to observe the effect of walnut male flower extract (WMFE) in differentiation and insulin secretion in βcells.Methods: Three doses of 1.4%, 3% and 6% for walnut male flowers were used. Bone marrow mesenchymal stem cells from rat was extracted by flushing and cell surface markers were analyzed by flow cytometry. Pancreatic tissue extract was prepared by use of protease inhibitor, and immediately were kept at -70 ° C for later use. BMMSCs were treated for RIPE + several doses and combination of WMFE, for 14 days. After two weeks, insulin secretion was measured by ELISA assay and insulin granules, in cell culture, were stained by dithizone dye. Gene related to βcells were evaluated by RT-PCR. PDX1, was evaluated by both immunocytochemistry and western blot. βcells was determined by stereology and cell count assay invivo and invitro.Results: Insulin was released almost twice more (1.94 time) in the cells treated for RIPE+WMFE (1.4 mg %), (0.72 ± 0.01 μg/L) than the cells treated for RIPE, (0.037± 0.01 µg/L), (P<0.05). RT-PCR results confirmed gene related to βcells: insulin, pro ins., PC1, PC2, Nkx 6.1, Glut2, Sst, Ngn3 in RIPE, RIPE+WMFE1.4 mg % groups. PDX1 protein was detected in differentiated groups by Western blot and immunocytochemistry. The results for stereology and cell count showed 44±2.5 differentiated cells per mm2 can be seen in the group of cells treated with RIPE in comparison with differentiated cells 51±2.0 per mm2 were observed in the RIPE + WMFE (1.4 mg %). The statistical analysis of the groups showed a significant increase (~7%) in RIPE+ WMFE (1.4 mg %) group in comparison with RIPE only (P= 0.034).The differentiated βcells from rat MSCs, was sub capsular transplanted into the STZ rat kidney to determine insulin secretion invivo. The results showed differentiated βcells had a good function and glucose level and insulin secretion back to the normal state 8 days after transplantation. Conclusion: WMFE can promote differentiation of BMMSCs and increases insulin secretion from the differentiated βcells invitro and invivo. Our study provides a new strategy to induce an efficient and directional differentiation of BMMSCs cells into βcells.

کلیدواژه ها:

beta cells ، differentiation ، BMMSCs