ABSTRACT
BACKGROUND:Previous studies have found that miR-21 expression is increased during osteogenic differentiation of bone marrow mesenchymal stem cells, but the action and molecular mechanism of miR-21 are stil unclear. OBJECTIVE:To verify the target gene of miR-21, Spry1, and to explore the role of Spry1 in osteogenic differentiation of human bone marrow mesenchymal stem cells. METHODS:Luciferase report was used to verify Spry1 gene targeted by miR-21, and western blot assay was used to detect the expression of Spry1 in the osteogenesis of human bone marrow mesenchymal stem cells. Spry1 expression vector was established and transfected into human bone marrow mesenchymal stem cells. Osteogenesis ability of human bone marrow mesenchymal stem cells was analyzed after Spry1 high expression by alkaline phosphatase, alizarin red staining, RT-PCR and western blot. RESULTS AND CONCLUSION:Luciferase report suggested that Spry1 was a target gene of miR-21. The expression level of Spry1 was decreased in the osteogenesis of human bone marrow mesenchymal stem cells. Increasing expression of Spry1 could inhibit osteogenic differentiation of human bone marrow mesenchymal stem cells. These results indicate that Spry1 as a target gene of miR-21 negatively regulates osteogenic differentiation of human bone marrow mesenchymal stem cells, and plays an important role in bone formation process.
ABSTRACT
BACKGROUND:In recent years a large number of studies have suggested that bone marrow mesenchymal stem cells can ease hyperglycemia of diabetic rats, but the related mechanism is unclear and controversial. OBJECTIVE:To investigate the relevant mechanism of bone marrow mesenchymal stem cells on pancreas microenvironment in vivo in diabetic rats. METHODS:Bone marrow mesenchymal stem cells were transfected with enhanced green fluorescent protein (EGFP) and administered to diabetic rats via the subcapsular pancreas. Blood glucose levels were monitored. The expressions of the key genes in islet development in these EGFP positive pancreatic cells were analyzed by Real-time quantitative PCR at different times. EGFP and insulin double-positive cells were detected by immunofluorescence. Flow cytometry was performed to analyze cellcycle and DNA ploidy. RESULTS AND CONCLUSION:Blood glucose levels were effectively reduced after transplantation. The expressions of the key genes in islet development reached their own peak values at different times after transplantation:Nestin at week 1, Nkx 2.2 at week 3, Pax 4 and Ngn 3 at week 4, insulin and glucagon at week 12, PDX-1 at week 8 until week 12. The cells double-positive for EGFP and insulin cells were observed. In the pancreas, EGFP positive cells at S+G 2/M phase were significantly increased, and there were no polyploid and aneuploid cells. In pancreas microenvironment, the bone marrow mesenchymal stem cells transplanted into the diabetic pancreas can differentiate into isletβ-like cells under gene control, but not through the fusion with tissue cells.