[Semaphorin 3A-stimulated bone marrow mesenchymal stem cells sheets promotes osteogenesis of type 2 diabetic rat].

To evaluate the effect of semaphorin 3A (Sema3A) pre-treated bone marrow mesenchymal stem cells (BMSC) sheets on new bone formation in type 2 diabetes mellitus rats. Type 2 diabetes mellitus (T2DM) were induced by injection of streptozotocin, and the BMSC were isolated, controlled, identified and induced into cell sheets. Fifteen T2DM rats were randomly divided into control, sheets and Sema3A-sheets group and the calvarial critical size defect (CSD) model of rats were established. The defect zone of rats from control group were implanted with bone powder. The defect zone of rats from sheets group were implanted with bone powder and BMSC sheets. The defect zone of rats from Sema3A-sheets group were implanted with bone powder and BMSC sheets pretreated with 1.0 mg/L Sema3A. After 8 weeks, the bone samples were harvested and analyzed by micro-CT scanning, HE staining for the evaluation of new bone formation, and the immunohistochemical analysis for the expression of osteogenesis-related proteins including type Ⅰ collagen (COL- Ⅰ ), bone morphogenetic protein-2 (BMP-2), and osteocalcin (OCN). The BMSC were isolated and cultured, and oil red O and Alizarin red S staining proved the multi-potential differentiation. Eight weeks after the establishment of calvarial CSD model, Sema3A-sheet group showed the most abundant new bone formation (0.516±0.070), with increased bone volume fraction, namely bone volume/tissue volume (BV/TV) compared with sheets group (0.319±0.050) and control group (0.224±0.037) (<0.05), and the sheets group showed increased BV/TV compared with control group (<0.05). While trabecular thickness (Tb.Th) control group showed no difference in three groups (>0.05). HE staining also confirmed that Sema3A-sheets group showed the most new bone formation. Sheet group (0.174±0.051) compared showed difference with control group (0.099±0.033) (< 0.05), and Sema3A-sheet group (0.421±0.069) showed increased bone formation compared with sheet group and control group (<0.05). Immunohistochemistry showed that BMSC sheet increased the expression of osteogenesis-related proteins including COL-Ⅰ, BMP-2 and OCN, while Sema3A pretreatment showed more obvious increase of the expression of COL-Ⅰ and OCN. The combined implantation of bone powder and Sema3A stimulated BMSC sheets significantly increased bone regeneration . Therefore, Sema3A pre-treated BMSC sheets transplantation provides a new strategy for restoring bone defect in T2DM.