Effect of microvascular endothelial cells on proliferation and apoptosis of osteoblast under condition of transwell chamber coculture

  Background  Bone capillaries play important roles in the maintenance of bone homeostasis and the phenomenon of “angiogenesis-osteogenesis coupling” has become the focus of clinicians. Revealing the mutual regulation relationship between microvascular endothelial cells and osteoblasts will provide new ideas for the clinical treatment of bone-related diseases.

  Objective  To investigate the effect of microvascular endothelial cells bEnd.3 on proliferation and apoptosis of osteoblast MC3T3-E1 cells under coculture conditions.

  Methods  The MC3T3-E1/bEnd.3 coculture system was established using transwell chambers. The cells were divided into control group (MC3T3-E1 cells were cultured separately) and coculture groups (MC3T3-E1 cells and bEnd.3 cells were co-incubated). CCK-8 assay was used to detect the changes in cell viability of MC3T3-E1 in the two groups. PI single-staining flow cytometry was applied to detect the changes in cell cycle of the two groups in MC3T3-E1, and Annexin V-FITC/PI double-staining flow cytometry for the changes of apoptosis rate of MC3T3-E1 in two groups. Western blotting was used to detect the expression changes of proliferation and apoptosis related-proteins (PCNA, Bax and Cleaved Caspase-3) in two groups of MC3T3-E1.

  Results  Compared with MC3T3-E1 cells cultured alone, the viability of MC3T3-E1 cells in coculture group enhanced significantly (P < 0.05), the ratio of S phase cells and G₂/M phase cells increased significantly (P < 0.01, P < 0.05 respectively), the expression level of PCNA increased (P < 0.01), while the rate of apoptosis decreased (P < 0.05), Bax and Cleaved Caspase-3 expression were down regulated (P < 0.05).

  Conclusion  Under the condition of coculture in transwell chamber, bEnd.3 cells can significantly promote the proliferation of MC3T3-E1 cells and inhibit its apoptosis.