Regulatory effect of CCN3 on proliferation of mouse embryonic fibroblasts and its mechanism / 南方医科大学学报

ABSTRACT

OBJECTIVE@#To investigate the role of NOV/CCN3 in regulating the proliferation of mesenchymal stem cells (MSCs) and its regulatory mechanism and assess the value of CCN3 as a proliferative factor in bone tissue engineering.@*METHODS@#Mouse embryonic fibroblasts (MEFs) were used as the MSC model, in which CCN3 expression was up-regulated and downregulated by transfection with the recombinant adenovirus vectors Ad-CCN3 and Ad-siCCN3, respectively. Flow cytometry was used to analyze the changes in cell cycle and apoptosis of the transfected cells. Western blotting was used to detect the expression levels of the proliferation indicators (PCNA, cyclin E, and cyclin B1) and the apoptosis indicators (Bax and Bcl-2) to assess the effect of modulation of CCN3 expression on MEF proliferation and apoptosis. CCN3 protein secretion by the cells was detected using ELISA. RT-qPCR and Western blotting were employed to analyze the changes in the expressions of Notch1, ligand DLL1, the downstream key proteins or genes (Hey1, P300, H3K9) and MAPK pathway-related proteins ERK1+2 and p-ERK1+2.@*RESULTS@#Flow cytometry showed that compared with the control cells, MEFs transfected with Ad-CCN3 exhibited significantly increased cell proliferation index (@*CONCLUSIONS@#CCN3 over-expression promotes the proliferation and inhibits apoptosis of MEFs possibly by inhibiting the classical Notch signaling pathway and activating the MAPK pathway