Effects of serum-free culture on apoptosis in intervertebral disc nucleus pulposus cells of rat and its primary mechanism

Objective To investigate the effects of serum-free culture on the apoptosis in intervertebral disc nucleus pulposus cells of rat and its primary mechanism. Methods The intervertebral disc nucleus pulposus cells of second generation from SD rats were randomly divided into the starvation group and the control group.The cells of the control group were culture in DMEM contain 10% serum while starvation group in Serum-free DMEM for 24 h.The cells survival were detected by CCK-8(Cell Counting Kit-8);The cells apoptosis was detected by TUNEL(terminal deoxynucleotidyl transferase mediated dUTP nick end labeling);The expression of p38MAPK protein and cleaved Caspase-3 protein was observed by immunocytochemistry;the changes of Bcl-2 mRNA,Caspase-3 mRNA and p38MAPK mRNA was measured by RT-PCR(reverse transcription-polymerase chain reaction). Results Compared with control group,the starvation group shows:①Cells survival was significantly decreased (P<0.01);②rate of cells apoptosis was significantly increased(P<0.01);③The expression of p38MAPK and cleaved Caspase-3 protein was significantly increased;④Bcl-2 mRNA was significantly decreased;Caspase-3 mRNA and p38MAPK mRNA was significantly increased(P<0.01). Conclusion Serum-free culture may lead to the apoptosis of intervertebral disc nucleus pulposus cells of rat and its mechanism is related to the down-regulation of Bcl-2 and activation of caspase 3 through the p38MAPK pathway.