Abstract
Objective To investigate the relationship between autophagy and the endplate chondrocytes degeneration with intermittent cyclic mechanical tension (ICMT) in vitro.Methods The primary lumbar endplate cartilage ceils of 10 SD rats were cultured and the P1 generation was selected to treat with ICMT (10％,0.5 Hz) for 0 h,3 h,12 h,24 h,48 h.The cell morphology changes were observed through inverted phase contrast microscope.The expression of cartilage marker gene type Ⅱ collagen,SOX-9 and proteoglycan expression were observed by RT-PCR.Beclin-1 and LC3 expression of end-plate chondrocytes were detected by RT-PCR and Western blotting.The monodansylcadaverine (MDC) staining was used to detect autophagy rate in the endplate chondrocytes.The cells activity was detected by MTT assay.Results After ICMT loading,the cells of 0 h group and 3 h group kept normal morphology,but the cells of 12 h group presented irregular shape.The cells of 24 b group and 48 h group tended to assume a spindle-shaped morphology.The expression of type Ⅱ collagen,proteoglycan and Sox-9 in 24 h group and 48 h group decreased obviously.The results of RT-PCR and Western blotting showed that autophagy-related genes LC3 and Beclin-1 expression in time dependent increasing.The results of MDC staining showed that the occurrence of autophagy were significantly increased in 24 h and 48 h groups after ICMT,but MTT analysis showed the cells viability was significantly decreased,and the cell viability was significantly decreased with the stimulation of 3-methyl-adenine.Conclusion After ICMT loading,the endplate chondrocytes will lose their phenotype gradually.The expression of autophagy-related genes LC3 and Beclin-1 are significantly increased,but the cell viability is significantly decreased.Inhibiting the autophagy can reduce the cell survival rate.Autophagy may participate in the endplate chondrocytes degeneration process induced by ICMT. Key words: Intervertebral disc degeneration; Chondrocytes; Autophagy
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