Effect of Bushen Huoxue Fang(BSHXF)on the expression of circadian rhythm molecules BMAL1 and CLOCK in endplate chondrocytes under mechanical loading
Objective To observe the role of BSHXF in regulating critical molecules in the circadian rhythms of mechanical load-degenerated endplate chondrocytes(CEPCs).Methods Dexamethasone synchronization treatment was used to detect the changes in the circadian rhythms of CEPCs over 48 h;different mechanical load conditions(5％or 10％draft force)were applied to explore the effects of mechanical load on the expression of rhythmic molecules in CEPCs.Control group,abnormal mechanical load group,intervention group of BHHXF and BHHXF control group were set up.The qRT-PCR was used to detect the expression of crucial rhythm molecules of CEPCs,brain and muscle Arnt-like 1(BMAL1)and circadian locomotor output cycles kaput(CLOCK),as well as the anabolic marker Aggrecan;apoptosis of cells in each group was detected by flow cytometry.Results CEPCs showed periodic rhythmic changes.Physiological loading caused elevated quantification of the phase,period,and amplitude of CEPCs'circadian rhythms.Abnormal mechanical loading(10％draft force,1 Hz,24 h)resulted in decreased expression of BMAL1 and CLOCK(P＜0.01),reduced expression of the Aggrecan(P＜0.01),and increased apoptosis(P＜0.05).The drug-containing serum of BSHXF elevated the expression of BMAL1,CLOCK,and Aggrecan(P＜0.05)and decreased apoptosis of CEPCs(P＜0.01)after mechanical loading regression.Conclusion BSHXF can effectively delay the mechanical load degeneration of CEPCs,and its mechanism of action is related to the regulation of BMAL1 and CLOCK.
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