Defective autophagy in vascular smooth muscle cells (VSMCs) in response to oxidative stress can lead to cellular apoptosis and plaque instability. Previous studies have revealed that the circadian clock system is involved in autophagic regulation and plaque progression. However, the mechanism by which circadian rhythmicity influences VSMC autophagy and plaque stability remains unclear. Our study described the circadian profiles in atheromatous plaques and verified the role of circadian misalignment in VSMC autophagy and apoptosis. We found that the mRNA expression levels of circadian locomotor output cycles protein kaput (CLOCK) and Beclin 1 were significantly decreased in unstable plaques compared with stable plaques. No significant differences were observed in other circadian rhythm genes. VSMCs treated with oxidized low-density lipoprotein (ox-LDL, 80 μg/ml) exhibited abnormal circadian rhythmicity and impaired autophagy, as evidenced by consistent decreases in CLOCK and Beclin 1 expression, suggesting a correlation between CLOCK and autophagy. CLOCK protein expression was inhibited by ox-LDL, accompanied by defective autophagy and an increased apoptosis rates (P < 0.05). Administration of rapamycin (10 nM) reversed the effect of ox-LDL on VSMC autophagy and apoptosis. Finally, CLOCK silencing led to a considerable decrease in autophagy. VSMCs with stable CLOCK silencing also showed an increased apoptosis rate. In addition, gene silencing of CLOCK in VSMCs counteracted the effects of moderate rapamycin concentrations on autophagy and apoptosis. In conclusion, these findings suggested that the CLOCK-dependent rapamycin signaling pathway is a critical mediator in ox-LDL-induced VSMCs with defective autophagy that exacerbates plaque destabilization.

血管平滑肌细胞 (VSMCs) 中响应氧化应激的缺陷自噬可导致细胞凋亡和斑块不稳定。以前的研究表明，生物钟系统参与自噬调节和斑块进展。然而，昼夜节律影响 VSMC 自噬和斑块稳定性的机制仍不清楚。我们的研究描述了动脉粥样硬化斑块的昼夜节律特征，并验证了昼夜节律失调在 VSMC 自噬和细胞凋亡中的作用。我们发现，与稳定斑块相比，不稳定斑块中昼夜运动输出周期蛋白 kaput (CLOCK) 和 Beclin 1 的 mRNA 表达水平显着降低。在其他昼夜节律基因中未观察到显着差异。用氧化低密度脂蛋白（ox-LDL，80 μg/ml）处理的 VSMC 表现出异常的昼夜节律和自噬受损，这可以通过 CLOCK 和 Beclin 1 表达的持续降低来证明，这表明 CLOCK 与自噬之间存在相关性。ox-LDL抑制CLOCK蛋白表达，伴有自噬缺陷和细胞凋亡率增加（P <0.05）。给予雷帕霉素 (10 nM) 可逆转 ox-LDL 对 VSMC 自噬和细胞凋亡的影响。最后，CLOCK 沉默导致自噬显着减少。具有稳定 CLOCK 沉默的 VSMC 也显示出增加的细胞凋亡率。此外，VSMC 中 CLOCK 的基因沉默抵消了中等浓度的雷帕霉素对自噬和细胞凋亡的影响。综上所述，