Atherosclerosis (AS) is commonly regarded as a key driver accounted for the leading causes of morbidity and mortality among cardiovascular and cerebrovascular diseases. A growing body of evidence indicates that autophagy in macrophages involved in AS might be a potential therapeutic target. C1q/TNF-related protein 9 (CTRP9) has been proven to delay the progression of cardiovascular diseases. However, the relations between CTRP9 and Sirt1, as well as their effects on macrophages autophagy have not been fully explored. Macrophages were differentiated from mononuclear cells collected from peripheral blood samples of healthy donors. The AS models were constructed by ox-LDL treatment. Cell viability was determined by CCK-8 assay. Immunofluorescence assay of LC3 was implemented for evaluating autophagy activity. Oil Red O staining was performed for lipid accumulation detection. ELISA, cholesterol concentration assay and cholesterol efflux analysis were conducted using commercial kits. Cycloheximide assay was implemented for revealing protein stability. RT-qPCR was used for mRNA expression detection, and western blotting was performed for protein level monitoring. CTRP9 attenuated impaired cell viability, autophagy inhibition and increased lipid accumulation induced by ox-LDL. Moreover, CTRP9 maintained Sirt1 protein level through enhancing its stability through de-ubiquitination, which was mediated by upregulated USP22 level. CRTP9 exerted its protective role in promoting autophagy and reducing lipid accumulation through the USP22/Sirt1 axis. Collectively, CTRP9 alleviates lipid accumulation and facilitated the macrophages autophagy by upregulating USP22 level and maintaining Sirt1 protein expression, thereby exerting a protective role in AS progression .

中文翻译：

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CTRP9 在体外激活 USP22 介导的 Sirt1 去泛素化，从而改变巨噬细胞的自噬状态，从而预防动脉粥样硬化进展

动脉粥样硬化（AS）通常被认为是心脑血管疾病发病和死亡的主要原因。越来越多的证据表明，与 AS 相关的巨噬细胞自噬可能是一个潜在的治疗靶点。 C1q/TNF 相关蛋白 9 (CTRP9) 已被证明可以延缓心血管疾病的进展。然而，CTRP9和Sirt1之间的关系以及它们对巨噬细胞自噬的影响尚未得到充分探讨。巨噬细胞是从健康捐献者外周血样本中收集的单核细胞分化而来的。通过ox-LDL治疗构建AS模型。通过CCK-8测定法测定细胞活力。 LC3 的免疫荧光测定用于评估自噬活性。进行油红O染色以检测脂质积累。使用商业试剂盒进行ELISA、胆固醇浓度测定和胆固醇流出分析。实施放线菌酮测定以揭示蛋白质稳定性。 RT-qPCR用于mRNA表达检测，Western blotting用于蛋白质水平监测。 CTRP9 减弱了 ox-LDL 诱导的细胞活力受损、自噬抑制和脂质积累增加。此外，CTRP9 通过去泛素化增强 Sirt1 蛋白的稳定性，从而维持 Sirt1 蛋白水平，而去泛素化是由上调的 USP22 水平介导的。 CRTP9通过USP22/Sirt1轴发挥其促进自噬和减少脂质积累的保护作用。总的来说，CTRP9 通过上调 USP22 水平和维持 Sirt1 蛋白表达来减轻脂质积累并促进巨噬细胞自噬，从而在 AS 进展中发挥保护作用。