Following a limited number of cell divisions, mesenchymal stem cells (MSCs) undergo senescence, and these senescent cells maintain metabolic modification and remain viable for long periods. Autophagy, an intracellular bulk degradation process, provides a survival effect for cells under stress. In this study, the effect of autophagy on senescent MSCs was analyzed. Following serial passaging, rat MSCs underwent replicative senescence, characterized by positive staining for senescence-associated β-galactosidase (SA-β-gal), and increased expression levels of p16 and p21. During MSC senescence, the levels of autophagic activity were increased, a greater number of autophagic vacuoles were observed in senescent MSCs by transmission electron microscopy, acridine orange staining was elevated and the expression levels of autophagy‑related proteins (microtubule‑associated protein 1A/1B‑light chain 3-II, Atg7 and Atg12) were increased. The role of autophagy in MSC senescence was further investigated through pharmacological inhibition of autophagy with bafilomycin A1 and 3-methyladenine. Inhibition of autophagy by pharmacological means reduced the rate of positive staining for SA-β-gal and the expression levels of senescence‑related proteins. In conclusion, these findings suggest that autophagy is activated during senescence and the autophagic activity may be a requirement for maintaining the senescent state of MSCs.

中文翻译：

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在长期培养过程中，自噬的抑制减轻了大鼠间充质干细胞的衰老状态。

在有限数量的细胞分裂后，间充质干细胞（MSC）发生衰老，这些衰老细胞保持代谢修饰并长期保持活力。自噬是细胞内大量降解的过程，可为处于压力下的细胞提供生存力。在这项研究中，分析了自噬对衰老间充质干细胞的影响。连续传代后，大鼠MSC经历复制性衰老，其特征是衰老相关的β-半乳糖苷酶（SA-β-gal）呈阳性染色，并且p16和p21的表达水平增加。在MSC衰老过程中，自噬活性水平增加，通过透射电镜观察到衰老MSC中有更多的自噬泡，cr啶橙染色增加，自噬相关蛋白（微管相关蛋白1A / 1B轻链3-II，Atg7和Atg12）的表达水平增加。通过药理学抑制bafilomycin A1和3-甲基腺嘌呤的自噬作用，进一步研究了自噬在MSC衰老中的作用。通过药理学方法抑制自噬会降低SA-β-gal的阳性染色率以及衰老相关蛋白的表达水平。总之，这些发现表明自噬在衰老过程中被激活，自噬活性可能是维持MSC衰老状态的必要条件。通过药理学抑制bafilomycin A1和3-甲基腺嘌呤的自噬作用，进一步研究了自噬在MSC衰老中的作用。通过药理学方法抑制自噬会降低SA-β-gal的阳性染色率以及衰老相关蛋白的表达水平。总之，这些发现表明自噬在衰老过程中被激活，自噬活性可能是维持MSC衰老状态的必要条件。通过药理学抑制bafilomycin A1和3-甲基腺嘌呤的自噬作用，进一步研究了自噬在MSC衰老中的作用。通过药理学方法抑制自噬会降低SA-β-gal的阳性染色率以及衰老相关蛋白的表达水平。总之，这些发现表明自噬在衰老过程中被激活，自噬活性可能是维持MSC衰老状态的必要条件。