H2O2诱导的2BS早老细胞中自噬体增多

自噬在细胞复制性衰老中起着重要的作用.然而,早老细胞中的自噬现象基本无相关的报道.本文通过外源性过氧化氢(H₂O₂)的诱导，构建人胚肺二倍体成纤维细胞（2BS细胞）早老模型.首先,通过SA-β-gal染色，验证细胞早老；从形态学和特异标志分子及雷帕霉素作用的靶位点(mTOR)信号通路不同角度检测自噬的变化，其中形态学检测包括丹（磺）酰戊二胺(MDC)自噬分子定量法及电镜自噬超微结构的观察；特异标志分子LC3的检测包括GFP-LC3自噬定位法和免疫印迹法检测LC3；及检测mTOR信号通路下游激酶p70S6蛋白的表达变化.结果表明，过氧化氢诱导的早老细胞中自噬体相对年轻细胞明显增多，且具有保护早老细胞的作用.

Autophagosomes Increased in H2O2-induced Premature Senescent 2BS Cells

Autophagy plays a critical role in replicative senescence cells. However, it remains poorly understood. Thereby we constructed the exogenous H2O2-nduced 2BS premature senescent cell model. And the β-Galactosidase (β-Gal) staining was conducted to confirm premature senescence. Morphological methods, specific markers for autophagy and mTOR signaling pathway were used to illuminate the change of autophagosomes. Monodansylcadaverine (MDC) staining and electron microscopy were firstly applied to detect the morphology of autophagosomes. GFP-LC3 was analyzed by fluorescence microscopy and immunoblotting was employedto examine the protein expression of LC3. Then, p70S6, the downstream effector of mTOR pathway, was examined by Western blotting. Consequently, we found that in the H2O2-induced premature senescent 2BS cell, autophagosomes increased in contrast to the young cells and have a protective function in premature senescence.