人胚肺成纤维细胞复制性衰老及过氧化氢诱导的早衰研究

Study on the replicative and premature senescence induced by hydrogen peroxide of human embryonic lung fibroblasts

目的研究关于人胚肺成纤维细胞复制性衰老及过氧化氢诱导的早衰。方法常规传代培养人胚肺二倍体成纤维细胞及应用过氧化氢染毒诱导细胞早衰发生,利用细胞衰老综合指标进行评价,包括细胞形态学改变、生长曲线、寿命周期、细胞周期分布及衰老相关β-半乳糖苷酶染色等,同时检测细胞衰老不同阶段增殖细胞核抗原PCNA的mRNA和蛋白水平表达变化。结果人胚肺二倍体成纤维细胞于52PDL(群体倍增水平)处于细胞复制性衰老状态,400μmol/L过氧化氢可短期内诱导细胞过早衰老,衰老的细胞变大扁平,饱和度降低,细胞不可逆的阻滞于G1期,衰老相关β-半乳糖苷酶染色阳性率增加,PCNA表达随着衰老程度的增加而逐渐下降。结论400μmol/L过氧化氢以一定方式作用于细胞可以诱导早衰发生,细胞复制性衰老与细胞早衰具有相同的生物学特征和增殖能力。

Objective To study both cellular replieative and prematuce senescence induced by hydrogen peroxide （H2 O2 ） of human embryonic lung fibroblasts （HEFs） . Methods HEFs were cultured conventionally and treated with H2O2 . Biomarkers of senescence were assessed, including cell morphology, growth curve, life span, distribution of cell cycle and senescence associated β-galactosidase （SA-β-gal）, and the expression of proliferating cell nuclear antigen （PCNA）. Results HEFs entered replieative senescence status at 52 population doubling levels （PDL） and 400μmo/L H2O2 induced premature senescence of HEFs in a short period. Senescent ceils were characterized as enlarged, flattened and reduced saturation density, retarded in the G1 phase with increased expression of SA-β-gal. The mRNA and protein Level of PCNA decreased gradually during cellular senescence. Conclusion Cellular premature senescence could be induced by 400μmol/L H2O2 . Premature senescence induced by H2O2 could share the morphology characteristics and proliferative capacity similar to those of replicative one.