目的研究p21野生型p53活化片段1/细胞周期蛋白依赖性激酶影响蛋白1/衰老细胞衍生抑制剂1(p21WAF1/CIP1/SDI1)在大鼠肾脏中随年龄增长的表达变化规律。方法取3月龄、12月龄及24月龄健康雄性Wistar大鼠肾组织进行衰老相关β-乳糖苷酶(SA-β-al)活性染色,TUNEL法检测细胞凋亡,采用逆转录多聚酶链反应(RTPCR)和Western印迹法(Western blot assay)分别在基因及蛋白质表达水平上检测肾脏组织中p21WAF1/CIP1/SDI1的表达变化,并用免疫组化法检测p21WAF1/CIP1/SDI1在肾脏组织中的表达与定位。结果大鼠肾脏组织SA-β-al活性随年龄增长逐渐增强,凋亡细胞也随年龄增长逐渐增加(P<0.05);p21WAF1/CIP1/SDI1 mRNA表达随年龄增长逐渐增强,不同月龄比较差异有统计学意义(P<0.05)。Western印迹亦显示p21WAF1/CIP1/SDI1蛋白表达随鼠龄增加逐渐增强(P<0.05)。免疫组化结果显示,p21WAF1/CIP1/SDI1蛋白表达于大鼠肾小球足细胞,其在肾小管与间质细胞中也有表达,且随年龄增长表达增加(P<0.05)。 结论p21WAF1/CIP1/SDI1在大鼠肾脏组织中的表达随年龄增加而增强,可作为肾脏组织中重要的衰老指标。
ObjectiveTo investigate the effect of p21WAF1/CIP1/SDI1 on the renal lesions in aging kidneys in rats. MethodsMale Wistar rats at the ages of 3, 12, 24 months were used in this study. The renal tissues were processed for senescence associated βgalactosidase (SA-β-al) staining. TUNEL assay was used to detect apoptosis of renal tubular epithelial cells. The expression and location of p21WAF1/CIP1/SDI1 in kidneys were studied by immunohistochemistry. The mRNA and protein levels of p21WAF1/CIP1/SDI1 were detected by Western blot and RTPCR, respectively. ResultsThe expression of neutral β-galactosidase activity was increased in aging kidneys. The number of apoptotic cells increased in kidney with advancing age(P〈0.05). The p21WAF1/CIP1/SDI1 staining appeared in the podocyte and interstitium in kidney. The staining intensity of p21WAF1/CIP1/SDI1 in kidneys in old animals was markedly increased, as compared to that in young and middleaged rats (P〈0.05).The mRNA level of p21WAF1/CIP1/SDI1 was significantly increased in aging kidneys, as compared to that at the age of 3 and 12 months, meanwhile the p21WAF1/CIP1/SDI1 proteins expression was dramatically increased with advancing age (P〈0.05). Conclusionsp21WAF1/CIP1/SDI1 expression increases with the aging of kidneys, which may serve as an important marker of organismal aging.