目的:初步探究Klotho在草酸钙肾结石大鼠模型肾组织中的表达及其意义.方法:将30只6～8周龄雄性SD大鼠随机分为结石组和对照组,每组15只,结石组采用乙二醇和氯化铵诱导法构建草酸钙肾结石模型.28 d造模完成后采集大鼠24 h尿液、血清标本,比较两组血肌酐(Cr)、尿素氮(BUN)、血Ca2+、24 h尿Ca2+及草酸(Ox)指标;采集肾组织标本,HE染色比较各组肾脏病理改变及草酸钙结晶的沉积情况,荧光定量PCR、Western印迹及免疫组织化学染色检测两组肾组织中Klotho mRNA及蛋白表达,并对肾脏氧化应激相关指标总抗氧化能力(total antioxidant capacity,T-AOC)、丙二醛(malondialdehyde,MDA)、过氧化氢酶(catalase,CAT)进行测定.结果:结石组在光学显微镜观察下可见草酸钙结石晶体沉积并有伴肾小管病理性改变,血清Cr,BUN,24 h尿Ca2+及Ox较对照组升高,差异有统计学意义(P＜0.01),血Ca2+差异无统计学意义(P＞0.05);结石组Klotho mRNA及蛋白表达较对照组下降,差异有统计学意义(P＜0.01);肾脏氧化应激相关指标中,结石组T-AOC,CAT较对照组降低,MDA较对照组升高,差异均有统计学意义(P＜0.01).结论:草酸钙肾结石的形成与Klotho表达异常及氧化应激反应有关;Klotho可能通过调控Ca2+代谢和氧化应激反应参与草酸钙肾结石的发病过程.%Objective:To make an initial study of the expression and significance of the Klotho in renal tissues of rat models with the calcium oxalate kidney stone.Methods:Thirty male SD rats that were 6-8 weeks old were randomly divided into a calculus group and a control group,15 rats in each group.The calculus group were fed by ethylene glycol and ammonium chloride to construct the calcium oxalate kidney stone rats.After 28 days,the 24-hour urine and blood serum of the two groups were collected and the level of serum creatinine (Cr),urea nitrogen (BUN),blood calcium,urinary calcium of 24 h and oxalic acid (Ox) were compared between the two groups;at the same time,renal tissue specimens were collected,and the pathological changes of the kidney and the deposition of calcium oxalate crystals in rats were compared by HE staining;expression ofKlotho mRNA and protein in renal tissue was detected by fluorescence quantitative PCR,Western-blotting and immunohistochemistry staining.Total antioxidant capacity (T-AOC),malondialdehyde (MDA) and catalase (CAT) about renal oxidative stress index were measured also.Results:The crystal deposition of calcium oxalate stones and the pathological changes of renal tubules was observed under optical microscope,and the level of Cr,BUN,urinary calcium of 24 h and Ox were higher than those in the control group,the difference was statistically significant (P＜0.01),however,the difference of blood calcium was not statistically significant (P＞0.05),the expression level of Klotho mRNA and protein in renal tissue in the calculus group were lower than the control group,the difference was statistically significant (P＜0.01),the level of T-AO C and CAT in the calculus group were lower,whereas the level of MDA was higher,the difference were statistically significant (P＜0.01).Conclusion:The formation of calcium oxalate kidney stones relate to the abnormal expression of Klotho and oxidative stress,Klotho may participate in the pathogenesis of calcium oxalate kidney stones by regulating urinary Ca2+ metabolism and oxidative stress reaction.
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