降尿酸方对高尿酸血症大鼠肾损伤及肾组织EZH2表达的影响

Effects of Jiangniaosuan Prescription on Renal Injuries and Expression of EZH2 in Kidney Tissue of Hyperuricemia Rats

目的观察降尿酸方对高尿酸血症大鼠肾损伤及zeste基因增强子人类同源物2(EZH2)表达的影响,并探讨其作用机制。方法40只SD大鼠随机分为正常组、模型组、降尿酸方组和别嘌醇组,每组10只。采用氧嗪酸钾(1500 mg/kg)灌胃制备高尿酸血症大鼠模型,降尿酸方组和别嘌醇组分别予降尿酸方16 g/kg、别嘌醇25 mg/kg灌胃,正常组和模型组予生理盐水灌胃。12周后检测大鼠血肌酐(SCr)、血尿素氮(BUN)、血尿酸(SUA)、24 h尿蛋白(UTP),HE和Masson染色观察肾组织病理变化,TUNEL染色检测肾组织细胞凋亡,免疫组化检测肾组织Ⅰ型胶原蛋白(ColⅠ)、Ⅳ型胶原蛋白(ColⅣ)、Cleaved Caspase-3、EZH2蛋白表达,Western blot检测肾组织Cleaved Caspase-3、Bax、Bcl-2、EZH2、组蛋白H3第27位赖氨酸三甲基化(H3K27me3)蛋白表达。结果与正常组比较,模型组大鼠SCr、BUN、SUA和24hUTP水平明显升高(P<0.05),肾小管管腔扩张伴炎性细胞浸润,肾间质纤维化,凋亡细胞明显增加,肾组织ColⅠ、ColⅣ、Cleaved Caspase-3、Bax、EZH2、H3K27me3蛋白表达升高,Bcl-2蛋白表达降低,差异均有统计学意义(P<0.01);与模型组比较,降尿酸方组大鼠SCr、BUN、SUA和24 h UTP水平明显降低(P<0.05),肾小管管腔扩张、炎性细胞浸润、纤维组织增生明显减轻,凋亡细胞明显减少,肾组织ColⅠ、ColⅣ、Cleaved Caspase-3、Bax、EZH2、H3K27me3蛋白表达降低,Bcl-2蛋白表达升高,差异均有统计学意义(P<0.05,P<0.01)。Cleaved Caspase-3和EZH2免疫组化结果与Western blot结果一致。结论降尿酸方能改善高尿酸血症大鼠肾损伤,抑制肾纤维化,有效保护肾功能,其机制可能与抑制细胞凋亡,降低肾组织EZH2表达有关。

Objective To observe the effects of Jiangniaosuan Prescription on renal injuries and expression of human enhancer of zeste homolog 2(EZH2)in hyperuricemia rats;To discuss its mechanism of action.Methods Totally 40 SD rats were randomly divided into normal group,model group,Jiangniaosuan Prescription group and allopurinol group,with 10 rats in each group.Rats model of hyperuricemia were prepared by intragastric administration of potassium oxoxazine(1500 mg/kg).Jiangniaosuan Prescription group and allopurinol group were intragastrically administered at Jiangniaosuan Prescription 16 g/kg and allopurinol 25 mg/kg,respectively.The normal group and the model group was given normal saline for gavage.The serum creatinine(SCr),blood urea nitrogen(BUN),blood uric acid(SUA),and 24 h urine total protein(UTP)were detected after 12 weeks;HE and Masson staining were used to observe the pathological changes of kidney tissue;TUNEL staining was used to detect apoptosis of renal tissue cells;immunohistochemical staining was used to detect the expressions of typeⅠcollagen(ColⅠ),typeⅣcollagen(ColⅣ),Cleaved Caspase-3,and EZH2 in kidney tissue;Western blot was used to detect the expressions of Cleaved Caspase-3,Bax,Bcl-2,EZH2,and H3K27me3 protein in kidney tissue.Results Compared with the normal group,the levels of SCr,BUN,SUA and 24 h UTP in the model group increased significantly(P<0.05);the renal tubule lumen dilation accompanied by inflammatory cell infiltration,renal interstitial fibrosis,and apoptotic cells increased significantly;the proteins expression of ColⅠ,ColⅣ,Cleaved Caspase-3,Bax,EZH2,and H3K27me3 in kidney tissue increased,and the protein expression of Bcl-2 decreased,with statistical significance(P<0.01).Compared with the model group,the levels of SCr,BUN,SUA,and 24 h UTP in Jiangniaosuan Prescription group were significantly reduced(P<0.05),the renal tubules dilation,inflammatory cells infiltration,fibrous tissue proliferation,and apoptotic cells were significantly reduced;the proteins expression of ColⅠ,ColⅣ,Cleaved Caspase-3,Bax,EZH2,and H3K27me3 in kidney tissue decreased,and the protein expression of Bcl-2 increased,with statistical significance(P<0.05,P<0.01).Cleaved Caspase-3 and EZH2 immunohistochemistry results were consistent with Western blot results.Conclusion Jiangniaosuan Prescription can improve renal injuries in rats with hyperuricemia,inhibit renal fibrosis,and effectively protect renal function.The mechanism may be related to the inhibition of cell apoptosis and reduction of EZH2 expression in renal tissue.