对香豆酸通过抑制RhoA/ROCK信号通路减轻糖尿病肾病病变  

p-coumaric acid alleviates diabetic nephropathy by inhibiting RhoA/ROCK signaling pathway

作　　者：王晶 朱燕亭 吴冰 金刚[1] 王琼[1] WANG Jing;ZHU Yanting;WU Bing;JIN Gang;WANG Qiong(Kidney Diseases and Hemodialysis Center,Shaanxi Provincial People′s Hospital,Xi′an 710068,China)

机构地区：[1]陕西省人民医院肾病血透中心,西安710068

出　　处：《山西医科大学学报》2024年第8期976-984,共9页Journal of Shanxi Medical University

基　　金：陕西省自然科学基金项目(2021JQ-905);陕西省人民医院科技发展孵化基金项目(2023YJY-60)。

摘　　要：目的探讨对香豆酸(p-CA)对糖尿病肾病(DN)大鼠的治疗作用及对Ras同源基因家族成员A(RhoA)/Rho相关卷曲螺旋蛋白激酶(ROCK)信号通路的影响。方法将大鼠分为6组:对照组(n=10)、DN组(n=10)、低剂量对香豆酸组(L-p-CA)(n=10)、中剂量对香豆酸组(M-p-CA)(n=10)、高剂量对香豆酸组(H-p-CA)(n=10)和RhoA激动剂U-46619组(U-46619)(n=12)。对照组大鼠为健康SD大鼠,其他组大鼠均为高脂饮食联合链脲佐菌素诱导的DN模型大鼠。造模后,对照组和DN组大鼠灌胃2 mL 0.5%羧甲基纤维素溶液,L-p-CA组、M-p-CA组和H-p-CA组大鼠分别灌胃2 mL剂量为50,100,200 mg/(kg·d)的对香豆酸溶液,U-46619组大鼠同时灌胃1 mL对香豆酸溶液(剂量为200 mg/(kg·d))以及1 mL剂量为30μg/(kg·d)的RhoA激动剂U-46619。各组大鼠均干预12周。治疗后检测各组大鼠生化指标水平:空腹血糖(FPG)、空腹胰岛素(FINS)、糖化血红蛋白(HbA1c)、尿素氮(BUN)、血肌酐(Cr)和24 h尿蛋白;以及血清氧化应激指标水平:超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、谷胱甘肽过氧化物酶(GSH-PX)和丙二醛(MDA)。通过苏木精-伊红(HE)和Masson三色染色评价大鼠肾脏损伤和纤维化。通过qRT-PCR检测肾脏肿瘤坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)和单核细胞趋化蛋白-1(MCP-1)转录活性。通过Western blot检测肾脏TNF-α、IL-1β、MCP-1、RhoA和ROCK1蛋白表达水平。结果与对照组比较,DN组大鼠的FPG、FINS、24 h尿蛋白、HbA1c、BUN和Cr水平升高(P<0.05);肾组织发生明显病变,肾组织纤维化面积升高(P<0.05);血清SOD、CAT和GSH-PX水平均降低(P<0.05),MDA水平升高(P<0.05);肾组织TNF-α、IL-1β和MCP-1的mRNA和蛋白水平均升高(P<0.05);肾组织RhoA和ROCK1蛋白表达水平升高(P<0.05)。与DN组比较,L-p-CA组、M-p-CA组和H-p-CA组大鼠的FPG、FINS、24 h尿蛋白、HbA1c、BUN和Cr水平降低(P<0.05);肾组织病变减轻,肾组织纤维化面积降低(P<0.05);血清SOD、CAT和GObjective To investigate the therapeutic effect of p-coumaric acid(p-CA)on diabetic nephropathy(DN)rats and its influence on Ras homolog gene family member A(RhoA)/Rho associated coiled-coil forming protein kinase(ROCK)signaling pathway.Methods Rats were divided into 6 groups:control group(n=10),DN group(n=10),low dose p-coumaric acid(L-p-CA)group(n=10),medium dose p-coumaric acid(M-p-CA)group(n=10),high dose p-coumaric acid(H-p-CA)group(n=10)and RhoA agonist U-46619 group(n=12).Rats in control group were healthy SD rats,and rats in the other groups were given high-fat diet combined with streptozotocin to induce DN models.After modeling,the rats in control group and DN group were intragastrically given 2 mL of 0.5%carboxymethyl cellulose solution,the rats in L-p-CA group,M-p-CA group and H-p-CA group were given intragastrically 2 mL of 50,100 and 200 mg/(kg·d)p-coumaric acid solution respectively,and the rats in U-46619 group were simultaneously and intragastrically given 1 mL of p-coumaric acid solution[200 mg/(kg·d)]and 1 mL of RhoA agonist U-46619[30μg/(kg·d)].All the rats were intervened for 12 weeks.After treatment,the levels of biochemical indexes[fasting blood glucose(FPG),fasting insulin(FINS),glycosylated hemoglobin(HbA1c),urea nitrogen(BUN),blood creatinine(Cr)and 24-hour urinary protein]and serum oxidative stress indexes[superoxide dismutase(SOD),catalase(CAT),glutathione peroxidase(GSH-PX)and malondialdehyde(MDA)]were mea-sured.HE staining and Masson trichrome staining were used to evaluate renal injury and fibrosis in rats.The transcriptional activities of tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β)and monocyte chemoattractant protein-1(MCP-1)in kidney were detected by qRT-PCR.The protein expression levels of TNF-α,IL-1β,MCP-1,RhoA and ROCK1 in kidney were detected by Western blot.Results Compared with control group,the levels of FPG,FINS,24 h urinary protein,HbA1c,BUN and Cr were increased in DN group(P<0.05),obvious pathological changes occurred in renal tissue,the area of renal

关 键 词：对香豆酸 糖尿病肾病 Ras同源基因家族成员A Rho相关卷曲螺旋蛋白激酶 氧化应激 炎症 纤维化 

分 类 号：R587.2[医药卫生—内分泌]