肾纤康通过调节Smad3介导的铁死亡缓解CKD小鼠肾纤维化  

Shen-Xiankang formula alleviates renal fibrosis in chronic kidney disease mice by regulating Smad3-mediated ferroptosis

作　　者：倪玉芳 张璐娜 张琼 王丽 李健春 NI Yufang;ZHANG Luna;ZHANG Qiong;WANG Li;LI Jianchun(Research Center of Integrated Traditional Chinese and Western Medicine,The Affiliated Traditional Chinese Medicine Hospital,Southwest Medical University,Luzhou 646000,China;Department of Nephrology,Affiliated Chinese Traditional Medicine Hospital,Southwest Medical University,Luzhou 646000,China)

机构地区：[1]西南医科大学附属中医医院中西医结合研究中心,四川泸州646000 [2]西南医科大学附属中医医院肾病科,四川泸州646000

出　　处：《中国病理生理杂志》2024年第6期1097-1104,共8页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.82104665);四川省科技厅项目(No.2023NSFSC1763,No.2022YFS0621);西南医科大学中西医结合基金资助项目(No.2023ZYYQ09)。

摘　　要：目的:观察肾纤康对单侧输尿管梗阻(unilateral ureteral obstruction,UUO)引起的小鼠肾间质纤维化的缓解效果,并探讨其作用机制。方法:将小鼠随机分为假手术组、模型组、低剂量(150 mg·kg^(−1)·d^(−1))肾纤康组和高剂量(450 mg·kg^(−1)·d^(−1))肾纤康组,每组8只。除假手术组外,其余各组均通过单侧输尿管结扎法建立慢性肾脏病(chronic kidney disease,CKD)模型。造模后,分别给予相应剂量的肾纤康,假手术组和模型组则灌胃等体积生理盐水,每天1次,连续7 d。实验结束后,收集小鼠肾脏组织进行苏木精-伊红(hematoxylin-eosin,HE)染色和Masson染色,观察肾脏组织损伤和纤维化程度;利用免疫组化和Western blot技术检测肾脏中纤维化、氧化应激及铁死亡相关蛋白水平的变化;蛋白质免疫共沉淀(co-immunoprecipitation,Co-IP)评估肾纤康对转录激活因子3(activating transcription factor 3,ATF3)与Smad3相互作用的影响。结果:在未经治疗的UUO模型组中,小鼠肾脏出现了如肾小管明显扩张和胶原沉积等典型的病理变化,肾纤康干预组病理改变程度和纤维化明显减轻(P<0.05)。在分子层面,肾纤康显著降低了UUO模型组小鼠中Smad3磷酸化水平,以及ATF3、4-羟基壬烯醛(4-hydroxynonenal,4-HNE)和NADPH氧化酶4(NADPH oxidase 4,NOX4)的异常表达,同时增加了谷胱甘肽过氧化酶4(glutathione peroxidase 4,GPX4)和溶质载体家族7成员11(solute carrier family 7 member 11,SLC7A11)表达。Co-IP实验结果表明,肾纤康显著影响了ATF3与Smad3的相互作用。结论:肾纤康有效缓解了UUO引起的小鼠肾间质纤维化,其潜在机制可能涉及对ATF3/Smad3相互作用的调节,进而减轻氧化应激和铁死亡,从而缓解肾脏纤维化。这些发现为肾纤康的进一步研究和临床应用提供了重要的科学依据。AIM:To evaluate the therapeutic effect of Shen-Xiankang formula on renal interstitial fibrosis induced by unilateral ureteral obstruction(UUO)in mice and to elucidate its underlying mechanisms.METHODS:Thirtytwo C57BL/6 mice were randomly divided into sham group,UUO model group,and Shen-Xiankang formula intervention groups receiving either a low dose(150 mg·kg^(−1)·d^(−1))or a high dose(450 mg·kg^(−1)·d^(−1)),with 8 mice in each group.All mice except those in sham group underwent UUO to establish chronic kidney disease(CKD)model.After modeling,corresponding doses of Shen-Xiankang or an equivalent volume of saline were administered daily for 7 d.Upon completion of treatment,renal tissues were collected for hematoxylin-eosin(HE)and Masson staining to assess tissue damage and fibrosis.Immunohistochemistry and Western blot analyses were used to detect the markers of fibrosis,oxidative stress,and fer‐roptosis.The effect of Shen-Xiankang formula on the interaction between activating transcription factor 3(ATF3)and Smad3 was assessed using co-immunoprecipitation(Co-IP).RESULTS:The untreated UUO model group exhibited notable pathological changes such as expanded renal tubules and collagen deposition.Shen-Xiankang treatment significantly alleviated these changes(P<0.05).It markedly reduced Smad3 phosphorylation,ATF3,4-hydroxynonenal(4-HNE),and NADPH oxidase 4(NOX4)aberrant expression,while increasing glutathione peroxidase 4(GPX4)and solute carrier family 7 member 11(SLC7A11)expression.Co-IP results indicated a significant modulation of the ATF3-Smad3 interaction by Shen-Xiankang.CONCLUSION:Shen-Xiankang formula effectively mitigates UUO-induced renal interstitial fibrosis in mice.The mechanism may involve modulating the ATF3/Smad3 interaction,which in turn attenuates oxidative stress and ferroptosis,consequently leading to the amelioration of renal fibrosis.These findings provide important insights for further research and clinical application of Shen-Xiankang formula.

关 键 词：肾纤康 慢性肾脏病 铁死亡 SMAD3蛋白 转录激活因子3 

分 类 号：R285.5[医药卫生—中药学] R363[医药卫生—中医学] R692.5