通络保肾方调控NLRP3/GSDMD通路抑制细胞焦亡改善IgA肾病大鼠足细胞损伤的机制  

Mechanistic Study of Tongluo Baoshen Formula in Ameliorating Podocyte Injury in IgA Nephropathy Rats by Regulating NLRP3/GSDMD Pathway to Inhibit Pyroptosis

作　　者：刘永芳[1] 周莉 刘晖扬 代建风 刘英华 陈帮明 林雪菲 杨太旺 刘星宇 LIU Yongfang;ZHOU Li;LIU Huiyang;DAI Jianfeng;LIU Yinghua;CHEN Bangming;LIN Xuefei;YANG Taiwang;LIU Xingyu(Jiujiang Traditional Chinese Medicine Hospital Affiliated to Jiangxi University of Chinese Medicine,Jiujiang 332000,China;Jiangxi University of Chinese Medicine,Nanchang 330004,China)

机构地区：[1]江西中医药大学附属九江市中医院,江西九江332000 [2]江西中医药大学,江西南昌330004

出　　处：《中医药信息》2025年第4期1-11,共11页Information on Traditional Chinese Medicine

基　　金：国家自然科学基金项目(82060839);江西省中医肾病临床医学研究中心建设项目(2019BCD42006);国家中医优势专科(肾病科)建设项目(赣中医药综合字〔2023〕3号);江西省中医优势病种肾衰病防治中心建设项目(赣中医药医政字〔2021〕8号);全国名老中医药专家付义传承工作室建设项目(国中医药人教函〔2022〕75号)。

摘　　要：目的:观察通络保肾方改善IgA肾病(IgAN)大鼠足细胞损伤的作用并探讨其作用机制。方法:130只SD大鼠随机选择20只作为正常组,其余110只为造模组;造模组采用牛血清白蛋白+四氯化碳+脂多糖方法建立IgAN模型,最终100只大鼠建模成功。100只IgAN模型大鼠随机分为模型组、氯沙坦钾组[5×10-3 g(/kg·d)]及通络保肾方低、中、高剂量组[5.3、10.6、21.2 g(/kg·d)],每组20只。各给药组给予相应剂量药物灌胃,正常组及模型组给予等体积蒸馏水,均每日灌胃1次,连续给药,于给药4周(各组随机选10只)、8周(剩余大鼠)分两批采集标本。ELISA法及肌氨酸氧化酶法检测尿微量白蛋白/尿肌酐比(ACR)、白细胞介素-18(IL-18)、胱天蛋白酶-1(Caspase-1)、GSDMD及血IgA;实时荧光定量法和蛋白免疫印迹法检测肾组织NOD样受体蛋白3(NLRP3)、GSDMD-NT、IL-18、Integrinα3、Integrinβ1 mRNA和蛋白表达;给药8周时,免疫组化法检测肾组织NLRP3、Caspase-1阳性表达,免疫荧光法检测GSDMD-NT表达;透射电镜观察足细胞超微结构改变。结果:与正常组比较,模型组大鼠尿ACR、Caspase-1、GSDMD、IL-18显著升高(P<0.01),且随成模时间延长,ACR、IL-18水平逐渐升高(P<0.05或P<0.01);血IgA无显著差异(P>0.05);肾组织NLRP3、GSDMD-NT、IL-18 mRNA和蛋白表达显著升高(P<0.01),Integrinα3、Integrinβ1 mRNA和蛋白表达显著降低(P<0.01),且随成模时间延长逐渐降低(P<0.01);免疫组化显示NLRP3、Caspase-1阳性面积占比显著增加(P<0.01),两者在肾小球内主要表达于毛细血管袢及系膜区,肾小管少量表达;免疫荧光显示GSDMD-NT平均荧光强度显著增强(P<0.01),主要表达于肾小球毛细血管袢及系膜区;超微结构显示足细胞胞体增大,足突部分融合,足突内微丝减少,溶酶体内可见膜样结构,节段胞膜破裂;与模型组比较,给药4周、8周时通络保肾方低、中、高剂量组和氯沙坦钾组尿ACR、IL-18、Caspase-1显著�Objective:To observe the effect of Tongluo Baoshen Formula(TLBSF)on improving podocyte injury in IgA nephropathy(IgAN)rats and explore its mechanism.Methods:A total of 130 SD rats were randomly divided,with 20 rats in the normal group and 110 in the modeling group. The IgAN model was established using bovine serum albumin,carbon tetrachloride, and lipopolysaccharide. Finally, 100 model rats were successfully constructed and randomlydivided into five groups: model group, losartan potassium group[ 5×10−³ g(/ kg·d)], and low, medium, and high-doseTLBSF groups [5. 3, 10. 6, 21. 2 g(/ kg·d)], each containing 20 rats. The treatment groups were administered therespective doses via gavage, while the normal and model groups received equal volumes of distilled water. After 4 and 8weeks of treatment, samples were collected from 10 randomly selected rats per group at each time point. ELISA andcreatinine oxidase methods were used to detect the urinary albumin-to-creatinine ratio( ACR), interleukin-18( IL-18), caspase-1, gasdermin D( GSDMD), and blood IgA levels. NOD-like receptor protein 3( NLRP3), GSDMDN-terminal (GSDMD-NT), IL-18, integrin α3, and integrin β1 mRNA and protein expression levels in kidneytissue were measured using real-time quantitative PCR and Western blot. Immunohistochemistry was used to detectNLRP3 and caspase-1 expression in kidney tissue, and immunofluorescence was employed to observe GSDMD-NTexpression. Transmission electron microscopy was utilized to assess podocyte ultrastructural changes. Results:Compared with the normal group, the model group showed significantly increased urinary ACR, caspase-1, GSDMD,and IL-18 levels (P < 0. 01), which progressively increased with prolonged modeling duration (P < 0. 05 or P <0. 01). However, blood IgA levels did not differ significantly (P > 0. 05). NLRP3, GSDMD-NT, IL-18 mRNA,and protein expression levels were significantly elevated in kidney tissue (P < 0. 01), while integrin α3 and integrin β1mRNA and protein levels were significantly reduced (P < 0.

关 键 词：通络保肾方 IGA肾病 足细胞损伤 NLRP3/GSDMD通路 细胞焦亡 

分 类 号：R285.5[医药卫生—中药学]