盐炙车前子调控肾小管上皮-间充质转化改善肾纤维化的机制研究  

Saltwater stir-fried Plantaginis Semen alleviates renal fibrosis by regulating epithelial-mesenchymal transition in renal tubular cells

作　　者：沈鑫蕾 朱清如 余文凯 周莉 单琪媛[1] 张易航 鲍旖旎 曹岗[1] SHEN Xin-lei;ZHU Qing-ru;YU Wen-kai;ZHOU Li;SHAN Qi-yuan;ZHANG Yi-hang;BAO Yi-ni;CAO Gang(School of Pharmacy,Zhejiang Chinese Medical University,Hangzhou 310053,China)

机构地区：[1]浙江中医药大学药学院,浙江杭州310053

出　　处：《中国中药杂志》2025年第5期1195-1208,共14页China Journal of Chinese Materia Medica

基　　金：国家自然科学基金青年科学基金项目(82204625);浙江省自然科学基金项目(LZ22H280001,LQ23H280013);浙江省中医药科技计划项目(2023ZR009);浙江中医药大学校级科研项目(2021RCZXZK14);浙江中医药大学研究生科学研究基金项目(2023YKJ10);浙江省大学生科技创新活动计划(新苗人才计划)项目(2024R410A034)。

摘　　要：探讨盐炙车前子对大鼠肾纤维化的影响及作用机制。选取36只SD大鼠,分成空白组,模型组,氯沙坦钾组,盐炙车前子低、中、高剂量组(15、30、60 g·kg^(-1))。除空白组外,其余各组采用单侧输尿管结扎(unilateral ureteral obstruction,UUO)手术建立大鼠肾纤维化模型,造模和灌胃给药周期均为14 d。连续给药14 d后,采用全自动生化仪检测各组大鼠血清肌酐(se-rum creatinine,Scr)、尿素氮(blood urea nitrogen,BUN)水平;通过苏木素-伊红(hematoxylin-eosin,HE)及Masson染色评价肾脏组织病理变化;Western blot及免疫荧光法检测肾脏组织纤连蛋白(fibronectin,FN)、Ⅰ型胶原蛋白(collagen Ⅰ)、波形蛋白(vimentin)、α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)的表达水平;采用RT-qPCR测定肾小管上皮-间充质转化(epithelial-mesenchymal transition,EMT)关键基因扭曲家族bHLH转录因子1(twist family bHLH transcription factor 1,TWIST1)、蜗牛家族转录抑制因子1(snail family transcriptional repressor 1,SNAI1)、E盒结合锌指蛋白1(zinc finger E-box binding homeobox 1,ZEB1),以及炎症因子白细胞介素-1β(interleukin-1β,IL-1β)、白细胞介素-6(interleukin-6,IL-6)、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)的基因表达水平。体外建立转化生长因子-β(transforming growth factor-β,TGF-β)诱导肾小管上皮HK-2细胞纤维化模型,考察盐炙车前子对肾小管上皮细胞EMT的抑制作用。结合网络药理学和Western blot进一步研究盐炙车前子缓解肾纤维化的分子机制。结果表明,与模型组比较,盐炙车前子能明显降低UUO大鼠血清Scr、BUN水平,减少肾小管损伤和胶原沉积;也可显著抑制UUO大鼠肾脏组织及TGF-β刺激HK-2细胞中FN、collagen Ⅰ、vimentin、α-SMA的蛋白表达,以及SNAI1、ZEB1、TWIST1、IL-1β、IL-6、TNF-α的基因表达。进一步研究发现盐炙车前子含量显著增加的成分主要富集丝裂原活化蛋白激酶(mitogen activated This study aimed to investigate the effect of saltwater stir-fried Plantaginis Semen(SPS)on renal fibrosis in rats and decipher the underlying mechanism.Thirty-six Sprague-Dawley rats were randomly assigned into control,model,losartan potassium,and low-,medium-,and high-dose(15,30,and 60 g·kg^(-1),respectively)SPS groups.Rats in other groups except the control group were subjected to unilateral ureteral obstruction(UUO)to induce renal fibrosis,and the modeling and gavage lasted for 14 days.After 14 consecutive days of treatment,the levels of serum creatinine(Scr)and blood urea nitrogen(BUN)in rats of each group were determined by an automatic biochemical analyzer.Hematoxylin-eosin(HE)and Masson staining were used to evaluate pathological changes in the renal tissue.Western blot and immunofluorescence assay were conducted to determine the protein levels of fibronectin(FN),collagen Ⅰ,vimentin,andα-smooth muscle actin(α-SMA)in the renal tissue.The mRNA levels of epithelial-mesenchymal transition(EMT)-associated transcription factors including twist family bHLH transcription factor 1(TWIST1),snail family transcriptional repressor 1(SNAI1),and zinc finger E-box binding homeobox 1(ZEB1),as well as inflammatory cytokines such as interleukin-1β(IL-1β),interleukin-6(IL-6),and tumor necrosis factor-α(TNF-α),were determined by RT-qPCR.Human renal proximal tubular epithelial(HK2)cells exposed to transforming growth factor-β(TGF-β)for the modeling of renal fibrosis were used to investigate the inhibitory effect of SPS on EMT.Network pharmacology and Western blot were employed to explore the molecular mechanism of SPS in alleviating renal fibrosis.The results showed that SPS significantly reduced Scr and BUN levels and alleviated renal injury and collagen deposition in UUO rats.Moreover,SPS notably down-regulated the protein levels of FN,collagen Ⅰ,vimentin,andα-SMA as well as the mRNA levels of SNAI1,ZEB1,TWIST1,IL-1β,IL-6,and TNF-α in the kidneys of UUO rats and TGF-β-treated HK-2 cells.In addition,compared

关 键 词：车前子 盐炙 肾纤维化 上皮-间充质转化(EMT) MAPK信号通路 

分 类 号：R285.5[医药卫生—中药学]