目的 探讨当归芍药散(DSS)含药血清及其有效成分阿魏酸(FA)、芍药苷(PF)对肾间质纤维化(RIF)大鼠的作用及机制。方法 制备当归芍药散药液，以低剂量(0。05 mL DSS药液/10 g体质量)、中剂量(0。1 mL DSS药液/10 g体质量)、高剂量(0。2 mL DSS药液/10 g体质量)每日同一时间灌胃雄性Sprague Dawley大鼠，持续7 d，腹主动脉取血获得相应低、中、高剂量DSS血清(DSSS)。将大鼠肾小管上皮细胞(NRK-52E)分成对照组、模型组(TGF-β处理)、siRNA转染组(TGF-β+siRNA处理)、FA组(TGF-β+FA处理)、PF组(TGF-β+PF处理)及DSSS低、中、高剂量组(TGF-β+DSSS-L、TGF-β+DSSS-M、TGF-β+DSSS-H分别处理)。筛选并确定DSSS、FA、PF、TGF-β对NRK-52E细胞的干预浓度，对以上各组NRK-52E细胞进行干预。采用Western blotting法检测各组NLRP3、半胱氨酸蛋白酶1(CASP-1)、Gasdermin D(GSDMD)、白细胞介素-18(IL-18)、纤连蛋白(FN)、E型钙黏蛋白(E-Cadherin)、N型钙黏蛋白(N-Cadherin)的蛋白表达变化。结果 相较于模型组，DSSS、FA、PF治疗组细胞焦亡相关蛋白NLRP3、CASP-1、GSDMD、IL-18表达均降低，差异均有统计学意义(P均<0。05)。DSSS中、高剂量及FA、PF治疗组相较于模型组纤维化相关黏附蛋白FN、E-Cadherin、N-Cadherin表达降低，差异均有统计学意义(P均<0。05)。结论 当归芍药散含药血清及有效成分FA、PF可抑制TGF-β诱导的NRK-52E细胞纤维化模型中NLRP3介导的焦亡蛋白激活，并能调节黏附蛋白表达和上皮-间质转化进展，缓解纤维化。
Effects of Danggui Shaoyao San-medicated serum and its active ingredients on renal interstitial fibrosis rats
Objective To investigate the interventional effects of Danggui Shaoyao San(DSS)-medicated serum and its active ingredients ferulic acid(FA)and paeoniflorin(PF)on renal interstitial fibrosis(RIF)rats and the mecha-nism.Methods DSS solution was prepared and administered to male Sprague Dawley rats by gavage at the same time of the day at low(0.05 mL DSS solution/10 g body mass),medium(0.1 mL DSS solution/10 g body mass),and high(0.2 mL DSS solution/10 g body mass)doses for 7 d.Blood was obtained from the abdominal aorta to obtain the equiva-lent low-,medium-,and high-dose of DSS serum(DSSS).Rat renal tubular epithelial cells(NRK-52E)were divided in-to the control group(Control),the model group(treated with TGF-β),the siRNA transfection group(treated with TGF-β +siRNA),the FA group(treated with TGF-β+FA),the PF group(treated with TGF-β+PF),and the low-,medium-,and high-dose DSSS groups(treated with TGF-β+DSS-L,TGF β+DSS-M,and TGF-β+DSS-H),respectively.The inter-vention concentrations of DSSS,FA,PF and TGF-β on NRK-52E cells were screened and determined for NRK-52E cells in the above groups.Western blotting was used to detect the changes in the protein levels of NLRP3,cysteine protease 1(CASP1),Gasdermin D(GSDMD),interleukin-18(IL-18),fibronectin(FN),E-Cadherin,and N-Cadherin in each group.Results Compared with the model group,the expression levels of pyroptosis-related proteins NLRP3,CASP1,GSDMD,and IL-18 decreased in the DSSS,FA,and PF groups,and the difference was statistically significant(all P<0.05).Compared with the model group,the expression levels of fibrosis-related adhesion proteins FN,E-Cadherin and N-Cadherin decreased in the medium-dose and high-dose DSSS and FA and PF groups,and the difference was statistically significant(all P<0.05).Conclusion DSS-medicated serum and its active ingredients,paeoniflorin and ferulic acid,can inhibit the activation of NLRP3-associated pyroptosis proteins in the TGF-β-induced fibrosis model of NRK-52E cells and regulate the expression of adhesion proteins and the progression of epithelial-mesenchymal transdifferentiation to allevi-ate the fibrosis.
万方数据
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