福建中医药2025，Vol.56Issue(8)：12-20,9.DOI:10.13260/j.cnki.jfjtcm.2025.08003
钩藤碱通过调控STAT3信号通路抑制血管紧张素Ⅱ诱导的心肌细胞肥大
Rhynchophylline Inhibits AngiotensinⅡ-Induced Cardiomyocyte Hypertrophy by Regulating STAT3 Signaling Pathway
摘要
Abstract
Objective:To investigate the mechanism of Rhynchophylline(Rhy)in inhibiting angiotensin Ⅱ(AngⅡ)-induced cardiomyocyte hypertrophy by regulating the signal transduction and transcription activator 3(STAT3)signaling pathway.Methods:Neonatal rat cardiomyocytes(NRCMs)were extracted from the heart tissue of 1-to 2-day-old newborn SD rats.The CCK-8 method was used to detect the cell viability of NRCMs intervened with 0,2,10,20,40,and 80 μmol/L Rhy,and the results showed that 40 μmol/L Rhy was the optimal intervention concentration.Western blot was used to detect the effects of different stimulation times(0,0.5,1,2,4,6,12,24 h)of AngⅡ on the nuclear and total protein expression levels of NRCMs STAT3 and pSTAT3.AutoDock Vina V1.2.x software was used for molecular docking to confirm the binding affinity between Rhy and STAT3.NRCMs were divided into 4 groups:1)the control group was cultured in normal medium;2)the AngⅡ group was cultured in medium containing 1 μmol/L AngⅡ;3)the Rhy group was cultured in medium containing 40 μmol/L Rhy;4)the AngⅡ+Rhy group was cultured in medium containing 1 μmol/L AngⅡ and 40 μmol/L Rhy.A separate experiment of NRCMs was divided into 5 groups:1)the control group was cultured in normal medium;2)the AngⅡ group was cultured in medium containing 1 μmol/L AngⅡ;3)the AngⅡ+Rhy group was cultured in medium containing 1 μmol/L AngⅡ and 40 μmol/L Rhy;4)the AngⅡ+S3I-201 group was cultured in medium con-taining 1 μmol/L AngⅡ and 10 μmol/L S3I-201;5)the AngⅡ+S3I-201+Rhy group was cultured in medium containing 1 μmol/L AngⅡ,40 μmol/L Rhy,and 10 μmol/L S3I-201.Immunofluorescence was applied to detect the expression of cardiac troponin T(cTnT)protein;qPCR was applied to detect the relative mRNA expression levels of atrial natriuretic peptide(ANP),brain natriuretic peptide(BNP),Collagen Ⅰ,and transforming growth factor-β1(TGF-β1);Western blot was applied to detect the nuclear protein expression levels of pSTAT3 and STAT3,as well as the total protein expression levels of p-STAT3,STAT3,and TGF-β1.Results:Intervention of NRCMs with AngⅡ at different time showed that,compared with the 0 h group,the expression levels of nuclear pro-tein p-STAT3 increased significantly in the 6,12,and 24 h groups(P<0.05);the nuclear protein expression levels of STAT3 increased significantly in the 2,4,6,and 12 h groups(P<0.05);the total protein expression levels of p-STAT3 increased significantly in the 4,6,12,and 24 h groups(P<0.05).Molecular docking results showed that the binding energy of the optimal conformation between STAT3 and Rhy was-7.3 kcal/mol.This conformation contained three hydrogen bond binding sites:arginine 518,serine 521,and glu-tamic acid 524.The hydrogen bond length with serine 521 was 2.2 Å,which falls within the range of strong hydrogen bonds(2.2-2.5 Å)and may significantly reduce the dissociation rate of the complex.In the 4-group experiment:compared with the control group,the AngⅡ group showed a significant increase in immunofluorescence area(P<0.05),a significant increase in mRNA expression lev-els of ANP,BNP,Collagen Ⅰ,and TGF-β1(P<0.05),a significant increase in nuclear protein expression levels of p-STAT3 and STAT3(P<0.05),and a significant increase in total protein expression levels of p-STAT3 and TGF-β1(P<0.05).No statistically sig-nificant differences were found in the Rhy group compared to the control group regarding immunofluorescence area,mRNA expres-sion levels of ANP,BNP,Collagen Ⅰ,and TGF-β1,nuclear protein expression levels of p-STAT3 and STAT3,and total protein ex-pression levels of p-STAT3 and TGF-β1(P>0.05).Compared with the AngⅡ group,the AngⅡ+Rhy group showed a significant re-duction in immunofluorescence area(P<0.05),a significant decrease in mRNA expression levels of ANP,BNP,Collagen Ⅰ,and TGF-β1(P<0.05),a significant decrease in nuclear protein expression levels of p-STAT3 and STAT3(P<0.05),and a significant de-crease in total protein expression levels of p-STAT3 and TGF-β1(P<0.05).In the 5-group experiment:compared with the control group,the AngⅡ group showed a significant increase in immunofluorescence area(P<0.05),a significant increase in mRNA expres-sion levels of ANP,BNP,Collagen Ⅰ,and TGF-β1(P<0.05),a significant increase in nuclear protein expression levels of p-STAT3 and STAT3(P<0.05),and a significant increase in total protein expression levels of p-STAT3 and TGF-β1(P<0.05).Compared with the AngⅡ group,the AngⅡ+Rhy,AngⅡ+S3I-201,and AngⅡ+S3I-201+Rhy groups all showed a significant reduction in im-munofluorescence area(P<0.05),a significant decrease in mRNA expression levels of ANP,BNP,Collagen Ⅰ,and TGF-β1(P<0.05),a significant decrease in nuclear protein expression levels of p-STAT3 and STAT3(P<0.05),and a significant decrease in total protein expression levels of p-STAT3 and TGF-β1(P<0.05).Compared with the AngⅡ+S3I-201 group,the AngⅡ+S3I-201+Rhy group showed no statistically significant differences in immunofluorescence area,mRNA expression levels of ANP,BNP,Colla-gen Ⅰ,and TGF-β1,nuclear protein expression levels of p-STAT3 and STAT3,and total protein expression levels of p-STAT3 and TGF-β1(P>0.05).Conclusion:Rhy can prevent AngⅡ-induced hypertrophy in NRCMs by inhibiting the STAT3 signaling pathway.关键词
心肌细胞肥大/钩藤碱/血管紧张素Ⅱ/信号转导与转录激活因子3/心肌细胞Key words
cardiomyocyte hypertrophy/Rhynchophylline angiotensin Ⅱ/signal transducer and activator of transcription 3/car-diomyocyte引用本文复制引用
龚雨航,马恩,陈进晓,沃达,任丹妮..钩藤碱通过调控STAT3信号通路抑制血管紧张素Ⅱ诱导的心肌细胞肥大[J].福建中医药,2025,56(8):12-20,9.基金项目
福建中医药大学青年科技创新培育计划项目(XQB202201) （XQB202201）
福建中医药大学高层次人才科研项目(X2024001-人才,X2024009-人才) （X2024001-人才,X2024009-人才）
