解放军医学杂志2024，Vol.49Issue(12)：1400-1407,8.DOI:10.11855/j.issn.0577-7402.0248.2024.0721
瞬时感受电位通道6在同型半胱氨酸诱导的小鼠肾脏足细胞自噬中的作用
Role of transient receptor potential channel 6 in homocysteine-induced podocyte autophagy of mouse kidney
摘要
Abstract
Objective To explore the regulatory role of transient receptor potential channel 6(TRPC6)on podocyte autophagy under the influence of homocysteine(Hcy)in mouse kidney.Methods Mouse renal podocytes were divided into control group and Hcy groups(stimulated by Hcy at 40,60,80 and 100 μmol/L for 48 h).The level of TRPC6 mRNA was assessed using quantitative reverse transcription polymerase chain reaction(qRT-PCR)to identify the optimal Hcy concentration for subsequent experiments.Western blotting was employed to evaluate the expression levels of autophagy-related proteins LC3 Ⅱ and p62,as well as the expression levels of podocyte structural proteins Nephrin and Podocin.The expression levels of TRPC6 mRNA and protein in both groups were determined using qRT-PCR,Western blotting and immunofluorescence.Transfections of cells with TRPC6 overexpression or interference were set as follows:(1)control group(untreated),negative control group of TRPC6 overexpression,and TRPC6 overexpression group;(2)control group(untreated),negative control group of TRPC6 interference,and TRPC6 interference group(si-1,si-2,si-3).The expression level of TRPC6 was detected using qRT-PCR.The cells after overexpressing or interfering of TRPC6 were further set as follows:(1)control group(untreated),Hcy group(80 μmol/L Hcy added),TRPC6 overexpression control+Hcy group,TRPC6 overexpression+Hcy group;(2)control group(untreated),Hcy group,TRPC6 interference control+Hcy group,and TRPC6 interference+Hcy group.The expression levels of p62,LC3 Ⅱ,and TRPC6 proteins were detected using Western blotting.Results qRT-PCR detection results showed that compared with control group,the expression level of TRPC6 mRNA in Hcy group increased with the increase of Hcy concentration,with the highest expression level observed at 80 μmol/L Hcy.Therefore,80 μmol/L Hcy was selected as the optimal concentration for intervention.At this time,the expression level of autophagy-related protein LC3 Ⅱ increased,and the expression level of p62 decreased(P<0.05).Western blotting results showed that compared with control group,the expression levels of podocyte-related proteins Nephrin and Podocin in Hcy group were significantly decreased(P<0.05).qRT-PCR results showed that compared with control group,the expression level of TRPC6 mRNA in Hcy group was significantly increased(P<0.05).Compared with negative control group for TRPC6 overexpression,both mRNA and protein expression levels of TRPC6 in TRPC6 overexpression group were significantly higher(P<0.05).Compared with negative control group for TRPC6 interference,both mRNA and protein expression levels of TRPC6 in TRPC6 interference group were significantly decreased(P<0.05).Western blotting results showed that compared with negative control group for TRPC6 overexpression,the expression level of autophagy-related protein LC3 Ⅱ in TRPC6 overexpression+Hcy group was significantly increased,and the expression level of p62 was significantly decreased(P<0.05).Compared with TRPC6 negative control+Hcy group for TRPC6 interference+Hcy,the expression level of autophagy-related protein LC3 Ⅱ in TRPC6 interference+Hcy group was significantly decreased,and the expression level of p62 was significantly increased(P<0.05).Conclusion Hcy can induce autophagy of renal podocytes.Inhibiting the expression of TRPC6 can significantly reduce the autophagy damage to podocytes.关键词
足细胞自噬/同型半胱氨酸/瞬时感受电位通道6Key words
podocyte autophagy/homocysteine/transient receptor potential channel 6分类
医药卫生引用本文复制引用
卢冠军,徐遥琴,石青,刘莉,张亚兰,白志刚,李淑娟,汪乐新,赵静,刘超,熊建团,焦运,杨安宁,姜怡邓,田宇佳..瞬时感受电位通道6在同型半胱氨酸诱导的小鼠肾脏足细胞自噬中的作用[J].解放军医学杂志,2024,49(12):1400-1407,8.基金项目
国家自然科学基金(82060139,81900273) （82060139,81900273）
宁夏回族自治区重点研发计划(2020BFH02003,2021BEG02033,2020BFH02001) （2020BFH02003,2021BEG02033,2020BFH02001）
中国医学科学院中央级公益性科研院所基本科研业务费项目(2019PT330002) This work was supported by the National Natural Science Foundation of China(82060139,81900273),the Key Research and Development Project of Ningxia Hui Autonomous Region(2020BFH02003,2021BEG02033,2020BFH02001),and the Basic Research Funding Project of Central Public Welfare Research Institutes,Chinese Academy of Medical Sciences(2019PT330002) （2019PT330002）
