Effect and mechanism of brain natriuretic peptide on aldosterone induced expression of natriuretic peptide receptor-A in renal tubular epithelial cells
Effect and mechanism of brain natriuretic peptide on aldosterone induced expression of natriuretic peptide receptor-A in renal tubular epithelial cells
李彤 1呼海娟 1刘德敏 1崔炜1
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作者信息
1. 050000 石家庄,河北医科大学第二医院心内一科 河北省心脑血管病研究所
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摘要
目的 研究B型利钠肽(BNP)对醛固酮(Ald)诱导的肾小管上皮细胞(RTEC)中利钠肽受体A(NPR-A)表达的影响,探寻肾脏发生BNP 抵抗的机制.方法 将正常状态的RTEC按BNP刺激浓度依次分为 10-7 M BNP组、10-6 M BNP组和 10-5 M BNP 组;将Ald诱导的RTEC按BNP刺激浓度依次分为Ald+10-7 M BNP组、Ald+10-6 M BNP组和Ald+10-5 M BNP组.通过实时定量聚合酶链反应检测RTEC上NPR-A的转录水平,蛋白质印迹法、免疫荧光染色技术及质膜蛋白分离技术检测NPR-A的蛋白表达水平,酶联免疫吸附法检测环磷酸鸟苷(cGMP)含量.结果 在正常RTEC中,随着BNP刺激浓度的升高,NPR-A的mRNA含量、蛋白表达量及cGMP含量均呈先增加后减少的趋势,高浓度BNP 会抑制NPR-A的基因转录和蛋白表达,降低cGMP 含量.在Ald诱导的RTEC中,随着BNP刺激浓度的升高,NPR-A的mRNA含量、蛋白表达量及cGMP含量均在轻微增加后急剧减少,高浓度BNP会明显抑制NPR-A的基因转录和蛋白表达,减少cGMP生成.当受到相同浓度BNP刺激时,Ald诱导的RTEC上NPR-A的转录水平及cGMP生成量均低于相应的正常RTEC.结论 在正常/Ald诱导的RTEC中,胞膜和胞质上NPR-A的蛋白表达水平的变化趋势与总蛋白水平相一致,即呈先升后降的动态变化特点,高浓度BNP会下调NPR-A水平,减少cGMP 生成.心力衰竭时发生的BNP抵抗现象与肾脏 NPR-A的变化密切相关,RTEC纤维化可能是 BNP 抵抗的机制之一.
Abstract
Objective To investigate the effect of brain natriuretic peptide(BNP)on the aldosterone(Ald)induced expression of natriuretic peptide receptor-A(NPR-A)in renal tubular epithelial cells(RTEC),and explore the mechanism of BNP resistance in the kidney.Methods RTEC in normal state was divided into 10-7 M BNP group,10-6 M BNP group and 10-5 M BNP group according to BNP stimulation concentration.Ald induced RTEC was divided into Ald+10-7 M BNP group,Ald+10-6 M BNP group and Ald+10-5 M BNP group according to BNP stimulation concentration.The transcription level of NPR-A on RTEC was detected by real-time quantitative polymerase chain reaction,the protein expression level of NPR-A was detected by western blot,immunofluorescence staining and plasma membrane protein separation techniques,and the cyclic guanosine monophosphate(cGMP)content was detected by enzyme-linked immunosorption assay.Results In normal RTEC,with the increase of BNP stimulation concentration,the mRNA content,protein expression level and cGMP content of NPR-A increased at the beginning and then decreased.High concentration of BNP inhibited gene transcription and protein expression of NPR-A and reduced cGMP content.In Ald induced RTEC,with the increase of BNP stimulation concentration,the mRNA content,protein expression and cGMP content of NPR-A decreased sharply after a slight increase.High concentration of BNP significantly inhibited gene transcription and protein expression of NPR-A and reduced the production of cGMP.When stimulated by the same concentration of BNP,the transcription level of NPR-A and the production of cGMP in Ald induced RTEC were lower than those in the corresponding normal RTEC group.Conclusions In normal/Ald induced RTEC,the change trend of NPR-A protein expression in the membrane and cytoplasm is consistent with that of total protein level,that is,the dynamic change characteristic is first increased and then decreased.High concentration of BNP could lower NPR-A level and reduce cGMP production.The phenomenon of BNP resistance in heart failure is closely related to the changes of kidney NPR-A,and RTEC fibrosis may be one of the mechanisms of BNP resistance.
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