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Qualitative level of GTP-binding proteins in diabetic rat myocardium and its biological significance in alteration of beta-adrenoceptor mediated cellular response.

糖尿病大鼠心肌中 GTP 结合蛋白的定性水平及其在改变 β-肾上腺素受体介导的细胞反应中的生物学意义。

基本信息

批准号：
08670098
负责人：
KANNO Morio
金额：
$ 1.41万
依托单位：
Hokkaido University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1996
资助国家：
日本
起止时间：
1996 至 1997
项目状态：
已结题

项目摘要

This research project has aimed to clarify which of ADP-ribosylation method or immunoblotting method is adequate for estimating GTP binding proteins (G) in diabetic rat myocardium. When estimated by ADP-ribosylation method, we found increased level of both Gs and Gi, but immunoblotting with anti-Gs alpha-and Gi alpha-subunit antiserum indicated decreased level of Gi and unchanged level of Gs. In diabetic rat myocardium, we observed : 1. augmented activation of adenylate cyclase (AC) , when stimulated by beta-adrenoceptors (R) , GppNHp, NaF and forskolin in comparison with the AC activities in control normal rat myocardium, 2. decreased level of GimRNA and unchanged level of Gs mRNA level. Under the hypothesis that the decreased level of Gi protein which gives tonic inhibitory influence on Gs-AC system, causes the alterations of AC activities in diabetic state, we examined influence of functional abolishment of Gi protein on R-Gs-AC system in PTX-treated rabbit myocardium. we found that PTX-treated myocardium produced augmentation of AC activities in response to beta-adrenoceptor stimulation and GppNHp, but, not to NaF and forskolin. Thus, even though there are a few discrepancy, the altered response of AC activities in diabetic myocardium is well explained by the decreased level of Gi. From these results, we conclude that G protein level estimated by immnoblotting method reflects well alterations in R-Gs-AC coupling in diabetic myocardium.

本课题旨在阐明adp -核糖基化法和免疫印迹法哪一种适合估测糖尿病大鼠心肌中GTP结合蛋白(G)。用adp -核糖基化法测定时，我们发现Gs和Gi水平均升高，但用抗Gs α和Gi α亚单位抗血清免疫印迹显示Gi水平降低，Gs水平不变。在糖尿病大鼠心肌中，我们观察到：β -肾上腺素受体(R)、GppNHp、NaF和forskolin刺激时腺苷酸环化酶（AC）的激活与正常对照大鼠心肌AC活性的增强，2。GimRNA水平降低，Gs mRNA水平不变。假设Gi蛋白水平降低，对Gs-AC系统产生强直抑制作用，导致糖尿病状态下AC活性的改变，我们研究了Gi蛋白功能消除对ptx治疗兔心肌R-Gs-AC系统的影响。我们发现，ptx处理的心肌在β -肾上腺素受体刺激和GppNHp的作用下产生AC活性的增强，但对NaF和forskolin没有反应。因此，尽管存在一些差异，但糖尿病心肌中AC活性的改变很好地解释了Gi水平的降低。从这些结果，我们得出结论，免疫印迹法估计的G蛋白水平很好地反映了糖尿病心肌R-Gs-AC偶联的变化。