Ohyanagi M, Yamamoto J, Nakamura K, Shibuya J, Morita M, Masutani M, Arii T, Iwasaki T
First Department of Internal Medicine, Hyogo College of Medicine, Nishinomiya, Japan.
J Mol Cell Cardiol. 1995 May;27(5):1131-9. doi: 10.1016/0022-2828(95)90049-7.
It has been reported that the function of the guanine-binding regulatory protein (G protein), especially the alpha subunit of the stimulatory G protein (Gs alpha), in myocardium is decreased with acute ischemia. However, it is unclear whether this decrease is due to transcriptional or post-transcriptional changes. Moreover, no studies have examined the distribution of G protein mRNA in ischemic myocardium using in situ hybridization. The purpose of this study was to explore alterations in mRNA of G proteins (Gs and Gi) in ischemic hearts using in situ hybridization. We measured the levels of mRNA for Gs alpha and Gi alpha in ischemic and non-ischemic myocardium by in situ hybridization using a radioisotope imaging system. We compare these mRNA levels in ischemic and non-ischemic myocardium with Northern blot analysis and the protein levels of G proteins by Western blot analysis. The mRNA for Gs alpha and Gi alpha was distributed diffusely in normal hearts. Levels of mRNA detected by in situ hybridization were substantially reduced by acute ischemia, and these results were confirmed by Northern and Western blot analysis. These results suggest that decreased levels of mRNA and protein for G proteins may underlie the impaired function of the receptor--G protein--adenylate cyclase system in ischemic myocardium. In addition, quantitative evaluation of mRNA is possible by in situ hybridization and correlates well with Northern analysis.
据报道，急性缺血时心肌中鸟嘌呤结合调节蛋白（G蛋白）的功能，尤其是刺激性G蛋白（Gsα）的α亚基功能会降低。然而，尚不清楚这种降低是由于转录变化还是转录后变化所致。此外，尚无研究使用原位杂交技术检测缺血心肌中G蛋白mRNA的分布情况。本研究的目的是利用原位杂交技术探究缺血心脏中G蛋白（Gs和Gi）mRNA的变化。我们使用放射性同位素成像系统，通过原位杂交测定缺血和非缺血心肌中Gsα和Giα的mRNA水平。我们通过Northern印迹分析比较缺血和非缺血心肌中的这些mRNA水平，并通过Western印迹分析比较G蛋白的蛋白质水平。Gsα和Giα的mRNA在正常心脏中呈弥漫性分布。急性缺血使原位杂交检测到的mRNA水平大幅降低，Northern和Western印迹分析证实了这些结果。这些结果表明，G蛋白mRNA和蛋白质水平的降低可能是缺血心肌中受体-G蛋白-腺苷酸环化酶系统功能受损的基础。此外，通过原位杂交可以对mRNA进行定量评估，且与Northern分析结果相关性良好。
