Misquitta C M, Samson S E, Grover A K
Department of Biomedical Sciences, HSC 4N41, McMaster University, Hamilton, Ontario, Canada.
Mol Cell Biochem. 1996 May 10;158(1):91-5. doi: 10.1007/BF00225887.
Densities of sarcoplasmic reticulum (SR) Ca(2+)-pump were compared in proximal and distal segments of pig left coronary artery using two biochemical methods: acylphosphate formation and immunoreactivity in Western blots, and a functional assay based on contraction to SR Ca(2+)-pump inhibitors. In the microsomes prepared from smooth muscle, the level of the 115 kDa SR Ca(2+)-pump acylphosphate was 7.1 +/- 0.3 -fold greater in distal than in proximal segments. Similarly in Western blots using these microsomes, the reactivity of the 115 kDa band to an anti-SR Ca(2+)-pump antibody was 5.3 +/- 0.8 -fold greater in distal than in proximal segments. Endothelium free coronary artery rings contracted to the SR Ca(2+)-pump inhibitors Cyclopiazonic acid (CPA, EC50 = 0.19 +/- 0.06 microM) and thapsigargin (EC50 = 0.0095 +/- 0.0035 microM). With 10 microM CPA, the force of contraction per tissue wet weight was 4.2 +/- 0.5 -fold greater in distal than in proximal rings, and with 1 microM thapsigargin it was 4.0 +/- 1.0 -fold greater. The contractions produced by 60 mM KCl were used as a control. In contrast to the CPA and thapsigargin, the force per mg tissue weight produced by 60 mM KCl did not differ significantly between the proximal and distal segments. Thus, the results from the two biochemical methods and those from the contractility data were all consistent with the smooth muscle in the distal segments of the coronary artery containing a higher density of the SR Ca(2+)-pump than the proximal segments.
使用两种生化方法比较猪左冠状动脉近端和远端节段肌浆网（SR）Ca(2+)泵的密度：酰基磷酸形成和蛋白质免疫印迹中的免疫反应性，以及基于对SR Ca(2+)泵抑制剂收缩反应的功能测定。在平滑肌制备的微粒体中，115 kDa SR Ca(2+)泵酰基磷酸水平在远端节段比近端节段高7.1±0.3倍。同样，在使用这些微粒体的蛋白质免疫印迹中，115 kDa条带与抗SR Ca(2+)泵抗体的反应性在远端节段比近端节段高5.3±0.8倍。无内皮冠状动脉环对SR Ca(2+)泵抑制剂环匹阿尼酸（CPA，EC50 = 0.19±0.06 microM）和毒胡萝卜素（EC50 = 0.0095±0.0035 microM）产生收缩反应。使用10 microM CPA时，每组织湿重的收缩力在远端环比近端环高4.2±0.5倍，使用1 microM毒胡萝卜素时高4.0±1.0倍。用60 mM KCl产生的收缩作为对照。与CPA和毒胡萝卜素不同，60 mM KCl产生的每毫克组织重量的力在近端和远端节段之间没有显著差异。因此，两种生化方法的结果以及收缩性数据的结果均与冠状动脉远端节段平滑肌中SR Ca(2+)泵密度高于近端节段一致。
