心力衰竭犬左心室心肌肌浆网钙摄取减少及受磷蛋白表达降低。

Reduced sarcoplasmic reticulum Ca(2+)-uptake and expression of phospholamban in left ventricular myocardium of dogs with heart failure.

作者信息

Gupta R C, Mishra S, Mishima T, Goldstein S, Sabbah H N

机构信息

Department of Medicine, Division of Cardiovascular Medicine, Detroit, Michigan 48202, USA.

出版信息

J Mol Cell Cardiol. 1999 Jul;31(7):1381-9. doi: 10.1006/jmcc.1999.0970.

DOI:10.1006/jmcc.1999.0970

PMID:10403755

Abstract

The purpose of this study was to examine the sarcoplasmic reticulum (SR) Ca(2+)-uptake and the expression of phospholamban (PLB) and Ca(2+)-ATPase (CAA) in left ventricular (LV) and right ventricular (RV) myocardium of 6 normal (NL) dogs and 6 dogs with chronic heart failure (HF). In addition, gene expression of PLB and CAA was also examined in LV myocardium of NL and HF dogs. HF (LV ejection fraction 23+/-2%) was produced by multiple sequential intracoronary microembolizations. Oxalate-dependent Ca(2+)-uptake was measured in isolated membrane vesicles. Using specific dog myocardial monoclonal antibody, the expression of CAA, PLB and calsequestrin (CSQ) were measured in sodium dodecyl sulfate extract prepared from LV and RV tissue. Steady-state mRNA levels were determined by Northern hybridization using specific cDNA clones of PLB, CAA, CSQ, and glyceraldehyde-3-phosphate dehydrogenase (GADPH), a house keeping gene. SR Ca(2+)-uptake of NL and HF dogs increased with increasing Ca(2+)concentrations and reached a plateau at 3 microm in both LV and RV. Total capacity (134+/-9 v 224+/-10 nmol(45)Ca/mg protein/10 min, P<0.05) and maximal velocity (15+/-2 v 2 nmol(45)Ca/mg protein/min, P<0.05) of the SR to sequester Ca(2+)was significantly lower in LV myocardium of HF dogs compared to NL, whereas the Hill coefficient and the affinity of the Ca(2+)-pump for Ca(2+)were unchanged. LV tissue levels of the PLB and CAA, normalized to noncollagen protein or to CSQ and the PLB and CAA mRNA levels, normalized to CSQ or GADPH mRNA, were also significantly lower in HF dogs compared to NL. In RV myocardial tissue, no significant differences in total capacity of SR to sequester Ca(2+), maximal velocity of SR Ca(2+)-uptake, the affinity and Hill Coefficient of the Ca(2+)-pump for Ca(2+), or tissue levels of PLB and CAA were observed between NL dogs compared to HF dogs. We conclude that SR Ca(2+)-uptake and SR PLB and CAA protein and gene expression levels are reduced in LV myocardium of dogs with chronic HF. These abnormalities can lead to Ca(2+)-overload and subsequent global LV dysfunction.

摘要

本研究旨在检测6只正常（NL）犬和6只慢性心力衰竭（HF）犬左心室（LV）和右心室（RV）心肌中肌浆网（SR）的Ca(2+)摄取以及受磷蛋白（PLB）和Ca(2+)-ATP酶（CAA）的表达。此外，还检测了NL犬和HF犬LV心肌中PLB和CAA的基因表达。通过多次连续冠状动脉内微栓塞制备HF（LV射血分数23±2%）模型。在分离的膜囊泡中测量草酸盐依赖性Ca(2+)摄取。使用特异性犬心肌单克隆抗体，在从LV和RV组织制备的十二烷基硫酸钠提取物中测量CAA、PLB和肌集钙蛋白（CSQ）的表达。使用PLB、CAA、CSQ和甘油醛-3-磷酸脱氢酶（GADPH，一种管家基因）的特异性cDNA克隆，通过Northern杂交测定稳态mRNA水平。NL犬和HF犬的SR Ca(2+)摄取均随Ca(2+)浓度增加而增加，在LV和RV中均在3微摩尔时达到平台期。与NL犬相比，HF犬LV心肌中SR摄取Ca(2+)的总容量（134±9对224±10纳摩尔(45)Ca/毫克蛋白/10分钟，P<0.05）和最大速度（15±2对2纳摩尔(45)Ca/毫克蛋白/分钟，P<0.05）显著降低，而Hill系数和Ca(2+)泵对Ca(2+)的亲和力未改变。与NL犬相比，HF犬中以非胶原蛋白或CSQ标准化的PLB和CAA的LV组织水平以及以CSQ或GADPH mRNA标准化的PLB和CAA mRNA水平也显著降低。在RV心肌组织中，与HF犬相比，NL犬在SR摄取Ca(2+)的总容量、SR Ca(2+)摄取的最大速度、Ca(2+)泵对Ca(2+)的亲和力和Hill系数或PLB和CAA的组织水平方面未观察到显著差异。我们得出结论，慢性HF犬LV心肌中的SR Ca(2+)摄取以及SR PLB和CAA蛋白及基因表达水平降低。这些异常可导致Ca(2+)超载及随后的全心LV功能障碍。