Reduced sarcoplasmic reticulum Ca(2+)-uptake and expression of phospholamban in left ventricular myocardium of dogs with heart failure.
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.
Gupta
R C
RC
Department of Medicine, Division of Cardiovascular Medicine, Detroit, Michigan 48202, USA.
Mishra
S
S
Mishima
T
T
Goldstein
S
S
Sabbah
H N
HN
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics; D000252:Calcium-Transporting ATPases / Q000096:biosynthesis / Q000235:genetics; D002155:Calsequestrin / Q000096:biosynthesis / Q000235:genetics; D004285:Dogs; D005260:Female; D006333:Heart Failure / Q000378:metabolism*; D006352:Heart Ventricles / Q000378:metabolism; D008297:Male; D009206:Myocardium / Q000378:metabolism*; D012333:RNA, Messenger; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*; D000099211:Phospholamban
D002118:Calcium; D002135:Calcium-Binding Proteins; D000252:Calcium-Transporting ATPases; D002155:Calsequestrin; D012333:RNA, Messenger; D000099211:Phospholamban