Altered expression of high-molecular-weight calmodulin-binding protein in human ischaemic myocardium

A high-molecular-weight calmodulin-binding protein (HMWCaMBP) was previousl y identified and purified from the cytosolic fraction of bovine heart. Ease d on the sequence homology, amino acid analysis, antibody reactivity, and c alpain inhibition, HMWCaMBP has been identified as a homologue of the calpa in inhibitor calpastatin. In the present study the expression of HMWCaMBP w as investigated in normal and ischaemic human myocardium. Western blot anal ysis of normal human cardiac muscle extract with the polyclonal antibody ra ised against bovine HMWCaMBP indicated a prominent immunoreactive band with a molecular mass of 140 kD. HMWCaMBP was localized in the cytoplasm and my ofilaments of cardiac myocytes. Furthermore, Western blot analysis of norma l and ischaemic cardiac tissues indicated a decrease in the expression of H MWCaMBP in ischaemic tissues. These studies were further substantiated by i mmunohistochemical studies, indicating strong to moderate HMWCaMBP immunore activity in normal cardiac muscle and poor to negative immunoreactivity in ischaemic muscle. The results obtained from the rat ischaemic model suggest ed that the expression of cardiac HMWCaMBP was significantly decreased duri ng ischaemia/reperfusion. In addition, mu-calpain and m-calpain expression was higher in ischaemic cardiac tissue samples than in normal controls. The calpain inhibitory activity of ischaemic cardiac tissues was significantly lower than normal cardiac tissue samples. Tn some cases of cardiac ischaem ia, HMWCaMBP highlighted the contraction band necrosis seen at the margins of a myocardial infarct. In vitro, NMWCaMBP was proteolysed by mu-calpain a nd m-calpain. These results indicate that HMWCaMBP could be susceptible to proteolysis by calpains during ischaemia or reperfusion and may play a cont ributory role in myocardial injury. Copyright (C) 2000 John Wiley & Sons, L td.