Reperfusion of rat heart after brief ischemia induces proteolysis of calspectin (nonerythroid spectrin or fodrin) by calpain.
Rat myocardium expresses the 240- and 235-kD polypeptides antigenically related to alpha- and beta-subunits of brain calspectin (nonerythroid spectrin or fodrin), respectively. In the subcellular fractions of the myocardium, alpha-calspectin was found in the 600g, 10,000g, and 100,000g pellets, whereas beta-calspectin was localized to the 10,000g pellet. On the basis of the Na+,K(+)-ATPase activity and the contents of a gap junction protein, the sarcolemma was distributed to the 10,000g and 100,000g pellets, and the intercalated disks were enriched in the 10,000g pellet. Both alpha- and beta-calspectin were proteolyzed by calpain in vitro. The two subunits were also proteolyzed in vivo, when the rat hearts underwent 10 to 60 minutes of global ischemia followed by 30 minutes of reperfusion. The reperfusion following the ischemia induced the proteolysis of alpha-calspectin in the 10,000g and 100,000g pellets, producing the 150-kD fragment. A synthetic calpain inhibitor, calpain inhibitor-1, suppressed the degradation of calspectin in vivo, which indicates that calpain is responsible for the reperfusion-induced proteolysis of calspectin. The inhibitor also improved myocardial stunning. Immunohistochemical study revealed that the proteolysis of alpha-calspectin occurs at the intercalated disks and the sarcolemma after postischemic reperfusion, in accord with the biochemical data. These results suggest that degradation of calspectin partly accounts for the contractile failure of the myocardium after postischemic reperfusion by disrupting the membrane skeleton and the intercalated disks.
Yoshida
K
K
Department of Legal Medicine, Yamaguchi University School of Medicine, Japan.
Inui
M
M
Harada
K
K
Saido
T C
TC
Sorimachi
Y
Y
Ishihara
T
T
Kawashima
S
S
Sobue
K
K
D000818:Animals; D002148:Calmodulin-Binding Proteins / Q000378:metabolism*; D002154:Calpain / Q000494:pharmacology*; D017202:Myocardial Ischemia / Q000378:metabolism*; D015425:Myocardial Reperfusion; D051381:Rats; D017208:Rats, Wistar
D002148:Calmodulin-Binding Proteins; D002154:Calpain