Anthracyclines induce calpain-dependent titin proteolysis and necrosis in cardiomyocytes.

Lim CC, Zuppinger C, Guo X, Kuster GM, Helmes M, Eppenberger HM, Suter TM, Liao R, Sawyer DB
J Biol Chem. 2004 279 (9): 8290-9

PMID: 14676206 · DOI:10.1074/jbc.M308033200

Titin, the largest myofilament protein, serves as a template for sarcomere assembly and acts as a molecular spring to contribute to diastolic function. Titin is known to be extremely susceptible to calcium-dependent protease degradation in vitro. We hypothesized that titin degradation is an early event in doxorubicin-induced cardiac injury and that titin degradation occurs by activation of the calcium-dependent proteases, the calpains. Treatment of cultured adult rat cardiomyocytes with 1 or 3 micromol/liter doxorubicin for 24 h resulted in degradation of titin in myocyte lysates, which was confirmed by a reduction in immunostaining of an antibody to the spring-like (PEVK) domain of titin at the I-band of the sarcomere. The elastic domain of titin appears to be most susceptible to proteolysis because co-immunostaining with an antibody to titin at the M-line was preserved, suggesting targeted proteolysis of the spring-like domain of titin. Doxorubicin treatment for 1 h resulted in approximately 3-fold increase in calpain activity, which remained elevated at 48 h. Co-treatment with calpain inhibitors resulted in preservation of titin, reduction in myofibrillar disarray, and attenuation of cardiomyocyte necrosis but not apoptosis. Co-treatment with a caspase inhibitor did not prevent the degradation of titin, which precludes caspase-3 as an early mechanism of titin proteolysis. We conclude that calpain activation is an early event after doxorubicin treatment in cardiomyocytes and appears to target the degradation of titin. Proteolysis of the spring-like domain of titin may predispose cardiomyocytes to diastolic dysfunction, myofilament instability, and cell death by necrosis.

MeSH Terms (22)

Animals Anthracyclines Antibiotics, Antineoplastic Apoptosis Calpain Caspase 3 Caspase Inhibitors Caspases Cells, Cultured Connectin Doxorubicin Enzyme Activation Enzyme Inhibitors Heart Ventricles Male Muscle Cells Muscle Proteins Myocardium Necrosis Protein Kinases Rats Rats, Sprague-Dawley

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