Proc Natl Acad Sci U S A. 2002 Apr 30;99(9):6252-6. Epub 2002 Apr 23.

Functional consequences of caspase activation in cardiac myocytes.

Communal C, Sumandea M, de Tombe P, Narula J, Solaro RJ, Hajjar RJ.

Source

Cardiovascular Research Center, Massachusetts General Hospital, Charlestown, MA 02129, USA.

Abstract

Cardiomyocyte apoptosis is present in many cardiac disease states, including heart failure and ischemic heart disease. Apoptosis is associated with the activation of caspases that mediate the cleavage of vital and structural proteins. However, the functional contribution of apoptosis to these conditions is not known. Furthermore, in cardiac myocytes, apoptosis may not be complete, allowing the cells to persist for a prolonged period within the myocardium. Therefore, we examined whether caspase-3 cleaved cardiac myofibrillar proteins and, if so, whether it affects contractile function. The effects of caspase-3 were studied in vitro on individual components of the cardiac myofilament including alpha-actin, alpha-actinin, myosin heavy chain, myosin light chain 1/2, tropomyosin, cardiac troponins (T, I, C), and the trimeric troponin complex. Exposure of the myofibrillar protein (listed above) to caspase-3 for 4 h resulted in the cleavage of alpha-actin and alpha-actinin, but not myosin heavy chain, myosin light chain 1/2, and tropomyosin, into three fragments (30, 20, and 15 kDa) and one major fragment (45 kDa), respectively. When cTnT, cTnI, and cTnC were incubated individually with caspase-3, there was no detectable cleavage. However, when the recombinant troponin complex was exposed to caspase-3, cTnT was cleaved, resulting in fragments of 25 kDa. Furthermore, rat cardiac myofilaments exposed to caspase-3 exhibited similar patterns of myofibrillar protein cleavage. Treatment with the caspase inhibitor DEVD-CHO or z-VAD-fmk abolished the cleavage. Myofilaments, isolated from adult rat ventricular myocytes after induction of apoptotic pathway by using beta-adrenergic stimulation, displayed a similar pattern of actin and TnT cleavage. Exposure of skinned fiber to caspase-3 decreased maximal Ca(2+)-activated force and myofibrillar ATPase activity. Our results indicate that caspase-3 cleaved myofibrillar proteins, resulting in an impaired force/Ca(2+) relationship and myofibrillar ATPase activity. Induction of apoptosis in cardiac cells was associated with similar cleavage of myofilaments. Therefore, activation of apoptotic pathways may lead to contractile dysfunction before cell death.

PMID:: 11972044; [PubMed - indexed for MEDLINE]
PMCID:: PMC122935

Free PMC Article

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Figure 6

Normalized pCa tension and pCa-ATPase relation in detergent-skinned fiber bundle were measured after caspase-3 treatment. Resting tension was fixed at 2.3 μm. (a) pCa tension of untreated fiber at t = 0 (solid line) and 45 min (dashed line); maximum tensions at pCa 4.5 at t = 0 and t = 45 min were 39.6 ± 4.4 mN/mm² and 34.0 ± 4.0 mN/mm², respectively. (b) pCa-ATPase of untreated fiber at t = 0 and 45 min; maximum ATP consumptions at pCa 4.5 at t = 0 and t = 45 min were 335 ± 29 pmol/μl per sec and 289 ± 24 pmol/μl per sec, respectively. (c) pCa-tension of fibers after t = 0 and 45 min of caspase-3 treatment; maximum tensions at pCa 4.5 at t = 0 and t = 45 min were 37.6 ± 5.0 mN/mm² and 16.6 ± 2.7 mN/mm², respectively. (d) pCa-ATPase of fibers after t = 0 and 45 min of caspase-3 treatment; maximum ATP consumptions at pCa 4.5 at t = 0 and t = 45 min were 297 ± 15 pmol/μl per sec and 149 ± 15 pmol/μl per sec, respectively. (e) pCa-tension of fibers after t = 0 and 45 min of caspase-3 and DEVD-CHO (10 μM) treatments; maximum tensions at pCa 4.5 at t = 0 and t = 45 min were 36.6 ± 6.9 mN/mm² and 27.2.0 ± 3.2 mN/mm², respectively. (f) pCa-ATPase of fibers after t = 0 and 45 min of caspase-3 and DEVD-CHO treatments; maximum ATP consumptions at pCa 4.5 at t = 0 and t = 45 min were 312 ± 8 pmol/μl per sec and 273 ± 3 pmol/μl per sec, respectively. Graph represents mean ± SE of percent change from t = 0 min from three to four fibers, from three to four different hearts for each case. After 45 min, caspase-3 treatment decreases maximal Ca²⁺-activated tension as compared with sham (respectively, at pCa 4.5, caspase-3 = 39.6 ± 6.3%* vs. Sham = 82.9 ± 0.7%, P < 0.01, n = 3). Furthermore, after 45 min, caspase-3 treatment decreases maximal Ca²⁺-activated ATPase as compared with sham (respectively, caspase-3 = 48.4 ± 6.0%* vs. Sham = 77.9 ± 2.0%, P < 0.05, n = 3). However, there was no change in pCa50.

Functional consequences of caspase activation in cardiac myocytes

Proc Natl Acad Sci U S A. 2002 April 30;99(9):6252-6256.