Myocardial infarction-induced N-terminal fragment of cardiac myosin-binding protein C (cMyBP-C) impairs myofilament function in human myocardium.

Myocardial infarction (MI) is associated with depressed cardiac contractile function and progression to heart failure. Cardiac myosin-binding protein C, a cardiac-specific myofilament protein, is proteolyzed post-MI in humans, which results in an N-terminal fragment, C0-C1f. The presence of C0-C1f in cultured cardiomyocytes results in decreased Ca(2+) transients and cell shortening, abnormalities sufficient for the induction of heart failure in a mouse model. However, the underlying mechanisms remain unclear. Here, we investigate the association between C0-C1f and altered contractility in human cardiac myofilaments in vitro. To accomplish this, we generated recombinant human C0-C1f (hC0C1f) and incorporated it into permeabilized human left ventricular myocardium. Mechanical properties were studied at short (2 µm) and long (2.3 µm) sarcomere length (SL). Our data demonstrate that the presence of hC0C1f in the sarcomere had the greatest effect at short, but not long, SL, decreasing maximal force and myofilament Ca(2+) sensitivity. Moreover, hC0C1f led to increased cooperative activation, cross-bridge cycling kinetics, and tension cost, with greater effects at short SL. We further established that the effects of hC0C1f occur through direct interaction with actin and α-tropomyosin. Our data demonstrate that the presence of hC0C1f in the sarcomere is sufficient to induce depressed myofilament function and Ca(2+) sensitivity in otherwise healthy human donor myocardium. Decreased cardiac function post-MI may result, in part, from the ability of hC0C1f to bind actin and α-tropomyosin, suggesting that cleaved C0-C1f could act as a poison polypeptide and disrupt the interaction of native cardiac myosin-binding protein C with the thin filament.

Pathogenic properties of the N-terminal region of cardiac myosin binding protein-C in vitro.

Cardiac myosin binding protein-C (cMyBP-C) plays a role in sarcomeric structure and stability, as well as modulating heart muscle contraction. The 150 kDa full-length (FL) cMyBP-C has been shown to undergo proteolytic cleavage during ischemia-reperfusion injury, producing an N-terminal 40 kDa fragment (mass 29 kDa) that is predominantly associated with post-ischemic contractile dysfunction. Thus far, the pathogenic properties of such truncated cMyBP-C proteins have not been elucidated. In the present study, we hypothesized that the presence of these 40 kDa fragments is toxic to cardiomyocytes, compared to the 110 kDa C-terminal fragment and FL cMyBP-C. To test this hypothesis, we infected neonatal rat ventricular cardiomyocytes and adult rabbit ventricular cardiomyocytes with adenoviruses expressing the FL, 110 and 40 kDa fragments of cMyBP-C, and measured cytotoxicity, Ca(2+) transients, contractility, and protein-protein interactions. Here we show that expression of 40 kDa fragments in neonatal rat ventricular cardiomyocytes significantly increases LDH release and caspase 3 activity, significantly reduces cell viability, and impairs Ca(2+) handling. Adult cardiomyocytes expressing 40 kDa fragments exhibited similar impairment of Ca(2+) handling along with a significant reduction of sarcomere length shortening, relaxation velocity, and contraction velocity. Pull-down assays using recombinant proteins showed that the 40 kDa fragment binds significantly to sarcomeric actin, comparable to C0-C2 domains. In addition, we discovered several acetylation sites within the 40 kDa fragment that could potentially affect actomyosin function. Altogether, our data demonstrate that the 40 kDa cleavage fragments of cMyBP-C are toxic to cardiomyocytes and significantly impair contractility and Ca(2+) handling via inhibition of actomyosin function. By elucidating the deleterious effects of endogenously expressed cMyBP-C N-terminal fragments on sarcomere function, these data contribute to the understanding of contractile dysfunction following myocardial injury.

Lovastatin attenuates nerve injury in an animal model of Guillain-Barré syndrome.

Statins, widely used as clinically effective inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase, exhibit anti-inflammatory properties that may be of therapeutic benefit for the management of some neurological disorders. In this study, a short-term course of lovastatin treatment is shown to markedly inhibit the development of experimental autoimmune neuritis (EAN) in the absence of hepatotoxic or myotoxic complications. Independent of cholesterol reduction, lovastatin treatment prevented EAN-induced peripheral nerve conduction deficits and morphologic nerve injury. Co-administration with mevalonate neutralized the prophylactic effects of lovastatin. When administered therapeutically, lovastatin significantly shortened the disease course. Autoreactive immunity, measured in vitro by myelin-stimulated proliferation of splenocytes, was significantly diminished by in vivo lovastatin treatment. Th1-dominant immune responses, measured by cytokine profiling, however, were not affected by lovastatin. Sciatic nerves of lovastatin-treated immunized rats showed markedly reduced levels of cellular infiltrates. Treating peripheral nerve endothelial monolayers with lovastatin significantly inhibited the in vitro migration of autoreactive splenocytes. Together, these data demonstrate that a short-term course of lovastatin attenuates the development and progression of EAN in Lewis rats by limiting the proliferation and migration of autoreactive leukocytes.