Ischemia/reperfusion-induced changes in cardiac contractile proteins

Abstract

Heart disease, often results in the development of ischemia/reperfusion (I/R) injury, and is one of the major causes of morbidity and mortality. I/R injury is usually characterized by the development of cardiac contractile dysfunction, which is one of the major indicators of recovery. The understanding of the mechanisms underlying the development cardiac contractile dysfunction in response to I/R is crucial for the prevention and treatment of I/R injury. Using a proteomics approach to an isolated heart model we identified myosin light chain 1 (MLC1) as an important contractile protein, phosphorylated by myosin light chain kinase (MLCK) and degraded by matrix metalloproteinase-2 (MMP-2) due to I/R. Prevention of MLC1 phosphorylation, with an MLCK inhibitor (ML-7) or a myosin light chain phosphatase – MLCP – stimulator (Y-27632), and degradation with an MMPs inhibitor (doxycycline) protects the heart from contractile dysfunction, associated with a preservation of MLC1 protein levels. The combined use of these three drugs (ML-7, Y-27632 and doxycycline) at subthreshold concentrations resulted in a full protection of both contractile function and MLC1 in hearts subjected to I/R. The MLCP indirect activator Y-27632 (a Rho kinase inhibitor) showed the best degree of protection of contractile function at full protective doses, independent of MLC1 preservation. A proteomics approach revealed that the administration of Y-27632 to hearts subjected to I/R results, not only in a protection of MLC1 from phosphorylation and degradation, but also a modulation of enzymes involved in energy production (lactate dehydrogenase and glyceraldehyde 3-phosphate dehydrogenase). This modulation of energy metabolism is potentially an importa effect of Rho kinase inhibitors, secondary to direct protection of cardiac contractile proteins from degradation in response to I/R. The data presented here show a novel paradigm in the development of I/R-induced cardiac contractile dysfunction. Moreover, we establish a new therapeutic approach (multidrug treatment) that can be of crucial importance in the development of new preventive or treatment strategies against cardiac injury.