Iwona Bil-Lula

Background: The fact that during myocardial ischemia/reperfusion (I/R) injury, myosin light chain 1 (MLC1) and troponin I (TnI) are degraded by matrix metalloproteases activity has already been well established in both in vitro and ex vivo studies. However, I/R injury is a complex issue based on several overlapping mechanisms. Increased activity of myosin light chain kinase and nitric oxide synthase due to oxidative stress leads to post-translational modifications of MLC1, thus leading to the increased degradation of these proteins. Methods: Wistar rats were subjected to left anterior descending coronary artery occlusion. To measure the pharmacological effect of doxycycline, transthoracic echocardiography as well as biochemical tests, concentrations of TnI, LDH, MLC1, MMP-2 and MMP-9 were performed. Gelatinize activity and cytotoxicity level were also assessed; Results: I.p., administration of doxycycline before LAD occlusion surgery increased TnI and LDH content in the heart and decreased cytotoxicity. A reduction of MMP-2 and MMP-9 concentration and MMP-2 activity after administration of Doxy was also observed, as well as improvement in echocardiographic parameters just 7 days after surgery. Conclusions: Inhibition of MMPs by doxycycline, in vivo, may serve as a protective agent in future therapy.

Many studies have proven the involvement of the RhoA/ROCK pathway in autoimmune and cardiovascular diseases and the beneficial effects of its downregulation. Here, we examined whether the effect of simvastatin on experimental autoimmune myocarditis (EAM) may be through targeting the Ras homolog family member A/Rho-associated coiled-coil containing kinases (RhoA/ROCK) pathway and whether previously shown downregulation of metalloproteinase 9 (MMP-9) could be associated with MLC phosphorylation. Two doses of simvastatin were administered to experimental rats with autoimmune myocarditis by gastric gavage for 3 weeks, at the stage of development of the inflammatory process. Immunohistochemical staining for RhoA and ROCK1 was evaluated semi-quantitatively with H-score. The RhoA staining showed no significant differences in expression between the groups, but the ROCK1 expression was significantly upregulated in the hearts of the EAM group and was not downregulated by simvastatin. The Western blotting analysis of the last downstream product of the RhoA/ROCK axis, phosphorylated myosin light chain (phospho-MYL9), revealed that protein content increased in EAM hearts and it was prevented by the highest dose of simvastatin. Our findings suggest that the RhoA/ROCK pathway is upregulated in EAM, and simvastatin in EAM settings inhibits the RhoA/ROCK pathway at the stage of phosphorylation of myosin light chains and provides a new insight into the molecular pathology of autoimmune myocarditis.