Analysis of RHO-RHO kinase pathway in a mouse model of hindlimb ischemia-reperfusion

Aim: Ischemia-reperfusion (I/R) damage is a broad and not fully understood physiopathological process in both clinical and molecular aspects; which is related to multiple clinical conditions such as infection and cancer. Rho-Rho Kinase signal transduction pathway has shown to be related to cerebral, renal, myocardial and hepatic ischemia-reperfusion. This study is planned to demonstrate alterations in Rho-Rho Kinase signal pathway in mouse skeletal muscle ischemia-reperfusion in both enzyme activity and protein expression levels. By means of the results, new strategies are to be planned to overcome ischemia-reperfusion injury in free muscle transfers. Additionally, Materials and Methods: Swiss albino mice weighing between 20-30 g were divided to 2 groups as control and study, 12 each. Study group was subjected to ischemia by application of tourniquet from great trochanteric level for 4 hours. At the end of 4 hours tourniquet was ended and reperfusion period started. By the end of totally 8 hours muscle specimens from biceps femoris muscle were taken and RhoA protein level, Rho Kinase enzyme level and Rho Kinase activities were assessed by ELISA. In control group only 8 hours of anesthesia was applied without tourniquet application, results of ELISA study were compared. Muscle biopsies were also taken in order to make a histopathological evaluation. Results: Rho Kinase activity is significantly higher in the study group. While no difference was observed in Rho Kinase levels, RhoA levels found to be lower in study group. Conclusions: Results of this study which used a mouse hindlimb ischemiareperfusion model demonstrates that Rho-Rho Kinase pathway is stimulated during skeletal muscle ischemia-reperfusion at kinase activity level. Rho kinase inhibitors may be useful agents by means of reducing both necrosis and apoptosis during free muscle transfers. Keywords: Ischemia-reperfusion, skeletal muscle, Rho, Rho-Kinase