Objective The aim of this study was to evaluate whether myosin light chain kinase (MLCK) knockdown attenuated H9C2 cell hypoxia/reoxygenation (H/R) injury and downstream signaling pathway. Methods The MLCK expression… Click to show full abstract
Objective The aim of this study was to evaluate whether myosin light chain kinase (MLCK) knockdown attenuated H9C2 cell hypoxia/reoxygenation (H/R) injury and downstream signaling pathway. Methods The MLCK expression in H/R injury model H9C2 cell was determined by western blot and qRT-PCR. H/R cells were transfected with si-MLCK in the presence of P38 inhibitor (SB203580) or ERK inhibitor (U0126). Then, cell apoptosis was verified by flow cytometry. Apoptosis-related proteins were detected by western blot. The contents of reactive oxygen species (ROS), lactate dehydrogenase (LDH), superoxide dismutase (SOD), interleukin-6 (IL-6), interleukin (IL)-1β (IL-1β), and tumor necrosis factor-α (TNF-α) were measured using flow cytometry and colorimetric assays, respectively. Results MLCK expression was higher in H/R cells. Knockdown of MLCK diminished the amounts of ROS, LDH, IL-6, IL-1β, and TNF-α and elevated the release of SOD in H/R model H9C2 cells. Additionally, H/R injury induced the cumulative expression and phosphorylation of ERK and the phosphorylation of P38, whereas MLCK siRNA-treated cells showed decreased ERK1/2 and P38 activation. Inversely, P38 inhibitor (SB203580) and ERK inhibitor (U0126) could reverse the cardioprotective effects induced by si-MLCK. Conclusion MLCK knockdown attenuated H/R injury in H9C2 cells via regulating the ERK/P38 signaling pathway. MLCK/ERK/p38 axis may provide novel insight into therapeutic targets to restrain I/R injury caused by revascularization therapy after acute myocardial infarction.
               
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