Inhibition of double stranded RNA dependent protein kinase (PKR) abrogates isoproterenol induced myocardial ischemia in vitro in cultured cardiomyocytes and in vivo in wistar rats.

Protein kinase R (PKR) plays a main role in inflammation, insulin resistance, and glucose balance. It is activated by various stress signals and is key mediators of diabetes and associated complications. In the present study, we investigated the effect of PKR inhibition on myocardial dysfunction, inflammatory, cell death and interrelated signalling pathways in isoproterenol induced myocardial ischemia in vivo in wistar rats and in vitro in cultured cardiomyocytes. H9C2 rat cardiomyocytes were treated with 10 μM Isoproterenol (ISO). For in vivo studies, rats were divided into 4 groups: control, ischemic group (ISO), preventive group, curative group and each group consist of 8 rats. Myocardial Ischemia (MI) was induced with two subsequent doses of ISO (100 mg/kg, s.c.). The rats were treated with PKR inhibitor, C16 (166.5 μg/kg, i.p.) for 14 days. Heart rate, systolic, diastolic and mean arterial pressures were measured by non-invasive BP apparatus. Cardiac biomarkers were measured by commercial kits. Ischemic Zone, Morphological abnormalities and fibrosis of heart was detected by TTC, haematoxylin & eosin staining, Masson's and Sirius red staining respectively. Protein expression was done by western blotting and immune histochemistry. mRNA expression was done by RT-PCR. MI was characterized by declined myocardial performance along with elevation of cardiac biomarkers and associated with increased expression of PKR, oxidative-nitrosative stress, activated various inflammatory pathways (nuclear factor kappa light chain enhancer of activated B cells -NF-κB); Mitogen-activated protein kinases-MAPK; c-Jun N-terminal kinase-JNK), increased expression of inflammatory markers (Tumour necrosis factor alpha-TNF-α), markers of fibrosis (Alpha smooth muscle actin -α-SMA; Transforming growth factor beta-TGF-β), enhanced cell death (Ischemic zone) and increased expression of extracellular regulated-kinases (ERK-1/2) and advanced glycation end products (AGE's). Interestingly, inhibition of PKR attenuated myocardial dysfunction, cardiac fibrosis, oxidative/nitrosative stress, inflammation, cell death, and inter-related signalling pathways. Our findings report that inhibition of PKR improves the ischemic mediated inflammation, apoptosis, cardiac hypertrophy and fibrosis in MI induced rats. Hence, inhibition of PKR might be one of intervention therapy for the treatment of myocardial ischemia.