p38 triggers late preconditioning elicited by anisomycin in heart - Involvement of NF-kappa B and iNOS

We investigated the role of stress-activated p38 MAP kinase (p38/SAPK-2) si gnaling in delayed preconditioning of the heart. Adult male out-bred ICR mi ce were treated with p38 activator, anisomycin (0.1 mg/kg IP), or vehicle ( 5% DMSO). Twenty-four hours later, hearts were perfused in Langendorff mode and subjected to 30 minutes of ischemia and 30 minutes of reperfusion. Imp rovement in postischemic recovery of end-diastolic pressure and reduction i n infarct size was observed, which was abolished by SB203580, a specific p3 8 inhibitor, and pyrrolidinediethyldithiocarbamate (PDTC), the NF-kappaB in hibitor, but not by PD 98059, a specific inhibitor for MEK1 or 2. Transient increase in p38 phosphorylation was observed 15 minutes after anisomycin t reatment which subsided by 30 minutes. Electrophoretic mobility shift assay demonstrated rapid activation of NF-kappaB DNA binding with anisomycin, pe aking at 30 minutes. Western blot confirmed the accumulation of p50 and p65 in nuclear extracts after anisomycin treatment. Anisomycin-induced NF-kapp aB DNA binding activity was inhibited by SB203580 and PDTC. Expression of i nducible nitric oxide synthase (iNOS) mRNA, protein, and nitric oxide (NO) synthesis were enhanced in anisomycin-treated mice. SB203580 and PDTC block ed the increased expression of iNOS and increase in synthesis of NO. Select ive iNOS inhibitor S-methylisothiourea abolished the protective effect of a nisomycin. Furthermore, postischemic cardioprotective effect of anisomycin was absent in mice with targeted ablation of iNOS gene but not in the wild- type B6.129 mice. For the first time, these results suggest that direct pha rmacological activation of p38 triggers delayed preconditioning by signalin g mechanism involving NF-kappaB activation and synthesis of NO from iNOS.