15-Deoxy-Δ12,14-prostaglandin J2-induced down-regulation of endothelial nitric oxide synthase in association with HSP70 induction

A natural ligand of peroxisome proliferator-activated receptor γ (PPARγ), 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2), decreases endothelial nitric oxide synthase (eNOS) expression by an unknown mechanism. Here we found that 15d-PGJ2-induced eNOS reduction is inversely associated with heat shock protein 70 (HSP70) induction in endothelial cells. Treatment of cells with 15d-PGJ2 decreased eNOS protein expression in a concentration- and time-dependent manner, but independently of PPARγ with no effect on mRNA levels. Although 15d-PGJ2 elicited endothelial apoptosis, inhibition of both pan-caspases and cathepsins failed to reverse reduction of eNOS protein. Interestingly, we observed that 15d-PGJ2 induced HSP70 in a dose-dependent manner. Immunoprecipitation and heat shock treatment demonstrated that eNOS reduction was strongly related to HSP70 induction. Cellular fractionation revealed that treatment with 15d-PGJ2 increased eNOS distribution 2.5-fold from soluble to insoluble fractions. These findings provide new insights into mechanisms whereby eNOS regulation by 15d-PGJ2 is related to HSP70 induction.