Objective: The protective properties of heme oxygenase 1 (HO-1) give reason to study this mechanism as a potential therapeutic target for inflammatory and cardiovascular diseases. Recent evidence suggests a possible interaction between the HO-1/CO- and the protein kinase Akt/NO-pathway. This study was designed to examine the effects of continuous HO-1 overexpression in endothelial cells.
Methods: Oncoretroviral vectors were constructed to achieve constitutive overexpression of HO-1, Akt, and green fluorescence protein in human umbilical vein endothelial cells. [(3)H]thymidine-incorporation and lipid-peroxidation were measured following exposure to heme and H(2)O(2). Expression of HO-1, Akt and its downstream-target endothelial NO-synthase were quantified by Western blot analysis. NO-synthase-activity was measured using the citrulline-conversion-assay.
Results: HO-1-overexpression reduced proliferative rates and DNA-synthesis of HUVEC, but provided potent protection from oxidative stress induced by heme and H(2)O(2). Phosphorylated-Akt and eNOS was downregulated in HO-1-HUVEC. eNOS-activity was reduced in HO-1-HUVEC. Co-infection with the Akt-retrovirus restored proliferative rates and eNOS-expression and -activity.
Conclusion: Continuously elevated HO-1-activity protects EC from oxidative stress but inhibits Akt-mediated proliferation and eNOS-expression. This inhibitory feedback mechanism could be a limitation of HO-1 as a target for the treatment of vascular disease.