HIF-regulated HO-1 gene transfer improves the post-ischemic limb recovery and diminishes TLR-triggered immune responses - Effects modified by concomitant VEGF overexpression

Heme oxygenase-1 (HO-1) mitigates cellular injury by antioxidant, anti-apoptotic, anti-inflammatory and proangiogenic effects. Vascular endothelial growth factor (VEGF) is a critical regulator of blood vessel growth. Their coordinated action was analyzed in a model of femoral artery ligation (FAL) in mice lacking HO-1 gene (HO-1 KO). Gastrocnemius skeletal muscles of HO-1 KO mice were preemptively injected with plasmids containing hypoxia-response element (HRE) driving the expression of only HO-1 (pHRE-HO1) or both HO-1 and VEGF (pHRE-HO1 VEGF). At day 14th the pHRE-HO1 vector increased an impaired post-ischemic blood flow recovery in HO-1 KO mice to the level observed in wild-type (WT) mice subjected to FAL and pHRE-HO1 VEGF restored it already at day 7. The pHRE-HO1 gene therapy diminished, when compared to control pHRE-empty-treated HO-1 KO mice, the expression of toll-like receptors (TLR4 and TLR9) and inflammatory cytokines (IL-1 beta, IL-6 and TNF alpha) at day 3, whereas opposite effects were observed following concomitant HO-1 and VEGF gene transfer. Moreover, HO-1 diminished ischemia-induced expression of MyoD involved in satellite cell differentiation in HO-1 KO mice. Our results confirm the therapeutic potential of HO-1 and VEGF against critical limb ischemia although, their concomitant delivery may have contradictory actions on the resolution of inflammation. (C) 2015 Elsevier Inc All rights reserved.