143: Therapeutic induction of heme-oxygenase-1 in the lung using a hemoglobin-based oxygen carrier

Abstract

Introduction: Success in lung transplantation is limited by obligatory ischemia and reperfusion (I/R) injury. Experimentally, endogenous and induced (hemin, hypoxia) overexpression of heme-oxygenase-1 (HO-1) provides protection against I/R injury, which is abolished with selective inhibition of this enzyme. Unfortunately, no clinically viable therapy exists to safely induce HO-1. We have previously reported that administration of a hemoglobin-based oxygen carrier (HBOC) upregulated HO-1 in isolated pulmonary microvascular endothelial cells. We therefore hypothesized that administration of a HBOC in vivo would induce expression of HO-1 in the lung. Methods: HBOC (PolyHeme®, human polymerized hemoglobin, pyridoxylated; Northfield Laboratories, Inc.) at 0% (control), 5% or 10% of calculated blood volume was administered intravenously (femoral vein) to anesthetized rats (N=5 per group). Recovering animals were subsequently maintained in a normothermic environment and sacrificed at 12 hours. Lungs were isolated, washed and then either flash frozen for tissue homogenization or frozen in embedding medium for histology. Immunoblot of tissue samples were probed for HO-1 and heat shock protein 72 (HSP72, stress-inducible). Densitometry was performed using ImageJ. Microscopic examination of tissue blocks were probed for HO-1 and CD 68 (anti-rat macrophage marker). Statistical analysis used ANOVA (*p<0.05). Results: Administration of HBOC produced nearly a 4-fold induction of lung HO-1 expression (* p<0.05) (see figure). However, there was no change in the stress-inducible HSP72. Immunohistochemistry confirmed upregulation of HO-1 and demonstrated the maximal upregulation of HO-1 to occur in macrophages. Conclusions: In this study, HBOC selectively induced expression of the protective enzyme HO-1 in the lung, without significant alteration of other stress-inducible heat-shock proteins. Significantly, we demonstrated that maximal production of HO-1 in response to treatment with HBOC occurs in macrophages. These data suggest a potential novel therapeutic strategy in lung transplantation. View Large Image Figure Viewer