This proposal is based upon three hypotheses: 1) The effective antioxidant activity of propranolol in vivo is primarily attributable to its biotransformation into more potent metabolites rather than to propranolol itself. 2) Lysosomes (particularly from endothelial cells) are a major source of iron, which enhances vascular injury in response to free radicals, and that concentrating antioxidants within lysosomes may provide enhanced tissue protection. 3) Modulation of lysosomal iron stores prior to oxidant exposure will alter the extent of injury induced by the oxidative stress. The applicant will utilize a variety of techniques to monitor: free radical production and oxidative injury (ESR spin trapping, lipid peroxidation assays, and endogenous antioxidant quantification); iron mobilization (histochemically and biochemically); and tissue and cellular toxicity (cellular viability and myocardial function) to investigate the following specific aims: 1. Determine the antioxidant properties and potency of 4-hydroxypropranolol (HOP) and a-naphthoxylacetic acid (NLA) (the two major biotransformed metabolites of propranolol). 2. Determine the effectiveness of the major metabolites of d-propranolol on ischemia/reperfusion-induced injury (hemodynamics; oxyradical, nitric oxide, and lipid radical production; etc.). 3. Determine if d,l- and d-propranolol and their major metabolites accumulate within the endothelial cell acidic compartments, and the mechanism and timing of this accumulation. 4. Examine the role of lysosomes and lysosomal iron in endothelial cell oxidative injury induced in response to applied (H2O2) and endogenously-induced (anoxia/reoxygenation) oxidants, and the effect of treatment with propranolol and its major metabolites on nitric oxide production in vivo. The applicant suggests that even mild iron excess in vivo may predispose cardiovascular patients to further risk, and that the major metabolites of propranolol may be cardioprotective clinically, in part, due to potent antioxidant properties.
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