Transplasma membrane electron transport (TPMET) systems in pulmonary endothelial cells effect reduction of extracellular electron acceptors via transport of intracellular donors to the extracellular acceptors. The physiological role and mechanisms of pulmonary TPMET are not well understood. A general hypothesis motivating the proposed research is that pulmonary endothelial TPMET influences the redox status of systemic arterial blood constituents. This function can have either antioxidant or prooxidant consequences depending on the nature of the electron acceptors arriving via the mixed venous blood. In its antioxidant role, pulmonary endothelial TPMET regeneration of plasma lipoprotein antioxidants contributes to antioxidant defense of the systemic circulation. This is, in part, a functional outcome of the large pulmomary endothelial surface area and the location of the lung in the circulatory system. The very properties of lung TPMET that promote its protective effects also have the potential to initiate oxidant injury when TPMET systems are presented with redox active toxins or free transition metal electron acceptors. The goal of the proposed studies is to examine these concepts and to further elucidate cellular mechanisms involved in pulmonary endothelial TPMET.
The specific aims of the proposed research are as follows: I. Determine the influence of pulmonary endothelial TPMET on the extracellular redox state of physiological or toxicological redox active compounds. The specific hypotheses are that pulmonary endothelial TPMET systems (1) reduce the oxidized form of coenzyme Q0 and Trolox C quinone to their antioxidant hydroquinone forms, (2) regenerate the reduced forms of the plasma lipoprotein antioxidant coenzyme Q10 as a mechanism underlying endothelial protection of plasma lipoprotein from oxidation, and 3) reduce the pulmonary toxin paraquat to its prooxidant monocation form. The general approach will be to demonstrate that reduction products of these electron acceptors appear in the extracellular medium when the intact cells are exposed to the oxidized forms, and to determine whether there are proteins on the cell surface that are capable of mediating the reduction carried out by the intact cells. II. Elucidate cellular mechanisms involved in TPMET. The specific hypotheses are that the activity of one of the pulmonary endothelial TPMET systems (1) depends on intracellular redox status as reflected in the intracellular NAD(P)H/NAD(P)+ ratios, and (2) involves a TPMET flavoprotein, and possibly other, redox centers. The general approach will be to correlate TPMET activity and pyridine nucleotide redox poise in intact cells and to determine some of the effects of redox prosthetic group inhibitors on the reductase activity of isolated plasma membrane redox components.
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