INACTIVATION OF HEART DIHYDROLIPOAMIDE DE HYDROGENASE BY LEUKOCYTE MYELOPEROXIDASE-DEPENDENT SYSTEMS - PREVENTION BY CAPTOPRIL AND OTHER THIOL COMPOUNDS

Myocardial dihydrolipoamide dehydrogenase (LADH) is inactivated after incubation at 30 degrees C, with myeloperoxidase (MPO)-dependent syste ms. The enzyme inactivation was a function of the pro-oxidant system c omposition and the time of incubation. The standard inactivating syste m contained 50 mM KH2PO4-K2HPO4, pH 7.4, 0.5-1.0 mu M LADH, and pro-ox idant system. After 30 or 60 min of incubation with the MPO/H2O2/NaCl system, LADH inactivation was 64 and 87%, respectively (Figure 1). In the absence of NaCl, inactivation values were 9 and 27%, respectively, whereas in the absence of MPO the inactivation Values were 4.0 and 11 %, respectively (Figure 1). Under similar experimental conditions, sod ium hypochlorite significantly inactivated LADH, thus supporting the r ole of hipochlorous acid as agent of the MPO/H2O2/ClNa system. With th e MPO/H2O2/Kl, MPO/H2O2/KSCN or the MPO/H2O2/NaNO2 systems LADH inacti vation depended on the anion nature, I being the most effective (Figur e 2). NaNO2 effectively replaced halides as pro-oxidant (Figure 3). Th e MPO/NADH/halide systems, where NADH replaced H2O2, also inactivated LADH. Native (not denatured) catalase completely prevented the MPO/NAD H/Kl system effect (Table 1), in close agreement with H2O2 production by the LADH-catalysed NADH oxidation and the role of H2O2 in LADH inac tivation. LADH was also inactivated after incubation with MPO-generate d free radicals such as the Chloropromazine and Paracetamol radicals ( Table 2). Thiol compounds (Captopril, penicillamine, cysteine, N-acety lcysteine and mercaptopropionylglycine) (Table 3 and Figure 4), as wel l as taurine, ascorbate (Table 4), GSSG and trypanothione (Figure 5), protected LADH against the MPO-dependent oxidizing systems, and also a gainst NaClO (Table 4). The summarized observations are discussed in r elation to MPO function in free radical production and pathologies suc h as ischemia-reperfusion injury and inflammation.