Manganese promotes increased formation of hydrogen peroxide by activated human macrophages and neutrophils in vitro.

Although pro-inflammatory mechanisms have been implicated in the pathogenesis of manganese (Mn²âº)-related neurological and respiratory disorders, relatively little is known about the potential of this metal to interact pro-oxidatively with human phagocytes. The primary objective of the current study was to investigate the effects of Mn²âº as MnCl2 (0.5-100 µM) on the generation of the reactive oxygen species (ROS), superoxide, hydrogen peroxide (H2O2), and hypohalous acids by isolated human blood neutrophils and monocyte-derived macrophages following activation of these cells with the chemotactic tripeptide, FMLP (1 µM), or the phorbol ester, PMA (25 ng/mL). Generation of ROS was measured using the combination of oxygen consumption, lucigenin/luminol-enhanced chemiluminescence, spectrofluorimetric detection of oxidation of 2,7-dichlorodihydrofluorescein, radiometric assessment of myeloperoxidase (MPO)-mediated protein iodination, release of MPO by ELISA, and spectrophotometric measurement of nitrite formation. Treatment of activated neutrophils with either FMLP or PMA resulted in significantly decreased reactivity of superoxide in the setting of increased formation of H2O2 and MPO-mediated iodination, with no detectable effects on either oxygen consumption or MPO release. Similar effects of the metal with respect to superoxide reactivity and H2O2 formation were observed with activated macrophages, while generation of NO was unaffected. Taken together with the findings of experiments using cell-free ROS-generating systems, these observations are compatible with a mechanism whereby Mn²âº, by acting as a superoxide dismutase mimetic, increases the formation of H2O2 by activated phagocytes. If operative in vivo, this mechanism may contribute to the toxicity of Mn²âº.

Potassium-Uptake Systems of Mycobacterium tuberculosis: Genomic and Protein Organization and Potential Roles in Microbial Pathogenesis and Chemotherapy

Mycobacterium tuberculosis (MTB) is a formidable microbial pathogen which uses multiple mechanisms to subvert host immune defences. These include the effective; protective barrier presented by the outer waxy coat; intracellular concealment from host defences; and the ability to enter a prolonged; dormant phase in the infected host. Priority strategies to combat the scourge of TB include the identification of novel and selective targets on/in MTB which are amenable to pharmacological or immune-mediated control. Because they are structurally different from their counterparts in eukaryotic cells and are likely to be essential for survival and growth; the major K+ transporters of MTB represent alternative and novel targets for drug and vaccine design. These K+-uptake systems of MTB are the primary focus of this review; with particular emphasis on their genomic and protein structures; properties and functions; and potential roles in intracellular survival

Effects of clofazimine on potassium uptake by a Trk-deletion mutant of Mycobacterium tuberculosis.

OBJECTIVES: This study was designed to investigate the effects of the membrane-active, anti-mycobacterial agent, clofazimine, on potassium (K+)-uptake by a mutant of Mycobacterium tuberculosis (MTB), in which the Trk system, the major K+ transporter of this microbial pathogen, had been selectively inactivated. METHODS: The ceoB and ceoC genes of MTB, which encode the TrkA proteins, CeoB and CeoC, were deleted by homologous recombination, and the double-knockout mutant and wild-type strains compared with respect to K+ uptake and growth in the presence and absence of clofazimine (0.015-2.5 mg/L) using radioassay procedures. RESULTS: Surprisingly, the magnitudes of K+ uptake and rate of growth of the ceoBC-knockout mutant were significantly (P < 0.05) greater than those of the wild-type strain, due, presumably, to induction of a back-up transporter. Exposure of both the wild-type strain and ceoBC-knockout mutant of MTB to clofazimine was accompanied by dose-related decreases in K+ uptake, as well as growth, which were of comparable magnitude for both strains. CONCLUSIONS: These observations demonstrate that the major K+ transporter of MTB, Trk, as well as an uncharacterized inducible back-up system, is equally sensitive to the inhibitory actions of clofazimine.