3-nitrotyrosine and Meningitis--Pneumococcal

Reactive oxygen and nitrogen species are released by immune-competent cells and contribute to cellular damage. On the other hand, certain pathogens, including Streptococcus pneumoniae, are known to produce hydrogen peroxide (H2O2), while production of nitrogen radicals by bacteria presumably occurs but has been poorly studied. We determined the relative contributions of bacterial versus host-derived oxygen and nitrogen radicals to cellular damage in pneumococcal infection. A special focus was placed on peroxynitrite as a hypothetical common product formed by the reaction of H2O2 and NO. In microglial cultures, reduction of the formation of 3-nitrotyrosine and cellular damage required H2O2-deficient (DeltaspxB or DeltacarB) pneumococci and inhibition of host NO synthesis with aminoguanidine. In infected C57BL/6 mice, neuronal loss and immunopositivity for nitrotyrosine in the dentate gyrus were markedly reduced with DeltaspxB or DeltacarB bacterial mutants and in inducible nitric oxide synthase knockout mice. We conclude that although host and bacteria both produce oxygen and nitrogen radicals, the interplay of prokaryotic H2O2 and eukaryotic NO is a major contributor to cellular damage in pneumococcal meningitis.

Topics: Animals; Carbamoyl-Phosphate Synthase (Ammonia); Cells, Cultured; Disease Models, Animal; Gene Deletion; Guanidines; Humans; Hydrogen Peroxide; Meningitis, Pneumococcal; Mice; Mice, Inbred C57BL; Neuroglia; Neurons; Nitric Oxide; Nitric Oxide Synthase; Peroxynitrous Acid; Pyruvate Oxidase; Streptococcus pneumoniae; Tyrosine

The aim of the study was to determine the differential expression of nitric oxide (NO) synthase (NOS) isoforms and the pathophysiologic relevance of inducible NOS (iNOS) in experimental pneumococcal meningitis. By use of reverse transcription-polymerase chain reaction analysis, immunohistochemistry, and Western blotting, increased brain mRNA and increased protein levels of endothelial NOS (eNOS) and iNOS were detected 24 h after intracisternal pneumococcal inoculation. In iNOS-deficient mice, disruption of the blood-brain barrier (BBB) was significantly reduced, compared with that in wild-type mice. This beneficial effect of iNOS deficiency was associated with a lack of nitrotyrosine immunoreactivity. Furthermore, brain protein levels of interleukin (IL)-1beta, IL-6, and tumor necrosis factor-alpha and brain mRNA levels of macrophage inflammatory protein (MIP)-1alpha and MIP-2 were significantly reduced in infected animals lacking iNOS. These findings suggest that (1) not only iNOS but also eNOS is up-regulated in the acute phase of experimental bacterial meningitis, and (2) iNOS-derived NO contributes to peroxynitrite formation and BBB breaching in this disease.

Topics: Animals; Blood-Brain Barrier; Blotting, Western; Chemokine CCL3; Chemokine CCL4; Chemokine CXCL2; Cytokines; Female; Gene Expression Regulation, Enzymologic; Immunohistochemistry; Macrophage Inflammatory Proteins; Male; Meningitis, Pneumococcal; Mice; Mice, Inbred C57BL; Monokines; Nitric Oxide Synthase; Protein Isoforms; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tyrosine; Up-Regulation

This study investigated the role of peroxynitrite in an adult rat model of pneumococcal meningitis. Immunohistochemically, nitrotyrosine residues, as a marker for peroxynitrite formation, were detected perivascularly and in proximity to inflammatory cells in the subarachnoid space. Nitrotyrosine immunoreactivity was colocalized with blood-brain barrier breaching, which was visualized by fluorescence microscopy after intravenous application of Evans blue. Treatment of infected rats with uric acid (300 mg/kg intraperitoneally), a scavenger of peroxynitrite, significantly attenuated intracranial pressure, cerebrospinal fluid white blood cell count, and blood-brain barrier leakage, as indicated by Evans blue concentration in the cerebrospinal fluid (21.6+/-9.3 mm Hg, 5776+/-1790 cells/microL, 9.7+/-6.4 microgram/mL in infected, untreated rats vs. 7.2+/-1.6 mm Hg, 2004+/-904 cells/microL, 1.1+/-1.0 microgram/mL infected, uric acid-treated rats, mean+/-SD, P<.05). These data suggest that peroxynitrite plays a central role in mediating pathophysiological alterations during bacterial meningitis.

Topics: Animals; Blood-Brain Barrier; Brain; Cerebrospinal Fluid; Free Radical Scavengers; Inflammation; Intracranial Pressure; Leukocytes; Male; Meningitis, Pneumococcal; Nitrates; Oxidants; Rats; Rats, Wistar; Subarachnoid Space; Tyrosine; Uric Acid