Modulation of inflammatory mediators during experimental bacterial meningitis

Abstract: Bacterial meningitis is a worldwide health problem with high morbidity and mortality mainly caused by Haemophilus influenzae, Neisseria meningitidis and Streptococcus pneumoniae. It is an inflammatory disease of the central nervous system (eNS) initiated by cell wall or membrane components from the bacteria. This stimulates an accumulation of polymorphonuclear leukocytes (PMN) in the subarachnoid space, where they are further stimulated by bacterial products or by cytokines to release inflammatory mediators including nitric oxide (NO) and chemokines. The role of these mediators in the inflammatory response in vivo is not fully understood. The aim of present thesis was to investigate the role of these inflammatory mediators using Haemophilus influenzae type b (Hib) in an experimental model for meningitis.Intraperitoneal inoculation of Hib into infant rats resulted in induction of iNOS mRNA that was detected in brain sections 12 hr post-inoculation (p.i). The number of iNOS mRNA-containing cells was gradually reduced and normalized by day 7 p.i. The numbers of intracerebral iNOS expressing cells was also detected after 12h p.i., but they were further elevated to a peak at 72h p.i. The iNOS positive brain sections also bound antibody specific for nitrotyrosine.Brains of infant rats challenged intraperitoneally with Hib resulted in a time-dependent expression of MIP-2, MIP-1α MCP-1 and RANTES, which were detected with maximum levels at 24-48 h p.i. There were significant increases in the number of neutrophils and macrophages in the meninges, the ventricular system and the periventricular area. The kinetics of MIP-2, MIP-α, MCP-1 and RANTES mRNA expression paralleled those of the recruitment of inflammatory cells and disease severity. Administration of anti-MIP-2 or anti-MIP-1α antibodies (Abs) reduced the number of infiltrated neutrophils, anti-MCP-1 Abs reducing macrophage infiltration.Hib infection resulted in 100% mortality by 72 h p.i. and pretreatment with CNI-1493, a tetravalent guanylhydrazone, resulted in 75% survival. This treatment also reduced the number of infiltrating granulocytes in the brain and reduced the number of cells producing TNF-α and IL-Iß in the spleen.Macrophages (RAW 246.7) preincubated with CNI-1493 prior to activation with LPS/IFN- γ had decreased NO production and reduced iNOS expression. The production of reactive oxygen species (ROS) was increased in FMLP-stimulated granulocytes following CNI-1493-treatment, whereas their F-actin content, motility and chemotaxis were decreased under the same condition.Conclusion: During experimental Hib meningitis iNOS was upregulated and associated with NO production. The chemokines MIP-2, MIP-1α MCP and RANTES were expressed and shown to be involved in neutrophil recruitment in vivo. Blockade of MIP-2 or MIP-1α bioactivity in vivo resulted in decreased neutrophil influx. CNI-1493 treatment increased animal survival by attenuating CNS inflammation and in vitro CNI-1493 had a direct effect on both macrophages and PMN.