The activation of protein kinase C induces higher production of reactive oxygen species by mononuclear cells in patients with multiple sclerosis thanin controls

Objective: Recent findings have increasingly shown the importance of reacti ve oxygen species (ROS) in causing oxidative damage to macromolecules and i n contributing to tissue degeneration in target organs of autoimmune diseas es. This study was aimed at comparing the base line and induced production of ROS by peripheral blood mononuclear cells (PB MNCs) of patients with mul tiple sclerosis (MS) in remission and relapse, of patients with other neuro logical diseases (OND) and of healthy controls. In addition, we analyzed th e underlying mechanism of ROS production. Methods: PB MNCs were separated from 28 MS patients in remission and 13 in relapse, and from 29 healthy controls and 10 OND. ROS was measured by spect rofluorometry. Expression of proinflammatory cytokines was assessed by semi quantitative reverse transcription polymerase chain reaction (RT-PCR). Mito chondrial (mt) DNA haplotypes were determined by using restriction site pol ymorphism analysis. Results: The base line and tumor necrosis factor (TNF)-alpha or interferon (IFN)-gamma induced ROS values were similar in the four groups, and the ind ividual measures did not show a correlation with MS associated mtDNA haplot ypes. Phorbol ester activation of protein kinase C (PKC) induced higher ROS production in all groups, however, with significantly greater values in th e MS remission group. Calphostine C, a PKC inhibitor decreased or eliminate d ROS production in a dose-dependent manner, suggesting further that it was predominantly or exclusively generated by PKC activated NADPH oxidase. A t rend of increased TNF-alpha and IFN-gamma expression was noted in the MS re lapse group, in contrast to the high ROS release in the MS remission group. Conclusion: The detected phase difference between the highest ROS productio n vs TNF-alpha expression is compatible with the hypothesis that different subpopulations of monocytes/macrophages are involved. We suggest that the R OS producing subpopulation preferentially migrates into the central nervous system (CNS) during a relapse. The present study together with our previou s observation on oxidative damage to DNA in active plaques delineates a mol ecular pathway likely involved in the histologic evolution of inflammatory demyelination.