Transforming growth factor beta concentrations and interferon gamma responses in cerebrospinal fluid of horses with equine protozoal myeloencephalitis

The following experiment was performed to test the hypothesis that transforming growth factor beta (TGF-beta) concentration varies in the cerebrospinal fluid and serum of horses with EPM and to determine if cerebrospinal fluid (CSF) alters the interferon-gamma (IFN-gamma) rersponse of equine peripheral blood mononuclear cells (PBMCs). The concentration of transforming growth factor-beta (TGF-beta (2)) was investigated in the serum and cerebrospinal fluid (CSF) of 18 horses (9 normal, 9 affected with equine protozoal myeloencephalitis [EPM]). The TGF-beta (2) assay was validated in a group of 6 normal horses. Intra-assay variability was 4.7 %, and interassay variability was 10.7 %. The slope of the curve of the unknown samples of various volumes demonstrated parallelism with a curve developed using equal volumes of assay kit standard. Assay of normal and EPM-affected horses found that TGF-beta (2) was present in both the serum and CSF of all animals. However, the concentration of TGF-beta (2) in the CSF was less (P = 0.03) in EPM-affected horses (144 pg/ml) than in normal horses (256 pg/ml). In addition, the effect of CSF from normal and EPM-affected horses on the production of interferon-gamma (IFN-gamma) by PHA-P stimulated PBMCs from normal horses was investigated using a bioassay. It was found that CSF from normal and EPM-affected horses enhanced IFN-gamma activity from PHA-P stimulated peripheral blood mononuclear cells (P less than or equal to 0.05); however, the response to CSF from EPM-affected horses was no different than the response to CSF from normal horses. Treatment of cells with anti-TGF-beta (2) monoclonal antibodies slightly increased the response when co-incubated with CSF from normal horses, and slightly decreased it when co-incubated with CSF from EPM-affected horses. These differences, however, did not achieve statistical significance (P >0.05). Results of this study indicated that production of TGF-beta (2) is altered in horses with EPM, and that CSF appears to contain substances which alter the inflammatory reaction to plant lectins. These findings confirm the immunomodulatory properties of CSF and suggest new techniques for future research regarding the pathophysiology of EPM.