Single wells which could not be enumerated because of confluence phenomena were assessed by using the highest numbers of cytokine-producing cells which could be regularly counted in other wells in the same assay as an approximated estimate

Single wells which could not be enumerated because of confluence phenomena were assessed by using the highest numbers of cytokine-producing cells which could be regularly counted in other wells in the same assay as an approximated estimate. Statistical analysis For statistical analysis, the two-sided em t /em -test for different variances (GraphPad PRISM 4 software, La Jolla, USA) was used. the influence of the Toll-like-receptor (TLR) ligand CpG oligonucleotide (CpG) on already established CNS autoimmunity in murine proteolipid protein (PLP)-induced EAE in SJL mice. CpG were found to co-stimulate PLPp-specific IFN- production in the peripheral immune system and in GNE-617 the CNS. However, CpG induced Interleukin (IL)-17 production in the inflamed CNS both alone and in combination with additional PLPp stimulation. These findings might indicate a mechanism by which systemic infections GNE-617 and the microbial stimuli associated with them may influence already existing CNS autoimmune pathology. Background Research on the role of GNE-617 T cells in CNS autoimmune disease both in human diseases as well as in their experimental animal models currently centers on the endogenous requirements which are necessary for T cell activation as well as on the exogenous factors which trigger it. Among the environmental elements, which can influence this process (both in a positive and negative way), infections are considered important [1;2]. These include both viral and bacterial infections as well as the pathogenic factors which are associated with them. The potential role of microbial pathogens in triggering autoimmune disease has been extensively studied both in humans as well as in experimental animal models. As an example for one of many infectious agents Rabbit Polyclonal to PTPRZ1 which have been discussed as the cause of MS, a positive association between Epstein-Barr virus (EBV) infection and increased risk of developing MS thereafter has been broadly discussed [3]. Such clinical observations have been supported by basic observations from cell culture models which try to explain how infectious agents can affect the CNS and facilitate key steps in MS pathogenesis, e.g. through creating a local GNE-617 proinflammatory milieu in the early stages of disease [4]. In the EAE mouse model, there is currently increasing evidence for a critical role of commensal gut microbiota in the initiation of CNS autoimmunity, as demonstrated by experiments in which reduction of the commensal microflora by antibiotic treatment inhibits the development of EAE [5]. However, despite the fact that disease progression or relapse is clinically to the same extent associated with bacterial or viral infection as the beginning of disease, in particular in the case of MS [6-9], the influence of these pathogenic conditions on already established autoimmune disease has received less attention. In EAE in the SJL mouse model lipopolysaccharide, a TLR 4 ligand, has been shown to be able to induce relapses via antigen presenting cell (APC)-dependent activation of autoantigen-specific T cells [10]. When studying the impact of infection on ongoing CNS autoimmunity, a differentiated setup has to be chosen. First, systemic effects of a microbial stimulus on the autoimmune T cell population might differ from its effects in the CNS [11;12]. Second, different T cell populations might be affected in a distinct way, which might also be different in the periphery and in the inflamed CNS. T cell populations that are currently implied in CNS autoimmune pathology are Th1 and Th17 cells, which are characterized by the production of IFN- and IL-17, respectively [13]. Both cytokines are important mediators of disease and tissue damage in CNS autoimmunity, albeit with different roles in the autoimmune process and different resulting pathology [14-16]. For this reason, it was the purpose of this study to investigate the influence of CpG as a paradigm of a microbial stimulus which is able to activate both APC [17] as well as T cells directly [18] on GNE-617 the PLPp-specific T cell cytokine production in EAE in SJL mice, in particular in regard of IFN- and IL-17. Methods Animals, antigens and treatments Female SJL/J.