pone-0021868-g002: RTL551 prevents migration of cells outside spleen.Splenocytes were counted from naïve mice and mice at indicated time points after EAE induction. As compared to naïve mice, spleen cell numbers increased after immunization and at onset of EAE followed by gradual decline with EAE progression in control groups of mice, possibly due to migration of cells outside the spleen towards the CNS. This trafficking of cells outside the spleen is prevented after RTL551 treatment. Data presented are the mean±SD of 8 mice per group except for naïve which had 5 mice. Significant differences between the groups (p≤0.05) were determined using Student's t test and are indicated by an asterisk.

Mentions: At onset of clinical signs of EAE prior to treatment, there was a significant 3X increase in the splenocyte numbers in mice immunized with MOG-35-55/CFA/Ptx (cell number in naïve vs. EAE mice, 45.5±19.9 vs. 144.8±54.4, Fig. 2). In the acute phase of EAE, cells are still trafficking from the periphery into the CNS, and as EAE progressed, there was a gradual decline in splenocyte numbers in Vehicle and RTL550 treated mice (Fig. 2), suggesting the migration of cells outside the spleen possibly towards the CNS. Interestingly, RTL551-treated mice retained the increased levels of cells in the spleen throughout the observation period.

Sinha S, Miller LM, Subramanian S, Burrows GG, Vandenbark AA, Offner H - PLoS ONE (2011)

Bottom Line: We showed that 1-5 daily injections of RTL551 (two-domain I-A(b) covalently linked to MOG-35-55 peptide), but not the control RTL550 ("empty" two-domain I-A(b) without a bound peptide) or Vehicle, reduced clinical signs of EAE, prevented trafficking of cells outside the spleen, significantly reduced the frequency of CD226 and T-bet expressing CD4+ T-cells in blood and inhibited expansion of CD44 expressing CD4+ T-cells in blood and spleen.Concomitantly, RTL551 selectively reduced CNS inflammatory lesions, absolute numbers of CNS infiltrating T-bet expressing CD4+ T-cells and IL-17 and IFN-γ secretion by CNS derived MOG-35-55 reactive cells cultured ex vivo.These novel results demonstrate that a major effect of RTL therapy is to attenuate Th1 specific changes in CD4+ T-cells during EAE and prevent expansion of effector T-cells that mediate clinical signs and CNS inflammation in EAE.

Affiliation: Neuroimmunology Research, Veterans Affairs Medical Center, Portland, Oregon, United States of America.

Abstract: Recombinant T cell receptor ligands (RTLs) that target encephalitogenic T-cells can reverse clinical and histological signs of EAE, and are currently in clinical trials for treatment of multiple sclerosis. To evaluate possible regulatory mechanisms, we tested effects of RTL therapy on expression of pathogenic and effector T-cell maturation markers, CD226, T-bet and CD44, by CD4+ Th1 cells early after treatment of MOG-35-55 peptide-induced EAE in C57BL/6 mice. We showed that 1-5 daily injections of RTL551 (two-domain I-A(b) covalently linked to MOG-35-55 peptide), but not the control RTL550 ("empty" two-domain I-A(b) without a bound peptide) or Vehicle, reduced clinical signs of EAE, prevented trafficking of cells outside the spleen, significantly reduced the frequency of CD226 and T-bet expressing CD4+ T-cells in blood and inhibited expansion of CD44 expressing CD4+ T-cells in blood and spleen. Concomitantly, RTL551 selectively reduced CNS inflammatory lesions, absolute numbers of CNS infiltrating T-bet expressing CD4+ T-cells and IL-17 and IFN-γ secretion by CNS derived MOG-35-55 reactive cells cultured ex vivo. These novel results demonstrate that a major effect of RTL therapy is to attenuate Th1 specific changes in CD4+ T-cells during EAE and prevent expansion of effector T-cells that mediate clinical signs and CNS inflammation in EAE.