Naturally occurring CD4+CD25+ regulatory T cells prevent but do not improve experimental myasthenia gravis.

In the current study, we investigated whether naturally occurring CD4(+)CD25(+) T cells, separated by immunomagnetic anti-CD4 and anti-CD25 Abs from naive animals, are able to protect from experimental autoimmune myasthenia gravis (EAMG) and modify the progression of ongoing disease when administered to Torpedo californica acetylcholine receptor (AChR)-immunized Lewis rats. Even though CD4(+)CD25(+) and CD4(+)CD25(high) T cell frequencies were similar in the spleens and lymph nodes of EAMG and healthy rats, we observed that CD4(+)CD25(+) T cells isolated from the spleens of naive animals inhibited in vitro the Ag-induced proliferation of T cell lines specific to the self-peptide 97-116 of the anti-AChR subunit (R97-116), an immunodominant and myasthenogenic T cell epitope, whereas CD4(+)CD25(+) T cells purified from the spleens of EAMG rats were less effective. CD4(+)CD25(+) T cells from EAMG rats expressed less forkhead box transcription factor P3 but more CTLA-4 mRNA than healthy rats. Naive CD4(+)CD25(+) T cells, obtained from naive rats and administered to T. californica AChR-immunized animals according to a preventive schedule of treatment, reduced the severity of EAMG, whereas their administration 4 wk postinduction of the disease, corresponding to the onset of clinical symptoms (therapeutic treatment), was not effective. We think that the exogenous administration of CD4(+)CD25(+) naive T cells prevents the early events underlying the induction of EAMG, events linked to the T cell compartment (Ag recognition, epitope spreading, and T cell expansion), but fails to ameliorate ongoing EAMG, when the IgG-mediated complement attack to the AChR at the neuromuscular junction has already taken place.

Effect of IgG immunoadsorption on serum cytokines in MG and LEMS patients.

We investigated the effect of IgG immunoadsorption (IA) on cytokine network in patients with treatment-resistant Myasthenia Gravis (MG) and Lambert-Eaton Syndrome (LEMS). We observed upregulation of interleukin (IL)-10, an anti-inflammatory and B cells growth factor, and reduction of pro-inflammatory factors such as IL-18 and IL-17, in both MG and LEMS after IA. Our observation suggests that the massive removal of antibodies might induce modifications of the cytokine balance linked to T and B cells mediated autoimmunity.

The kinesin superfamily motor protein KIF4 is associated with immune cell activation in idiopathic inflammatory myopathies.

The idiopathic inflammatory myopathies (IIMs) dermatomyositis, polymyositis, and inclusion body myositis are characterized by myofiber degeneration and inflammation. The triggering factors of muscle autoaggression in these disorders are unknown, but infiltrating T cells may be activated locally and proliferate in situ. T-cell polarization involving reorientation of cytoskeleton and microtubule-organizing centers mediated by motor proteins may occur within inflammatory cells in the muscle. We therefore analyzed ubiquitous and neuronal kinesin superfamily (KIF) members KIF-5, dynein, and KIF4 in IIM muscle biopsies and in activated peripheral blood lymphocytes from healthy donors. Only KIF-4 was altered. Transcript levels were significantly higher in IIM muscle than in controls, and KIF4 inflammatory cells were found in IIM muscles. In polymyositis and inclusion body myositis, KIF4 cells were mainly located around individual muscle fibers, whereas in dermatomyositis, they were also near blood vessels. KIF4 cells were not specific to any immune lineage, and some were Ki67. In peripheral blood lymphocytes stimulated with mitogens, interleukin 2 or anti-CD3/CD28 antibodies, KIF4 expression was upregulated, and the protein was localized in the cytoplasm in association with lysosome-associated membrane protein 1 and perforin lysosomal vesicles. These results imply that KIF4 is associated with activated T cells, irrespective of their functional phenotype, and that it is likely involved in cytoskeletal modifications associated with in situ T-cell activation in IIM.

Pixantrone (BBR2778) reduces the severity of experimental autoimmune myasthenia gravis in Lewis rats.

Pixantrone (BBR2778) (PIX) and mitoxantrone share the same mechanism of action because both drugs act as DNA intercalants and inhibitors of topoisomerase II. PIX is an interesting candidate immunosuppressant for the treatment of autoimmune diseases because of its reduced cardiotoxicity compared with mitoxantrone. The clinical response to conventional immunosuppressive treatments is poor in some patients affected by myasthenia gravis (MG), and new but well-tolerated drugs are needed for treatment-resistant MG. PIX was tested in vitro on rat T cell lines specific for the immunodominant peptide 97-116 derived from rat acetylcholine receptor (AChR), and showed strong antiproliferative activity in the nanomolar range. We demonstrate in this study that PIX administration reduced the severity of experimental autoimmune MG in Lewis rats. Biological and immunological analysis confirmed the effect of PIX, compared with vehicle-treated as well as mitoxantrone-treated experimental autoimmune MG rats. Anti-rat AChR Abs were significantly reduced in PIX-treated rats, and AChR content in muscles were found increased. Torpedo AChR-induced T cell proliferation tests were found reduced in both in vitro and ex vivo experiments. The effectiveness and the reduced cardiotoxicity make PIX a promising immunosuppressant agent suitable for clinical investigation in MG, although additional experiments are needed to confirm its safety profile in prolonged treatments.

Allorecognition of human neural stem cells by peripheral blood lymphocytes despite low expression of MHC molecules: role of TGF-beta in modulating proliferation.

Neural stem cells (NSCs) transplantation has been proposed as a means of restoring damaged brain tissue, a possibility rendered more likely by reports of low NSCs immunogenicity in various experimental models because of low expression of MHC class I and II as well as co-stimulatory molecules. We investigated the immunogenicity of a human NSC line grown in normal culture conditions and in the presence of pro-inflammatory cytokines IFN-gamma and tumor necrosis factor alpha by one-way mixed lymphocyte reaction (MLR) experiments with peripheral blood lymphocytes from eight HLA-incompatible donors. NSCs stimulated lymphocyte proliferation in almost all donors tested, with stimulation indices in the range of the low-end distribution curve of MLR between donors. The healthy subject that gave negative MLR results was the best compatible donor with respect to NSC haplotype. Since we observed low MLR responses overall, we studied if NSCs might exert any immunomodulatory activity. We detected transcription and release of the immunomodulatory molecule transforming growth factor beta (TGF-beta)-1; moreover, the addition of TGF-beta1 in MLR experiments down-regulated proliferative responses. To further confirm the immunological potential of human NSCs, we studied xenogeneic recognition of NSCs by immunocompetent cells derived from C57BL/6 mice, showing that NSCs can elicit an allo(xeno) response ex vivo. Our data indicate that NSCs have low but not negligible immunogenic potential that is sufficient to activate peripheral lymphocytes. Secretion of TGF-beta1 might balance the immunogenicity of NSCs. Nevertheless, the possibility that allo-NSCs grafting might induce in the long term an immune activation, thus vanishing their therapeutical effect, should not be overlooked and deserves further investigation.