ABSTRACT
Autoimmune diseases of the central nervous system (CNS) such as multiple sclerosis (MS) are only partially represented in current experimental models and the development of humanized immune mice is crucial for better understanding of immunopathogenesis and testing of therapeutics. We describe a humanized mouse model with several key features of MS. Severely immunodeficient B2m-NOG mice were transplanted with peripheral blood mononuclear cells (PBMCs) from HLA-DRB1-typed MS and healthy (HI) donors and showed rapid engraftment by human T and B lymphocytes. Mice receiving cells from MS patients with recent/ongoing Epstein-Barr virus reactivation showed high B cell engraftment capacity. Both HLA-DRB1*15 (DR15) MS and DR15 HI mice, not HLA-DRB1*13 MS mice, developed human T cell infiltration of CNS borders and parenchyma. DR15 MS mice uniquely developed inflammatory lesions in brain and spinal cord gray matter, with spontaneous, hCD8 T cell lesions, and mixed hCD8/hCD4 T cell lesions in EAE immunized mice, with variation in localization and severity between different patient donors. Main limitations of this model for further development are poor monocyte engraftment and lack of demyelination, lymph node organization, and IgG responses. These results show that PBMC humanized mice represent promising research tools for investigating MS immunopathology in a patient-specific approach.
Subject(s)
Brain , CD8-Positive T-Lymphocytes , Disease Models, Animal , HLA-DRB1 Chains , Multiple Sclerosis , Spinal Cord , Animals , Humans , Multiple Sclerosis/immunology , Multiple Sclerosis/genetics , Mice , HLA-DRB1 Chains/genetics , CD8-Positive T-Lymphocytes/immunology , Spinal Cord/immunology , Spinal Cord/pathology , Brain/pathology , Brain/immunology , Encephalomyelitis, Autoimmune, Experimental/immunology , Encephalomyelitis, Autoimmune, Experimental/genetics , Encephalomyelitis, Autoimmune, Experimental/pathology , CD4-Positive T-Lymphocytes/immunology , FemaleABSTRACT
Autoimmune diseases affecting the CNS not only overcome immune privilege mechanisms that protect neural tissues but also peripheral immune tolerance mechanisms towards self. Together with antigen-specific T cells, myeloid cells are main effector cells in CNS autoimmune diseases such as multiple sclerosis, but the relative contributions of blood-derived monocytes and the tissue resident macrophages to pathology and repair is incompletely understood. Through the study of oxidized mannan-conjugated myelin oligodendrocyte glycoprotein 35-55 (OM-MOG), we show that peripheral maturation of Ly6ChiCCR2+ monocytes to Ly6ChiMHCII+PD-L1+ cells is sufficient to reverse spinal cord inflammation and demyelination in MOG-induced autoimmune encephalomyelitis. Soluble intradermal OM-MOG drains directly to the skin draining lymph node to be sequestered by subcapsular sinus macrophages, activates Ly6ChiCCR2+ monocytes to produce MHC class II and PD-L1, prevents immune cell trafficking to spinal cord, and reverses established lesions. We previously showed that protection by OM-peptides is antigen specific. Here, using a neutralizing anti-PD-L1 antibody in vivo and dendritic cell-specific Pdl1 knockout mice, we further demonstrate that PD-L1 in non-dendritic cells is essential for the therapeutic effects of OM-MOG. These results show that maturation of circulating Ly6ChiCCR2+ monocytes by OM-myelin peptides represents a novel mechanism of immune tolerance that reverses autoimmune encephalomyelitis.
Subject(s)
Encephalomyelitis, Autoimmune, Experimental , Encephalomyelitis , Animals , Antigens, Ly , Encephalomyelitis/pathology , Immune Tolerance , Mannans , Mice , Mice, Knockout , Monocytes , Myelin-Oligodendrocyte Glycoprotein , Peptides , Receptors, CCR2ABSTRACT
CNS autoantigens conjugated to oxidized mannan (OM) induce antigen-specific T cell tolerance and protect mice against autoimmune encephalomyelitis (EAE). To investigate whether OM-peptides treat EAE initiated by human MHC class II molecules, we administered OM-conjugated murine myelin oligodendrocyte glycoprotein peptide 35-55 (OM-MOG) to humanized HLA-DR2b transgenic mice (DR2b.Ab°), which are susceptible to MOG-EAE. OM-MOG protected DR2b.Ab° mice against MOG-EAE by both prophylactic and therapeutic applications. OM-MOG reversed clinical symptoms, reduced spinal cord inflammation, demyelination, and neuronal damage in DR2b.Ab° mice, while preserving axons within lesions and inducing the expression of genes associated with myelin (Mbp) and neuron (Snap25) recovery in B6 mice. OM-MOG-induced tolerance was peptide-specific, not affecting PLP178-191-induced EAE or polyclonal T cell proliferation responses. OM-MOG-induced immune tolerance involved rapid induction of PD-L1- and IL-10-producing myeloid cells, increased expression of Chi3l3 (Ym1) in secondary lymphoid organs and characteristics of anergy in MOG-specific CD4+ T cells. The results show that OM-MOG treats MOG-EAE in a peptide-specific manner, across mouse/human MHC class II barriers, through induction of a peripheral type 2 myeloid cell response and T cell anergy, and suggest that OM-peptides might be useful for suppressing antigen-specific CD4+ T cell responses in the context of human autoimmune CNS demyelination.
Subject(s)
Axons/drug effects , Encephalomyelitis, Autoimmune, Experimental/drug therapy , Immunosuppressive Agents/pharmacology , Myeloid Cells/drug effects , Spinal Cord/drug effects , T-Lymphocytes/drug effects , Adult , Animals , Axons/immunology , Axons/metabolism , Axons/pathology , Case-Control Studies , Cell Proliferation/drug effects , Cells, Cultured , Encephalomyelitis, Autoimmune, Experimental/genetics , Encephalomyelitis, Autoimmune, Experimental/immunology , Encephalomyelitis, Autoimmune, Experimental/metabolism , Female , Gene Expression Regulation , Greece , HLA-DR Antigens/genetics , HLA-DR Antigens/metabolism , Humans , Interleukin-2/metabolism , Interleukin-2 Receptor alpha Subunit/metabolism , Lymphocyte Activation/drug effects , Male , Mice, Inbred C57BL , Mice, Transgenic , Middle Aged , Myelin Basic Protein/genetics , Myelin Basic Protein/metabolism , Myeloid Cells/immunology , Myeloid Cells/metabolism , Spinal Cord/immunology , Spinal Cord/metabolism , Spinal Cord/pathology , Synaptosomal-Associated Protein 25/genetics , Synaptosomal-Associated Protein 25/metabolism , T-Lymphocytes/immunology , T-Lymphocytes/metabolism , Young AdultABSTRACT
Mannan (polysaccharide) conjugated with a myelin oligodendrocyte glycoprotein (MOG) peptide, namely (KG)5MOG35-55, represents a potent and promising new approach for the immunotherapy of Multiple Sclerosis (MS). The MOG35-55 epitope conjugated with the oxidized form of mannan (poly-mannose) via a (KG)5 linker was found to inhibit the symptoms of MOG35-55-induced experimental autoimmune encephalomyelitis (EAE) in mice using prophylactic and therapeutic vaccinated protocols. Deamidation is a common modification in peptide and protein sequences, especially for Gln and Asn residues. In this study, the structural solution motif of deaminated peptides and their functional effects in an animal model for MS were explored. Several peptides based on the MOG35-55 epitope have been synthesized in which the Asn53 was replaced with Ala, Asp, or isoAsp. Our results demonstrate that the synthesized MOG peptides were formed to the deaminated products in basic conditions, and the Asn53 was mainly modified to Asp. Moreover, both peptides (wild type and deaminated derivative) conjugated with mannan (from Saccharomyces cerevisiae) independently inhibited the development of neurological symptoms and inflammatory demyelinating spinal cord lesions in MOG35-55-induced EAE. To conclude, mannan conjugated with a deamidated product did not affect the efficacy of the parent peptide.
