Exploiting the preferential phagocytic uptake of nanoparticle-antigen conjugates for the effective treatment of autoimmunity.

Tolerance induction is central to the suppression of autoimmunity. Here, we engineered the preferential uptake of nano-conjugated autoantigens by spleen-resident macrophages to re-introduce self-tolerance and suppress autoimmunity. The brain autoantigen, myelin oligodendrocyte glycoprotein (MOG), was conjugated to 200 or 500 nm silica nanoparticles (SNP) and delivered to the spleen and liver-resident macrophages of experimental autoimmune encephalomyelitis (EAE) mice, used as a model of multiple sclerosis. MOG-SNP conjugates significantly reduced signs of EAE at a very low dose (50 µg) compared to the higher dose (>800 µg) of free-MOG. This was associated with reduced proliferation of splenocytes and pro-inflammatory cytokines secretion, decreased spinal cord inflammation, demyelination and axonal damage. Notably, biodegradable porous SNP showed an enhanced disease suppression assisted by elevated levels of regulatory T cells and programmed-death ligands (PD-L1/2) in splenic and lymph node cells. Our results demonstrate that targeting nano-conjugated autoantigens to tissue-resident macrophages in lymphoid organs can effectively suppress autoimmunity.

Functional characterization of reappearing B cells after anti-CD20 treatment of CNS autoimmune disease.

The anti-CD20 antibody ocrelizumab, approved for treatment of multiple sclerosis, leads to rapid elimination of B cells from the blood. The extent of B cell depletion and kinetics of their recovery in different immune compartments is largely unknown. Here, we studied how anti-CD20 treatment influences B cells in bone marrow, blood, lymph nodes, and spleen in models of experimental autoimmune encephalomyelitis (EAE). Anti-CD20 reduced mature B cells in all compartments examined, although a subpopulation of antigen-experienced B cells persisted in splenic follicles. Upon treatment cessation, CD20+ B cells simultaneously repopulated in bone marrow and spleen before their reappearance in blood. In EAE induced by native myelin oligodendrocyte glycoprotein (MOG), a model in which B cells are activated, B cell recovery was characterized by expansion of mature, differentiated cells containing a high frequency of myelin-reactive B cells with restricted B cell receptor gene diversity. Those B cells served as efficient antigen-presenting cells (APCs) for activation of myelin-specific T cells. In MOG peptide-induced EAE, a purely T cell-mediated model that does not require B cells, in contrast, reconstituting B cells exhibited a naive phenotype without efficient APC capacity. Our results demonstrate that distinct subpopulations of B cells differ in their sensitivity to anti-CD20 treatment and suggest that differentiated B cells persisting in secondary lymphoid organs contribute to the recovering B cell pool.

Redirecting adult mesenchymal stromal cells to the brain: a new approach for treating CNS autoimmunity and neuroinflammation?

Mesenchymal stromal cells or stem cells (MSCs) have been shown to participate in tissue repair and are immunomodulatory in neuropathological settings. Given this, their potential use in developing a new generation of personalized therapies for autoimmune and inflammatory diseases of the central nervous system (CNS) will be explored. To effectively exert these effector functions, MSCs must first gain entry into damaged neural tissues, a process that has been demonstrated to be a limiting factor in their therapeutic efficacy. In this review, we discuss approaches to maximize the therapeutic efficacy of MSCs by altering their intrinsic trafficking programs to effectively enter neuropathological sites. To this end, we explore the significant role of chemokine receptors and adhesion molecules in directing cellular traffic to the inflamed CNS and the capacity of MSCs to adopt these molecular mechanisms to gain entry to this site. We postulate that understanding and exploiting these migratory mechanisms may be key to the development of cell-based therapies tailored to respond to the migratory cues unique to the nature and stage of progression of individual CNS disorders.

Treatment of spontaneous EAE by laquinimod reduces Tfh, B cell aggregates, and disease progression.

OBJECTIVE: To evaluate the influence of oral laquinimod, a candidate multiple sclerosis (MS) treatment, on induction of T follicular helper cells, development of meningeal B cell aggregates, and clinical disease in a spontaneous B cell-dependent MS model. METHODS: Experimental autoimmune encephalomyelitis (EAE) was induced in C57BL/6 mice by immunization with recombinant myelin oligodendrocyte glycoprotein (rMOG) protein. Spontaneous EAE was evaluated in C57BL/6 MOG p35-55-specific T cell receptor transgenic (2D2) × MOG-specific immunoglobulin (Ig)H-chain knock-in (IgHMOG-ki [Th]) mice. Laquinimod was administered orally. T cell and B cell populations were examined by flow cytometry and immunohistochemistry. RESULTS: Oral laquinimod treatment (1) reduced CD11c+CD4+ dendritic cells, (2) inhibited expansion of PD-1+CXCR5+BCL6+ T follicular helper and interleukin (IL)-21-producing activated CD4+CD44+ T cells, (3) suppressed B cell CD40 expression, (4) diminished formation of Fas+GL7+ germinal center B cells, and (5) inhibited development of MOG-specific IgG. Laquinimod treatment not only prevented rMOG-induced EAE, but also inhibited development of spontaneous EAE and the formation of meningeal B cell aggregates. Disability progression was prevented when laquinimod treatment was initiated after mice developed paralysis. Treatment of spontaneous EAE with laquinimod was also associated with increases in CD4+CD25hiFoxp3+ and CD4+CD25+IL-10+ regulatory T cells. CONCLUSIONS: Our observations that laquinimod modulates myelin antigen-specific B cell immune responses and suppresses both development of meningeal B cell aggregates and disability progression in spontaneous EAE should provide insight regarding the potential application of laquinimod to MS treatment. Results of this investigation demonstrate how the 2D2 × Th spontaneous EAE model can be used successfully for preclinical evaluation of a candidate MS treatment.

B cell-derived transforming growth factor-ß1 expression limits the induction phase of autoimmune neuroinflammation.

Studies in experimental autoimmune encephalomyelitis (EAE), a murine model of multiple sclerosis (MS), have shown that regulatory B cells modulate the course of the disease via the production of suppressive cytokines. While data indicate a role for transforming growth factor (TGF)-ß1 expression in regulatory B cell functions, this mechanism has not yet been tested in autoimmune neuroinflammation. Transgenic mice deficient for TGF-ß1 expression in B cells (B-TGF-ß1-/-) were tested in EAE induced by recombinant mouse myelin oligodendrocyte glycoprotein (rmMOG). In this model, B-TGF-ß1-/- mice showed an earlier onset of neurologic impairment compared to their littermate controls. Exacerbated EAE susceptibility in B-TGF-ß1-/- mice was associated with augmented CNS T helper (Th)1/17 responses. Moreover, selective B cell TGF-ß1-deficiency increased the frequencies and activation of myeloid dendritic cells, potent professional antigen-presenting cells (APCs), suggesting that B cell-derived TGF-ß1 can constrain Th1/17 responses through inhibition of APC activity. Collectively our data suggest that B cells can down-regulate the function of APCs, and in turn encephalitogenic Th1/17 responses, via TGF-ß1, findings that may be relevant to B cell-targeted therapies.

CNS accumulation of regulatory B cells is VLA-4-dependent.

OBJECTIVE: To investigate the role of very late antigen-4 (VLA-4) on regulatory B cells (Breg) in CNS autoimmune disease. METHODS: Experimental autoimmune encephalomyelitis (EAE) was induced in mice selectively deficient for VLA-4 on B cells (CD19cre/α4(f/f)) by immunization with myelin oligodendrocyte glycoprotein (MOG) peptide (p)35-55 or recombinant human (rh) MOG protein. B-cell and T-cell populations were examined by flow cytometry and immunohistochemistry. Breg were evaluated by intracellular IL-10 staining of B cells and, secondly, by coexpression of CD1d and CD5. RESULTS: As previously reported, EAE was less severe in B-cell VLA-4-deficient vs control CD19cre mice when induced by rhMOG, a model that is B-cell-dependent and leads to efficient B-cell activation and antibody production. Paradoxically, B-cell VLA-4-deficient mice developed more severe clinical disease than control mice when EAE was induced with MOG p35-55, a B-cell-independent encephalitogen that does not efficiently activate B cells. Peripheral T-cell and humoral immune responses were not altered in B-cell VLA-4-deficient mice. In MOG p35-55-induced EAE, B-cell VLA-4 deficiency reduced CNS accumulation of B but not T cells. Breg were detected in the CNS of control mice with MOG p35-55-induced EAE. However, more severe EAE in B-cell VLA-4-deficient mice was associated with virtual absence of CNS Breg. CONCLUSIONS: Our results demonstrate that CNS accumulation of Breg is VLA-4-dependent and suggest that Breg may contribute to regulation of CNS autoimmunity in situ. These observations underscore the need to choose the appropriate encephalitogen when studying how B cells contribute to pathogenesis or regulation of CNS autoimmunity.

Myelin-reactive antibodies initiate T cell-mediated CNS autoimmune disease by opsonization of endogenous antigen.

In the pathogenesis of central nervous system (CNS) demyelinating disorders, antigen-specific B cells are implicated to act as potent antigen-presenting cells (APC), eliciting waves of inflammatory CNS infiltration. Here, we provide the first evidence that CNS-reactive antibodies (Ab) are similarly capable of initiating an encephalitogenic immune response by targeting endogenous CNS antigen to otherwise inert myeloid APC. In a transgenic mouse model, constitutive production of Ab against myelin oligodendrocyte glycoprotein (MOG) was sufficient to promote spontaneous experimental autoimmune encephalomyelitis (EAE) in the absence of B cells, when mice endogenously contained MOG-recognizing T cells. Adoptive transfer studies corroborated that anti-MOG Ab triggered activation and expansion of peripheral MOG-specific T cells in an Fc-dependent manner, subsequently causing EAE. To evaluate the underlying mechanism, anti-MOG Ab were added to a co-culture of myeloid APC and MOG-specific T cells. At otherwise undetected concentrations, anti-MOG Ab enabled Fc-mediated APC recognition of intact MOG; internalized, processed and presented MOG activated naïve T cells to differentiate in an encephalitogenic manner. In a series of translational experiments, anti-MOG Ab from two patients with an acute flare of CNS inflammation likewise facilitated detection of human MOG. Jointly, these observations highlight Ab-mediated opsonization of endogenous CNS auto-antigen as a novel disease- and/or relapse-triggering mechanism in CNS demyelinating disorders.

Graphene Functionalized Scaffolds Reduce the Inflammatory Response and Supports Endogenous Neuroblast Migration when Implanted in the Adult Brain.

Electroactive materials have been investigated as next-generation neuronal tissue engineering scaffolds to enhance neuronal regeneration and functional recovery after brain injury. Graphene, an emerging neuronal scaffold material with charge transfer properties, has shown promising results for neuronal cell survival and differentiation in vitro. In this in vivo work, electrospun microfiber scaffolds coated with self-assembled colloidal graphene, were implanted into the striatum or into the subventricular zone of adult rats. Microglia and astrocyte activation levels were suppressed with graphene functionalization. In addition, self-assembled graphene implants prevented glial scarring in the brain 7 weeks following implantation. Astrocyte guidance within the scaffold and redirection of neuroblasts from the subventricular zone along the implants was also demonstrated. These findings provide new functional evidence for the potential use of graphene scaffolds as a therapeutic platform to support central nervous system regeneration.

Immunosuppressive potential of human amnion epithelial cells in the treatment of experimental autoimmune encephalomyelitis.

BACKGROUND: Multiple sclerosis (MS) is an autoimmune inflammatory disease of the central nervous system (CNS). In recent years, it has been found that cells such as human amnion epithelial cells (hAECs) have the ability to modulate immune responses in vitro and in vivo and can differentiate into multiple cell lineages. Accordingly, we investigated the immunoregulatory effects of hAECs as a potential therapy in an MS-like disease, EAE (experimental autoimmune encephalomyelitis), in mice. METHODS: Using flow cytometry, the phenotypic profile of hAECs from different donors was assessed. The immunomodulatory properties of hAECs were examined in vitro using antigen-specific and one-way mixed lymphocyte proliferation assays. The therapeutic efficacy of hAECs was examined using a relapsing-remitting model of EAE in NOD/Lt mice. T cell responsiveness, cytokine secretion, T regulatory, and T helper cell phenotype were determined in the peripheral lymphoid organs and CNS of these animals. RESULTS: In vitro, hAECs suppressed both specific and non-specific T cell proliferation, decreased pro-inflammatory cytokine production, and inhibited the activation of stimulated T cells. Furthermore, T cells retained their naïve phenotype when co-cultured with hAECs. In vivo studies revealed that hAECs not only suppressed the development of EAE but also prevented disease relapse in these mice. T cell responses and production of the pro-inflammatory cytokine interleukin (IL)-17A were reduced in hAEC-treated mice, and this was coupled with a significant increase in the number of peripheral T regulatory cells and naïve CD4+ T cells. Furthermore, increased proportions of Th2 cells in the peripheral lymphoid organs and within the CNS were observed. CONCLUSION: The therapeutic effect of hAECs is in part mediated by inducing an anti-inflammatory response within the CNS, demonstrating that hAECs hold promise for the treatment of autoimmune diseases like MS.

Lithium chloride improves the efficiency of induced pluripotent stem cell-derived neurospheres.

Induced pluripotent stem cell (iPSC)-derived neurospheres, which consist mainly of neural progenitors, are considered to be a good source of neural cells for transplantation in regenerative medicine. In this study, we have used lithium chloride, which is known to be a neuroprotective agent, in an iPSC-derived neurosphere model, and examined both the formation rate and size of the neurospheres as well as the proliferative and apoptotic status of their contents. Our results showed that lithium enhanced the formation and the sizes of the iPSC-derived neurospheres, increased the number of Ki67-positive proliferating cells, but reduced the number of the TUNEL-positive apoptotic cells. This increased number of Ki67 proliferating cells was secondary to the decreased apoptosis and not to the stimulation of cell cycle entry, as the expression of the proliferation marker cyclin D1 mRNA did not change after lithium treatment. Altogether, we suggest that lithium enhances the survival of neural progenitors and thus the quality of the iPSC-derived neurospheres, which may strengthen the prospect of using lithium-treated pluripotent cells and their derivatives in a clinical setting.

B-cell very late antigen-4 deficiency reduces leukocyte recruitment and susceptibility to central nervous system autoimmunity.

Natalizumab, which binds very late antigen-4 (VLA-4), is a potent therapy for multiple sclerosis (MS). Studies have focused primarily upon its capacity to interfere with T-cell migration into the central nervous system (CNS). B cells are important in MS pathogenesis and express high levels of VLA-4. Here, we report that the selective inhibition of VLA-4 expression on B cells impedes CNS accumulation of B cells, and recruitment of Th17 cells and macrophages, and reduces susceptibility to experimental autoimmune encephalomyelitis. These results underscore the importance of B-cell VLA-4 expression in the pathogenesis of CNS autoimmunity and provide insight regarding mechanisms that may contribute to the benefit of natalizumab in MS, as well as candidate therapeutics that selectively target B cells.

In vivo multi-modal imaging of experimental autoimmune uveoretinitis in transgenic reporter mice reveals the dynamic nature of inflammatory changes during disease progression.

BACKGROUND: Experimental autoimmune uveoretinitis (EAU) is a widely used experimental animal model of human endogenous posterior uveoretinitis. In the present study, we performed in vivo imaging of the retina in transgenic reporter mice to investigate dynamic changes in exogenous inflammatory cells and endogenous immune cells during the disease process. METHODS: Transgenic mice (C57Bl/6 J Cx 3 cr1 (GFP/+) , C57Bl/6 N CD11c-eYFP, and C57Bl/6 J LysM-eGFP) were used to visualize the dynamic changes of myeloid-derived cells, putative dendritic cells and neutrophils during EAU. Transgenic mice were monitored with multi-modal fundus imaging camera over five time points following disease induction with the retinal auto-antigen, interphotoreceptor retinoid binding protein (IRBP1-20). Disease severity was quantified with both clinical and histopathological grading. RESULTS: In the normal C57Bl/6 J Cx 3 cr1 (GFP/+) mouse Cx3cr1-expressing microglia were evenly distributed in the retina. In C57Bl/6 N CD11c-eYFP mice clusters of CD11c-expressing cells were noted in the retina and in C57Bl/6 J LysM-eGFP mice very low numbers of LysM-expressing neutrophils were observed in the fundus. Following immunization with IRBP1-20, fundus examination revealed accumulations of Cx3cr1-GFP(+) myeloid cells, CD11c-eYFP(+) cells and LysM-eGFP(+) myelomonocytic cells around the optic nerve head and along retinal vessels as early as day 14 post-immunization. CD11c-eYFP(+) cells appear to resolve marginally earlier (day 21 post-immunization) than Cx3cr1-GFP(+) and LysM-eGFP(+) cells. The clinical grading of EAU in transgenic mice correlated closely with histopathological grading. CONCLUSIONS: These results illustrate that in vivo fundus imaging of transgenic reporter mice allows direct visualization of various exogenously and endogenously derived leukocyte types during EAU progression. This approach acts as a valuable adjunct to other methods of studying the clinical course of EAU.

Application of human induced pluripotent stem cells for modeling and treating neurodegenerative diseases.

The advent of human induced pluripotent stem cells (hiPSCs), reprogrammed in vitro from both healthy and disease-state human somatic cells, has triggered an enormous global research effort to realize personalized regenerative medicine for numerous degenerative conditions. hiPSCs have been generated from cells of many tissue types and can be differentiated in vitro to most somatic lineages, not only for the establishment of disease models that can be utilized as novel drug screening platforms and to study the molecular and cellular processes leading to degeneration, but also for the in vivo cell-based repair or modulation of a patient's disease profile. hiPSCs derived from patients with the neurodegenerative diseases amyotrophic lateral sclerosis, Parkinson's disease, Alzheimer's disease and multiple sclerosis have been successfully differentiated in vitro into disease-relevant cell types, including motor neurons, dopaminergic neurons and oligodendrocytes. However, the generation of functional iPSC-derived neural cells that are capable of engraftment in humans and the identification of robust disease phenotypes for modeling neurodegeneration still require several key challenges to be addressed. Here, we discuss these challenges and summarize recent progress toward the application of iPSC technology for these four common neurodegenerative diseases.

Single ß³-amino acid substitutions to MOG peptides suppress the development of experimental autoimmune encephalomyelitis.

CD4(+) T-cells play a key role in the pathogenesis of multiple sclerosis (MS). Altered peptide ligands capable of modulating T-cell autoreactivity are considered a promising strategy for development of antigen-specific therapies for MS. Since peptides are inherently unstable, the current study explored single ß-amino acid substitution as a means of stabilizing an epitope of myelin oligodendrocyte glycoprotein. ß-Amino acid substitution at position 44, the major T-cell receptor contact residue, increased the half-life of active metabolites. Vaccination with one altered peptide, MOG44ßF, conferred protection from EAE, decreased T-cell autoreactivity and pro-inflammatory cytokine production. Additional studies using MOG44ßF in an oral treatment regimen, administered after EAE induction, also attenuated disease severity. Thus, altered peptides such as those reported here may lead to the development of novel and more specific treatments for MS.

Intrathecal anti-CD20 efficiently depletes meningeal B cells in CNS autoimmunity.

Clinical trials revealed that systemic administration of B-cell-depleting anti-CD20 antibodies can hold lesion formation in the early relapsing-remitting phase of multiple sclerosis (MS). Throughout the secondary-progressive (SP) course of MS, pathogenic B cells may, however, progressively replicate within the central nervous system (CNS) itself, which is largely inaccessible to systemic anti-CD20 treatment. Utilizing the murine MS model of experimental autoimmune encephalomyelitis, we show that intrathecal (i.t.) administration of anti-CD20 alone very efficiently depletes meningeal B cells from established CNS lesions. In SP-MS patients, adding i.t. administration of anti-CD20 to its systemic use may be a valuable strategy to target pathogenic B-cell function.

MOG transmembrane and cytoplasmic domains contain highly stimulatory T-cell epitopes in MS.

OBJECTIVE: Recently, we reported that the 218 amino acid murine full-length myelin oligodendrocyte glycoprotein (MOG) contains novel T-cell epitopes p119-132, p181-195, and p186-200, located within its transmembrane and cytoplasmic domains, and that p119-132 is its immunodominant encephalitogenic T-cell epitope in mice. Here, we investigated whether the corresponding human MOG sequences contain T-cell epitopes in patients with multiple sclerosis (MS) and healthy controls (HC). METHODS: Peripheral blood T cells from patients with MS and HC were examined for proliferation to MOG p119-130, p181-195, p186-200, and p35-55 by fluorescence-activated cell sorting analysis using carboxylfluorescein diacetate succinimidyl ester dilution assay. Intracellular production of proinflammatory cytokines was analyzed by flow cytometry. RESULTS: MOG p119-130, p181-195, and p186-200 elicited significantly greater T-cell responses than p35-55 in patients with MS. T cells from patients with MS proliferated significantly more strongly to MOG p119-130 and p186-200 than did T cells from HC. Further, MOG p119-130-specific T cells exhibited Th17 polarization, suggesting this T-cell epitope may be relevant to MS pathogenesis. CONCLUSIONS: Transmembrane and cytoplasmic MOG domains contain potent T-cell epitopes in MS. Recognition of these determinants is important when evaluating T-cell responses to MOG in MS and may have implications for development of myelin antigen-based therapeutics.

Immunodominant T-cell epitopes of MOG reside in its transmembrane and cytoplasmic domains in EAE.

OBJECTIVE: Studies evaluating T-cell recognition of myelin oligodendrocyte glycoprotein (MOG) in multiple sclerosis (MS) and its model, experimental autoimmune encephalomyelitis (EAE), have focused mostly on its 117 amino acid (aa) extracellular domain, especially peptide (p) 35-55. We characterized T-cell responses to the entire 218 aa MOG sequence, including its transmembrane and cytoplasmic domains. METHODS: T-cell recognition in mice was examined using overlapping peptides and intact full-length mouse MOG. EAE was evaluated by peptide immunization and by adoptive transfer of MOG epitope-specific T cells. Frequency of epitope-specific T cells was examined by ELISPOT. RESULTS: Three T-cell determinants of MOG were discovered in its transmembrane and cytoplasmic domains, p119-132, p181-195, and p186-200. Transmembrane MOG p119-132 induced clinical EAE, CNS inflammation, and demyelination as potently as p35-55 in C57BL/6 mice and other H-2(b) strains. p119-128 contained its minimal encephalitogenic epitope. p119-132 did not cause disease in EAE-susceptible non-H-2(b) strains, including Biozzi, NOD, and PL/J. MOG p119-132-specific T cells produced Th1 and Th17 cytokines and transferred EAE to wild-type recipient mice. After immunization with full-length MOG, a significantly higher frequency of MOG-reactive T cells responded to p119-132 than to p35-55, demonstrating that p119-132 is an immunodominant encephalitogenic epitope. MOG p181-195 did not cause EAE, and MOG p181-195-specific T cells could not transfer EAE into wild-type or highly susceptible T- and B-cell-deficient mice. CONCLUSIONS: Transmembrane and cytoplasmic domains of MOG contain immunodominant T-cell epitopes in EAE. A CNS autoantigen can also contain nonpathogenic stimulatory T-cell epitopes. Recognition that a myelin antigen contains multiple encephalitogenic and nonencephalitogenic determinants may have implications for therapeutic development in MS.

Fine structure of neurally differentiated iPS cells generated from a multiple sclerosis (MS) patient: a case study.

We compared the characteristics of neural cells derived from induced pluripotent stem (iPS) cells from a patient with multiple sclerosis versus neurally differentiated control iPS cells of a healthy individual. The iPS cells were differentiated toward the oligodendrocyte lineage using a four-step protocol established for the differentiation of embryonic stem cells. The resulting cell population was immunostained on day 112 of differentiation for the presence of oligodendrocytes and analyzed by transmission electron microscopy (TEM). Both patient and control samples resembled a mixed population of neural cells rather than oligodendroglia of high purity, including neural stem cell-like cells and possibly oligodendrocytes demonstrable by TEM.

Quantitative proteome profiling of CNS-infiltrating autoreactive CD4+ cells reveals selective changes during experimental autoimmune encephalomyelitis.

Experimental autoimmune encephalomyelitis (EAE) is a murine model of multiple sclerosis, a chronic neurodegenerative and inflammatory autoimmune condition of the central nervous system (CNS). Pathology is driven by the infiltration of autoreactive CD4(+) lymphocytes into the CNS, where they attack neuronal sheaths causing ascending paralysis. We used an isotope-coded protein labeling approach to investigate the proteome of CD4(+) cells isolated from the spinal cord and brain of mice at various stages of EAE progression in two EAE disease models: PLP139-151-induced relapsing-remitting EAE and MOG35-55-induced chronic EAE, which emulate the two forms of human multiple sclerosis. A total of 1120 proteins were quantified across disease onset, peak-disease, and remission phases of disease, and of these 13 up-regulated proteins of interest were identified with functions relating to the regulation of inflammation, leukocyte adhesion and migration, tissue repair, and the regulation of transcription/translation. Proteins implicated in processes such as inflammation (S100A4 and S100A9) and tissue repair (annexin A1), which represent key events during EAE progression, were validated by quantitative PCR. This is the first targeted analysis of autoreactive cells purified from the CNS during EAE, highlighting fundamental CD4(+) cell-driven processes that occur during the initiation of relapse and remission stages of disease.

Molecular grafting onto a stable framework yields novel cyclic peptides for the treatment of multiple sclerosis.

Multiple sclerosis (MS) is an inflammatory disease of the central nervous system (CNS) and is characterized by the destruction of myelin and axons leading to progressive disability. Peptide epitopes from CNS proteins, such as myelin oligodendrocyte glycoprotein (MOG), possess promising immunoregulatory potential for treating MS; however, their instability and poor bioavailability is a major impediment for their use clinically. To overcome this problem, we used molecular grafting to incorporate peptide sequences from the MOG35-55 epitope onto a cyclotide, which is a macrocyclic peptide scaffold that has been shown to be intrinsically stable. Using this approach, we designed novel cyclic peptides that retained the structure and stability of the parent scaffold. One of the grafted peptides, MOG3, displayed potent ability to prevent disease development in a mouse model of MS. These results demonstrate the potential of bioengineered cyclic peptides for the treatment of MS.