Abigail C. Buenafe

Treatment of multiple sclerosis with T–cell receptor peptides: Results of a double–blind pilot trial

A T–cell receptor (TCR) peptide vaccine from the Vβ5.2 sequence expressed in multiple sclerosis (MS) plaques and on myelin basic protein (MBP)–specific T cells boosted peptide–reactive T cells in patients with progressive MS. Vaccine responders had a reduced MBP response and remained clinically stable without side effects during one year of therapy, whereas nonresponders had an increased MBP response and progressed clinically. Peptide–specific T helper 2 cells directly inhibited MBP–specific T helper 1 cells in vitro through the release of interleukin–10, implicating a bystander suppression mechanism that holds promise for treatment of MS and other autoimmune diseases.

Estrogen inhibition of EAE involves effects on dendritic cell function

Estrogen has been found to have suppressive effects on the induction of experimental autoimmune encephalomyelitis (EAE), an animal model for the human disease multiple sclerosis. We have investigated the effects of 17β‐estradiol (E2) treatment on dendritic cells (DCs) in two different mouse models of EAE. The frequency of CD11b+/CD11c+ DCs was significantly decreased in the brain of mice protected from EAE induction by E2 treatment. In addition, the frequency of CD11c+/CD8α+ DCs producing tumor necrosis factor (TNF)α and interferon (IFN)γ in the spleen of E2‐treated mice was dramatically decreased compared to that in control mice with EAE, demonstrating an effect of E2 on DC function. In order to examine E2 effects on DCs in more detail, splenic DCs were cultured in the presence of granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) and interleukin (IL)‐4to promote maturation. E2 pretreatment was found to suppress the ability of cultured DCs bearing a mature phenotype to present Ag to myelin basic protein (MBP)‐specific T cells. Analysis of cytokine production demonstrated that E2 decreased TNFα, IFNγ and IL‐12 production in mature DCs. In addition, MBP‐specific T cells cocultured with E2‐pretreated mature DCs in the presence of antigen demonstrated a shift towards production of Th2 cytokines IL‐4 and IL‐10 and a concomitant decrease in the production of Th1 cytokines TNFα and IFNγ. Thus, E2 treatment appears to have multiple effects on the DC population, which may contribute to a down‐regulation or block in the activation of Th1 cells involved in the induction of EAE.

Multiple sclerosis: the frequency of allelic forms of tumor necrosis factor and lymphotoxin-alpha

The cytokines LTa and TNF have been implicated as major mediators of tissue injury in multiple sclerosis (MS). In this study we have assessed the frequency of specific polymorphisms for these genes in MS (n = 53) and controls (n = 81) using a highly sensitive, two stage nested polymerase chain reaction (PCR), with the second stage using mutation-specific primers. Genomic DNA was extracted from blood cells and the results confirmed by direct dideoxy chain termination sequencing. The frequency of the -308 G to A mutation in the TNF promoter region in normal controls was 15% and in MS was 24%. For LTa gene the exon 3 polymorphism allele A was detected in 36% of controls and 34% of the MS patients. In MS, the combined genotype TNF G + A and LTa C + C was present 6 times more frequently (12%) than in controls (2%), and patients with this genotype showed the highest EDSS scores. We found the TNF and LTa polymorphisms to occur independently from the HLA class II DR2 allele distribution in MS. Whilst the G - A polymorphism in TNF gene promoter has been studied previously in MS, with conflicting results, this is the first study that has addressed the exon 3 polymorphism in LTa in MS. The results indicate that this polymorphism is not linked with the higher genetic predisposition for MS, but that combined TNF G + A and LTa C + C genotype might contribute to development of the disease.

Lipopolysaccharide pretreatment modulates the disease course in experimental autoimmune encephalomyelitis

Treatment with the bacterial product lipopolysaccharide (LPS) prior to the induction of experimental autoimmune encephalomyelitis (EAE) consistently led to a delayed onset of disease but not to a reduction in disease severity. T cell proliferation was reduced in LPS-treated mice, due at least in part to a loss in antigen presenting cell function. T cell and macrophage infiltration into the CNS was delayed and TNFalpha production was diminished in LPS pre-treated mice, consistent with the delay in disease onset. Real-time PCR analysis of gene expression in the CNS of LPS or saline pre-treated mice demonstrated an early induction of TNFalpha, TGFbeta, IFNbeta, and SOCS3 in the LPS pre-treated mice. Thus, exposure to LPS prior to EAE induction affects antigen presentation and may modulate the expression of inflammatory regulators that impact the autoimmune disease course.

Vβ CDR3 motifs associated with BP recognition are enriched in OX-40+ spinal cord T cells of Lewis rats with EAE

Spinal cord (SC) T cells were isolated at the onset of actively induced experimental autoimmune encephalomyelitis (EAE) and sorted for the presence of the OX‐40 activation marker. Previously, we reported an enhanced bias in Vβ8.2 expression as well as enhanced proliferative responses to basic protein antigens among the OX‐40+ SC T cells. Here we demonstrate that CDR3 motifs associated with EAE are present at a significantly higher frequency in Vβ8.2 sequences of OX‐40+ SC T cells (16/17) compared with those of OX‐40− SC T cells (5/17). Thus, the OX‐40 antigen may be useful as a marker to isolate and characterize autoantigen‐specific T cells from the site of inflammation in T‐cell‐mediated autoimmune diseases.

A novel regulatory pathway for autoimmune disease: Binding of partial MHC class II constructs to monocytes reduces CD74 expression and induces both specific and bystander T-cell tolerance

Treatment with partial (p)MHC class II-β1α1 constructs (also referred to as recombinant T-cell receptor ligands - RTL) linked to antigenic peptides can induce T-cell tolerance, inhibit recruitment of inflammatory cells and reverse autoimmune diseases. Here we demonstrate a novel regulatory pathway that involves RTL binding to CD11b(+) mononuclear cells through a receptor comprised of MHC class II invariant chain (CD74), cell-surface histones and MHC class II itself for treatment of experimental autoimmune encephalomyelitis (EAE). Binding of RTL constructs with CD74 involved a previously unrecognized MHC class II-α1/CD74 interaction that inhibited CD74 expression, blocked activity of its ligand, macrophage migration inhibitory factor, and reduced EAE severity. These findings implicate binding of RTL constructs to CD74 as a key step in both antigen-driven and bystander T-cell tolerance important in treatment of inflammatory diseases.

Specificity of regulatory CD4+CD25+ T cells for self-T cell receptor determinants.

Although the phenotypic and regulatory properties of the CD4+CD25+ T cell lineage (Treg cells) have been well described, the specificities remain largely unknown. We demonstrate here that the CD4+CD25+ Treg population includes the recognition of a broad spectrum of human TCR CDR2 determinants found in the germline V gene repertoire as well as that of a clonotypic nongermline‐encoded CDR3β sequence present in a recombinant soluble T cell receptor (TCR) protein. Regulatory activity was demonstrated in T cell lines responsive to TCR but not in T cell lines responsive to control antigens. Inhibitory activity of TCR‐reactive T cells required cell–cell contact and involved CTLA‐4, GITR, IL‐10, and IL‐17. Thus, the T–T regulatory network includes Treg cells with specificity directed toward self‐TCR determinants.

IL-7 enhances Ag-specific human T cell response by increasing expression of IL-2R α and γ chains

Abstract Interleukin-7 has demonstrated potent enhancing effects on the growth and differentiation of several immature cell types, including thymocytes, and on survival of resting and antigen activated T cells. In this study, we evaluated the effects of IL-7 on post-thymic antigen-specific T cells from human blood. IL-7 was found to enhance proliferation responses and IFN-γ secretion of myelin or recall Ag-specific Th1 cells through the selective up-regulation of the IL-2Rα and γ but not β chains in both an Ag-dependent and Ag-independent manner, but did not affect monocytes, B cells, or NK cells. These functions of IL-7 enhanced the detection of Th1 but not Th2 cell frequency by >2.5 fold, and promoted selection of Ag-specific Th1 cells by the limiting dilution method. Moreover, IL-7 pretreatment conferred increased resistance of CD4+ T cells to CD8+ cell lysis. These studies demonstrate that IL-7 promotes the growth and survival of circulating Ag-specific human Th1 cells through a mechanism that probably involves the γc common receptor for IL-2 family members that includes IL-7.

Preformed CD40L Is Stored in Th1, Th2, Th17, and T Follicular Helper Cells as Well as CD4+8− Thymocytes and Invariant NKT Cells but Not in Treg Cells

CD40L is essential for the development of adaptive immune responses. It is generally thought that CD40L expression in CD4+ T cells is regulated transcriptionally and made from new mRNA following antigen recognition. However, imaging studies show that the majority of cognate interactions between effector CD4+ T cells and APCs in vivo are too short to allow de novo CD40L synthesis. We previously showed that Th1 effector and memory cells store preformed CD40L (pCD40L) in lysosomal compartments and mobilize it onto the plasma membrane immediately after antigenic stimulation, suggesting that primed CD4+ T cells may use pCD40L to activate APCs during brief encounters. Indeed, our recent study showed that pCD40L is sufficient to mediate selective activation of cognate B cells and trigger DC activation in vitro. In this study, we show that pCD40L is present in Th1 and follicular helper T cells developed during infection with lymphocytic choriomeningitis virus, Th2 cells in the airway of asthmatic mice, and Th17 cells from the CNS of animals with experimental autoimmune encephalitis (EAE). pCD40L is nearly absent in both natural and induced Treg cells, even in the presence of intense inflammation such as occurs in EAE. We also found pCD40L expression in CD4 single positive thymocytes and invariant NKT cells. Together, these results suggest that pCD40L may function in T cell development as well as an unexpectedly broad spectrum of innate and adaptive immune responses, while its expression in Treg cells is repressed to avoid compromising their suppressive activity.

MHC‐restriction, cytokine profile, and immunoregulatory effects of human T cells specific for TCR Vβ CDR2 peptides: Comparison with myelin basic protein‐specific T cells

HLA‐DR2+ patients with multiple sclerosis (MS) that respond to vaccination with TCR Vβ5.2‐38‐58 peptides have increased frequencies of TCR peptide‐specific T cells, reduced frequencies of myelin basic protein (MBP)‐specific T cells, and a better clinical course than non‐responders. To evaluate possible network regulation of MBP responses by TCR peptide‐specific T cells, we compared properties of both cell types. Both MBP‐ and TCR peptide‐specific T cell clones were CD4+ and predominantly HLA‐DR restricted. HLA‐DR2, which is in linkage disequilibrium in MS patients, preferentially restricted TCR peptide‐specific clones as well as MBP‐specific responses in HLA‐DR2 and DR2,3+ donors. Within the DR2 haplotype, however, both DRβ1*1501 and DRβ5*0101 alleles could restrict T cell responses to Vβ CDR2 peptides, whereas responses to MBP were restricted only by DRβ5*0101. TCR peptide‐specific clones expressed message for Th2 cytokines, including IL‐4, IL‐5, IL‐6, IL‐10, and TGF‐β, whereas MBP‐specific T cell clones expressed the Th1 cytokines IFN‐γ and IL‐2. Consistent with the Th2‐like cytokine profile, TCR peptide‐specific T cell clones expressed higher levels of CD30 than MBP‐specific T cells. Culture supernatants from TCR peptide‐specific T cell clones, but not from MBP‐ or Herpes simplex virus‐specific T cells, inhibited both proliferation responses and cytokine message production of MBP‐specific T cells. These results demonstrate distinct properties of MBP and TCR peptide‐specific T cells, and indicate that both target and bystander Th1 cells can be inhibited by Th2 cytokines secreted by activated TCR peptide‐specific T cells. These data support the rationale for TCR peptide vaccination to regulate pathogenic responses mediated by oligoclonal T cells in human autoimmune diseases.