M. Vainiene

MYELIN BASIC PROTEIN SPECIFIC T-HELPER CELLS INDUCE EXPERIMENTAL ANTERIOR UVEITIS

Immunopathological changes in the eyes were examined in Lewis rats after active and passive induction of experimental autoimmune encephalomyelitis (EAE) with myelin basic proteins (MBP) at various stages of EAE. The onset of anterior uveitis (AU) coincided with hind limb paralysis, but uveitis persisted after clinical signs of EAE had subsided. A mild form of uveitis was characteristic for the majority of rats. The changes within the iris and ciliary body consisted of an accumulation of inflammatory cells lining the anterior surface of iris, the trabecular meshwork, and, in some cases, within the ciliary body and the aqueous humor. A similar histopathological picture was observed when rats were injected with the secondary encephalitogenic determinant for Lewis rats, MBP peptide 87–99. Flow cytometry analysis of T cells from the anterior segment of the inflamed eyes after immunization with MBP revealed the presence of CD4+ cells exclusively expressing Vβ8.2 and OX‐40 markers. Our data suggest that MBP are encephalitogenic and uveitogenic in Lewis rats and that the Vβ8.2‐positive T cells in the eye represent encephalitogenic T cells. Many of those T cells were distributed in the iris and the anterior chamber. These findings indicate that these MBP‐specific T cells may play a critical role in EAE as well as in AU.

Prevention and treatment of relapsing autoimmune encephalomyelitis with myelin peptide-coupled splenocytes

Injection of antigen cross‐linked accessory cells has proven to be an efficient and highly selective approach for inducing epitope‐specific peripheral tolerance. This approach has been used successfully to inhibit induction of experimental autoimmune encephalomyelitis (EAE) and to dissect the relative dominance of component encephalitogenic determinants that contribute to both acute and relapsing EAE. In this study, we evaluated the tolerogenic effect of the dominant encephalitogenic epitope for SJL/J mice, residues 139–151 of myelin proteolipid protein (PLP), on the induction and relapses of EAE induced actively with PLP139‐151/CFA. Our results demonstrate the powerful protective effect of treating mice before induction of EAE with PLP139–151‐conjugated splenocytes (SPL) on the incidence and severity of both the initial episode and the first relapse of EAE. Moreover, treatment of mice on the first day of onset of clinical signs of EAE reduced the severity of the first relapse, apparently by reducing T cell recognition of PLP139–151, although no significant therapeutic effect was observed during the initial treated clinical episode. These data demonstrate the utility of using neuroantigen‐coupled accessory cells to prevent and treat relapsing EAE.

TCR peptide therapy decreases the frequency of encephalitogenic T cells in the periphery and the central nervous system

The V beta 8 CDR2 consensus peptide, residues 44-54, is highly effective in the treatment of clinical experimental autoimmune encephalomyelitis (EAE) in Lewis rats. To monitor immunological changes during EAE resulting from TCR peptide therapy, the frequencies of encephalitogenic and regulatory T cells were quantitated in lymph nodes, blood, and spinal cord. The frequency of T cells specific for basic protein and its major encephalitogenic epitope, residues 72-89, increased during EAE to about 1 cell per 100,000 lymph node or blood cells at the peak of clinical disease, and then declined. In contrast, the frequency of these T cells in spinal cord was highest, 50 per 100,000, prior to onset of clinical signs, and then decreased rapidly prior to spontaneous recovery. Injection of 100 micrograms of TCR V beta 8-44-54 peptide caused a decrease within 1-5 days in the frequencies of guinea pig basic-protein (GP-BP) and 72-89-reactive T cells in blood and spinal cord, and in the total number of infiltrating cells in spinal cord. In lymph nodes, 72-89-reactive T cells decreased as T cells specific for a protective epitope, residues 55-69 of GP-BP increased, suggesting epitope switching at the site of GP-BP immunization. Conversely, the frequency of T cells specific for the V beta 8-44-54 peptide increased, especially in blood and spinal cord, whereas T cell frequencies to control antigens were unchanged. These data document the critical presence of encephalitogenic T cells within the spinal cord during clinical EAE, and demonstrate that rapid and profound changes in T cell frequencies in the periphery and spinal cord are triggered by TCR peptide therapy.

Lymphokine mRNA expression in the spinal cords of Lewis rats with experimental autoimmune encephalomyelitis is associated with a host recruited CD45R hi/CD4+ population during recovery

To evaluate CD4+ T cell subpopulations involved in the induction and recovery from experimental autoimmune encephalomyelitis (EAE), the CD45R phenotype and lymphokine mRNA profile was evaluated for encephalitogenic CD4+ T cell lines in vitro and compared to CD4+ T cells isolated from the spinal cord of Lewis rats with EAE. All of the myelin basic protein (MBP)-specific T cell lines and clones that adoptively transferred EAE were > 90% CD4+ and > 90% CD45R lo. A time course of EAE disease progression was monitored as a function of the percentage of CD45R hi/CD4+ T cells isolated from the spinal cords of diseased animals. The majority of CD4+ T cells found in the central nervous system during the early phase of passive EAE were CD45R lo (the same as the encephalitogenic lines/clones). A large increase of the CD45R hi/CD4+ T cells (up to 45%) was observed during the peak and recovery phases of EAE. Lymphokine mRNA production was analyzed from antigen-stimulated MBP-specific lines, and from spinal cord lymphocytes isolated from rats with EAE. The BP-specific lines produced Th1 lymphokines (IL-2, IFN-gamma, and TNF-alpha), while the spinal cord lymphocytes produced the same Th1 lymphokines as well as IL-4 and IL-10. The CD45R hi/CD4+ T cells isolated from the spinal cords were larger and expressed more lymphokine RNA per cell than the CD45R lo/CD4+ T cells. The encephalitogenic cells (CD45R lo) were detected in the spinal cords of rats with a fluorescent dye and by allelic transfers and all of the CD45R hi/CD4+ T cells were found to be host recruited. Thus, it appears that the CD45R hi/CD4+ lymphocytes found in the spinal cord represent a host-recruited, activated cellular infiltrate that increased in number in the recovery phase of EAE and synthesized both Th1 and Th2 lymphokines.

The effect of TCR Vβ8 peptide protection and therapy on T cell populations isolated from the spinal cords of Lewis rats with experimental autoimmune encephalomyelitis

Vaccination or treatment of Lewis rats with TCR V beta 8 peptides can prevent or reverse the clinical signs of experimental autoimmune encephalomyelitis (EAE) which is mediated predominantly by V beta 8.2+ CD4+/CD45R lo T cells. However, rats protected or treated with V beta 8 peptides still developed histological lesions in the spinal cord (SC), even though they remained clinically well. We sought to discern phenotypic changes characteristic of these SC infiltrating lymphocytes. In particular, we focused on whether the immunoregulatory mechanism induced by TCR peptides caused a reduction of V beta 8.2+ T cells, or induced changes in CD45R lo or hi/CD4+ subpopulations that have been associated respectively with EAE induction or recovery. In the V beta 8 peptide vaccinated rats there was a dramatic decrease in the number of V beta 8.2+ T cells isolated from the SC early in disease. During the recovery phase, however, the number of V beta 8.2+ SC T cells was similar in protected and control groups; in contrast, there was striking reduction in the number and size of CD45R hi/CD4+ T cells in the protected animals. In rats treated with V beta 8.2 peptide, no changes were observed in the number of SC V beta 8.2+ T cells or expression of V beta 8.2 message, but similar to vaccinated rats, there was a marked decrease in the number of CD45R hi/CD4+ T cells. These data suggest that vaccination with TCR peptides prevented the initial influx of encephalitogenic V beta 8.2+ T cells into the central nervous system (CNS), whereas treatment appeared to inactivate V beta 8.2+ T cells already present in the CNS. In both cases, TCR peptide-induced inhibition of the encephalitogenic T cells apparently preempted the need for CD45R hi/CD4+ T cells that may normally be necessary to resolve the disease.

Natural immunodominant and experimental autoimmune encephalomyelitis-protective determinants within the lewis rat vβ8.2 sequence include CDR2 and framework 3 idiotopes

The Vβ8.2‐39‐59 peptide has served as a prototypic natural regulatory idiotope in Lewis rats developing experimental autoimmune encephalomyelitis (EAE). The purpose of the present study was to determine if additional regulatory regions were contained within the Vβ8.2 sequence expressed by most encephalogenic T cells. A comprehensive strategy utilizing Vβ8+ hybridomas, a recombinant (r) Vβ8.2 molecule, and overlapping synthetic Vβ8.2 peptides reconfirmed the natural recognition of the 39‐59 idiotope, and revealed a second immunodominant and EAE‐protective determinant residing within residues 71‐90. Both the Vβ8.2‐39‐59 and Vβ8.2‐71‐90 peptides were immunogenic, and each was recognized after immunization of Lewis rats with Vβ8+ cells or rVβ8.2, indicating the preservation of these epitopes during the processing of the Vβ8.2 chain. Moreover, both epitopes were recognized naturally by T cells from rats developing or recovering from EAE that had never been purposefully immunized with Vβ8.2 peptides or rVβ8.2. Of additional interest, the Vβ8.2‐31‐50 peptide was recognized by T cells from some rats immunized with complete Freunds adjuvant (CFA) alone. This peptide possessed mildly protective activity against EAE and thus could account for sporadic reports of CFA interference in EAE.

Protection against experimental encephalomyelitis: Idiotypic autoregulation induced by a non-encephalitogenic T cell clone expressing a cross-reactive T cell receptor V gene

The recovery process in experimental autoimmune encephalomyelitis (EAE) in Lewis rats is characterized by an increasing diversity of T cell clones directed at secondary epitopes of myelin basic protein. Of particular interest, residues 55 to 69 of guinea pig basic protein could induce protection against EAE. A nonencephalitogenic T cell clone, C455-69, that was specific for this epitope transferred protection against both active and passive EAE. Clone C4 was found to express V beta 8.6 in its Ag receptor, and residues 39 to 59 of the TCR V beta 8.6 sequence were found to be highly crossreactive with the corresponding residues 39 to 59 of TCR V beta 8.2, which is known to induce protective anti-idiotypic T cells and antibodies. Like the TCR V beta 8.2 peptide, the V beta 8.6 sequence induced autoregulation and provided effective treatment of established EAE. Thus, the EAE-protective effect of the guinea pig basic protein 55-69 sequence was most likely mediated by T cell clones such as C4 that could efficiently induce anti-TCR immunity directed at a cross-reactive regulatory idiotope.

Subacute sclerosing panencephalitis: influence of the clinical course and treatment with isoprinosine on non‐specific cell‐mediated and humoral immunity

ABSTRACT‐ Cell‐mediated and humoral immunity were studied in 30 patients with subacute sclerosing panencephalitis (SSPE). Marked changes in cell‐mediated immunity were observed, manifested as a decrease in total lymphocyte count, decrease of proportion of T cells forming late E rosettes, depression in lymphocyte blastogenesis, production of migration inhibition factor and delayed‐type skin reactivity. The humoral immune responses was not so markedly changed and only increased levels of serum IgG, IgM and the C3c fragment of complement were noticed. The changes in immunity were more pronounced in patients with a more advanced stage of the disease. 19 patients were investigated 3 times at 7‐week intervals during treatment with isoprinosine. In the group of patients (10 cases) whose clinical status was rapidly deteriorating, a marked decline of cell‐mediated immunity was observed. In the group of patients showing a stationary course of the disease, cellular immunity was improving. The mechanism of the disturbances of non‐specific immunity in SSPE and the influence of isoprinosine on the immune answer are discussed.

Immunoregulatory determinants on rat TCR Vβ8.2.

Antineulrophil cytoplasmic antibodies (ANCA) have been detected in patients with Wegener granulomatosis, various forms of necrotizing vasculitis and other autoimmune diseases such as connective tissue diseases, inflammatory bowel disease and autoimmune liver disease. The humoral immune response seems to play a role in the pathogenesis of MS. The blood brain barrier and particulary the endothelial cell may be a potential target for this immune response. The possible expression of proteinase-3 (target for C-ANCA) and of myeloperoxidase (target for P-ANCA) at the surface of activated endothelial cells might allow these antibodies to injure the endothelium. It is from this standpoint that we have investigated the prevalence of ANCA in the sera of 50 consecutive patients with clinically definite MS (28 women, 22 men; mean age 35.1 years; mean time from disease’s onset 6 years; 27 relapsing-remitting and 23 chronic progressive). ANCA were investigateg by indirect immunefluorescence test and confirm by ELBA. Three out of 50 (6%) had a positive titre of ANCA although at low levels (one P-ANCA, two C-ANCA). No correlation between ANCA and any clinical parameter was found. ANCA do not seem to play a role in the pathogenesis of MS.