Modulation of experimental allergic neuritis in rats by in vivo treatment with monoclonal anti T cell antibodies

Monoclonal antibodies (MCA) to different T lymphocyte cell surface antigens have been used to treat rats during different phases of the development of experimental allergic neuritis (EAN). The effects of this treatment were followed by clinical evaluation and in some instances by immunohistochemical analysis of lymphoid organs and affected nerves of the antibody-treated rats. Several MCA, W3/13 (pan T cell reactive), W3/25 (anti-rat CD4), Ox 8 (anti-rat CD8) as well as Ox 6 (anti-Ia) partly prevented clinical signs of EAN when given shortly before expected onset of disease, whereas W3/13 and Ox 8 given at the height of disease did not further affect disease development. However, Ox 19 (anti-rat CD5) given at the same time as immunization partly prevented clinical signs of EAN, while Ox 19 given shortly before expected onset of disease or during height of disease drastically exaggerated disease symptoms. Immunohistochemical studies after Ox 8 or Ox 19 treatment showed a complete absence of staining for the respective antibodies, while staining was preserved with the other MCA. It is concluded that: (1) Ox 8 positive "suppressor/cytotoxic" T lymphocytes do not exert any suppressive effects on EAN during the now investigated phases of disease, and that (2) anti T lymphocyte antibodies (here Ox 19) may exert opposite effects on autoimmune disease when given at different phases of disease development. This may have implications for potential therapeutic trials of MCA therapy for putative autoimmune demyelinating diseases in man.     

Restraint stress suppresses experimental allergic encephalomyelitis in Lewis rats

We studied the effect of restraint stress on experimental allergic encephalomyelitis (EAE), a model for the human disease multiple sclerosis (MS), in Lewis rats. Rats were subjected to 12 h restraint stress for 3 consecutive nights from the first (day 1) or the eighth day (day 8) after sensitization with the antigen (guinea-pig spinal cord). All controlled rats exhibited clinical and histologie signs of EAE. The mean ± SD of incubation time, clinical score (0–4) and histologic score (0–3) for this group were 12.8 ± 1.0 days, 2.7 ± 0.6, and 2.3 ± 0.7, respectively. Restraint stress from day 8 significantly suppressed EAE: the mean ± SD of incubation time, clinical score, and histologic score for this group were 17.2 ± 2.2 days (p < 0.001), 1.8 ± 1.3 (p < 0.05), and 1.5 ± 0.9 (p < 0.02), respectively. Restraint stress from day 1 did not modify EAE. The findings suggest that stressful factors may exert an influence on the clinical course of MS.     

Reinduction of experimental allergic encephalomyelitis in convalescent Lewis rats with cyclophosphamide

Experimental allergic encephalomyelitis (EAE) in Lewis rats is an acute, monophasic autoimmune disease, and rats recovering from EAE are generally resistant to active reinduction. However, following immunosuppression with a high dose (100-200 mg/kg) of cyclophosphamide (CY), EAE was reinduced in convalescent rats, irrespective of whether or not they were rechallenged with the encephalitogenic inoculum. Reinduction of EAE was not observed in rats treated with low doses (20 mg/kg or less) of CY. A selective inhibition of suppressor cells during the convalescent stage may be responsible for the reinduction of EAE in Lewis rats.     

T cell immunity and interferon-gamma secretion during experimental allergic encephalomyelitis in Lewis rats.

An immunospot assay that detects single secretory cells was used to enumerate interferon-gamma secreting cells (IFN-gamma-sc) in mononuclear cell suspensions from the central nervous system (CNS) and peripheral lymphoid organs after actively induced experimental allergic encephalomyelitis (EAE) in Lewis rats. In the CNS compartment there was a significant increase in the number of IFN-gamma-sc preceding the onset of the clinical signs of EAE. Both in rats with EAE and rats immunized with Freund's complete adjuvant (FCA) the number of IFN-gamma-sc increased in peripheral lymphoid organs, as compared to non-immunized controls. In view of the potent immunoregulatory effects of IFN-gamma, its intra-CNS secretion may play a crucial role for clinicopathological events in EAE. To study the numbers of primed T cells that in response to myelin antigens produced IFN-gamma, mononuclear cell suspensions from peripheral lymphoid organs were precultured to allow for antigen uptake, presentation and T cell triggering, followed by enumeration of IFN-gamma-sc. T cells responding to a peptide of myelin basic protein (MBP) that previously have been shown encephalitogenic in Lewis rats, appeared initially and were quantitatively dominant over the course of EAE. Later, T cell reactivities to multiple regions of MBP appeared, showing that the concept of immunodominance in EAE is non-absolute and time dependent. Splenocyte cultures from EAE rats exposed to the different antigens showed a reduced number of IFN-gamma-sc compared to cultures not exposed to antigen, suggesting an antigen-induced suppression of T cell effector molecules.     

The beta-adrenergic agonist isoproterenol suppresses experimental allergic encephalomyelitis in Lewis rats

Treatment with the beta-adrenergic agonist isoproterenol suppresses clinical and histological experimental allergic encephalomyelitis in Lewis rats. The effect of isoproterenol treatment is greater when the drug is given from the time of immunization through the acute phase of the illness or from 8 to 14 days post-immunization than when given for the first 7 days after immunization.     

Pregnancy and the susceptibility of Lewis rats to experimental allergic encephalomyelitis

Pregnant Lewis rats challenged with encephalitogen during the second or third week of gestation were afforded a high level of protection against experimental allergic encephalomyelitis, whilst females sensitised during the first week of pregnancy enjoyed only limited protection from clinical signs of disease. When rechallenged with encephalitogen, females which had been sensitised during pregnancy were marginally more susceptible to reinduction of clinical signs of disease than their virgin counterparts. Mothers inoculated during the first week of gestation were the only group to produce abnormal young.     

Suppression of acute experimental allergic encephalomyelitis in Lewis rats with cerebral contusion

We previously reported that the thymolysis was observed in acute experimental allergic encephalomyelitis (EAE) rats immunized with brain tissue homogenates in complete Freund adjuvant. This observation has led us to suppose that the central nervous system (CNS) has high antigenicity for self. To keep from the high antigenicity of CNS, the blood brain barrier (BBB) and cerebrospinal fluid (CSF) which maintain CNS in isolated state are essential for self. If the BBB is destroyed and brain tissue is disseminated in CSF, is oneself sensitized by CNS? In this experiment, we produced cerebral contusion in rats and the BBB was partially destroyed and brain tissue was disseminated in CSF. No clinical signs of EAE were observed by cerebral contusion was exposed to encephalitogen one week later, EAE was remarkably suppressed compared with those in controls or sham operated groups (p less than 0.001). Almost 80% of the rats with cerebral contusion showed no response to EAE. These results seen to show that self is sensitized by autologous brain tissue and CNS has high antigenicity.     

Reactive gliosis in the brains of Lewis rats with experimental allergic encephalomyelitis

This paper assesses reactive gliosis in the optic tracts and other regions of brain in Lewis rats with experimental autoimmune encephalomyelitis (EAE). Enhanced immunostaining for glial fibrillary acidic protein (GFAP) in brains from rats with EAE occurred primarily in the white-matter tracts and was not restricted to sites of inflammation. Immunocytochemical staining for other putative astrocytic antigens demonstrated glutathione-S-transferase (Yb isoenzyme) to be localized extensively in GFAP-positive cells and vimentin to be present both in inflammatory cells and in some GFAP-positive astroglial cells. Positive staining for carbonic anhydrase and glutamine synthetase was observed in oligodendrocytes. In the optic tracts glutamine synthetase, but not carbonic anhydrase, was also observed in some astrocytes.     

T cell vaccination in monoclonal antibody-induced hyperacute experimental allergic encephalomyelitis

Demyelinating inflammatory disease of the central nervous system-(CNS) can be a multifactorial process mediated by cellular and antibody-mediated immune processes. In rats, hyperacute disease progression and severe demyelination can be induced in experimental allergic encephalomyelitis (EAE)-diseased animals by injection of a monoclonal antibody, 8-18C5, specific for an oligodendrocyte cell surface glycoprotein. Here we demonstrate that this antibody-induced hyperacute EAE can be prevented by 'vaccination' with myelin basic protein (MBP)-specific T cells. Thus, the 8-18C5 antibody-mediated disease process is critically dependent on inflammatory processes induced by T lymphocytes and T cell vaccination is highly effective in preventing the development of demyelinating CNS lesions.     

Treatment of clinical experimental allergic encephalomyelitis in the rat with monoclonal antibodies

An anti-CD4 monoclonal antibody of the IgG2a subclass, OX35, and an anti-I-A monoclonal antibody (IgG1) were used in vivo to treat experimental allergic encephalomyelitis in the Lewis rat. The anti-CD4 antibody was as effective in shortening the duration of the disease as the previously reported use of W3/25 (Brostoff and Mason, J. Neuroimmunol., 10 (1984) 331–340). It did not appear necessary for the antibody treatment to remove the CD4⁺ cells from the circulation in order to show an effect on the clinical course of the disease. The anti-I-A antibody did not have any noticeable effect in this disease model.     

Mast cells in brains during experimental allergic encephalomyelitis in Lewis rats.

We studied the number of mast cells and their extent of degranulation in brains of Lewis rats with acute experimental allergic encephalomyelitis (EAE), activity induced with guinea pig spinal cord and Freund's complete adjuvant. Non-immunized controls and EAE rats were killed on days 10, 11, 12, and 16 post-immunization (p.i.). The percentage of degranulated mast cells was significantly increased in EAE brains. Signs of degranulation were observed as early as day 10 p.i. Clinical EAE signs appeared from day 10 p.i. A significant change in mast cell number was not observed. The percentage of degranulated cells was largest at day 16 p.i., at a time when the inflammation had reached the thalamus. This indicates that mast cell degranulation may occur as a result of the inflammation. Collectively, the data suggest that mast cells may play a role in the pathogenesis of EAE.     

Neurovascular fibrinolytic activity in normal Lewis rats and rats with cell-transferred experimental allergic encephalomyelitis

Fibrinolytic activity in the form of plasminogen activator (PA) was assessed using a histochemical fibrin slide technique in spinal cords of normal Lewis rats and rats with the cell-transferred form of experimental allergic encephalomyelitis (EAE). PA was localized exclusively to blood vessels. Vessels in the leptomeninges had maximum activity. A precipitous decrease in PA activity occurred in recipient rats which coincided with onset of clinical neurologic signs. A subsequent return in activity occurred in association with clinical remission of disease but remained well below the activity level of normal rats for as long as the recipient animals were followed. Vessels containing perivascular cellular infiltrates of EAE had little or no detectable PA activity. Furthermore, PA could not be demonstrated to be associated with infiltrating inflammatory cells, including macrophages. These findings provide further support for involvement of the coagulation and fibrinolytic systems in the early clinical manifestations of EAE in Lewis rats.     

Experimental allergic encephalomyelitis induced by proteolipid apoprotein in Lewis rats

Experimental allergic encephalomyelitis (EAE) was induced in inbred Lewis rats by sensitization with bovine white matter proteolipid apoprotein (PLP). 18-61 days after a single injection of 100 micrograms of PLP, 12 of 31 rats (39%) developed clinical EAE and 18 of 23 (78%) showed pathologic EAE with significant demyelination. Lymphocyte proliferative responses and antibodies to PLP were elevated but did not correlate with the clinical or pathologic state. This is the first demonstration of PLP-induced EAE with significant demyelination in rats and will contribute to the study of autoimmune demyelination.     

Inhibition of experimental allergic encephalomyelitis in the Lewis rat by paclitaxel

Experimental allergic encephalomyelitis (EAE), an animal model for multiple sclerosis (MS), is useful for preclinical testing for agents to be considered for treatment for this human demyelinating disease. Microtubules in lymphocytes play an important role in the cascade of human T cell activation, and paclitaxel (PTX), a microtubule stabilizer, can inhibit T cell function. A new formulation of micellar PTX, free of Cremophor and ethanol, was tested for its effect on the induction of EAE in Lewis rats. Adoptive EAE was induced with an encephalitogenic T cell line activated with guinea pig myelin basic protein (GP MBP) peptide 68-88. PTX (10 mg/kg) was administered 24 and 72 h after cell transfer. The clinical signs, fulminating in controls, were completely blocked by PTX, but mild CNS inflammation remained unaltered. A similar dose of PTX, given on days 6 and 8 to animals developing active EAE after immunization with GP MBP peptide 68-88 in complete Freund's adjuvant, greatly reduced the severity of paralysis and delayed the onset of disease by 8-9 days. Marked weight loss and severe toxicity were noted with higher and more prolonged administration. In vitro micellar PTX inhibited activation of encephalitogenic T cells by both specific antigen and mitogen. Lower doses and longer treatment programs may provide effective treatment with acceptable adverse effects with this agent in the treatment of inflammatory demyelinating disease.     

Lovastatin treatment decreases mononuclear cell infiltration into the CNS of Lewis rats with experimental allergic encephalomyelitis

Mononuclear cell infiltration into the CNS and induction of inflammatory cytokines and iNOS in diseases like multiple sclerosis (MS) and experimental allergic encephalomyelitis (EAE) have been implicated in subsequent disease pathogenesis and progression. We report that Lovastatin treatment blocks the clinical disease and induction of inflammatory cytokines and iNOS in spinal cords of MBP induced EAE rats. A significant number of the infiltrating cells in CNS were ED1+ cells of monocyte/macrophage lineage. To understand the mechanism of efficacy of Lovastatin against EAE, we examined the effect of Lovastatin on the transmigration of mononuclear cells into EAE spinal cord. The data presented here documents that Lovastatin treatment attenuates the transmigration of mononuclear cells possibly by down regulating the expression of LFA-1, a ligand for ICAM, in endothelial-leukocyte interaction. These results indicate that Lovastatin treatment prevents infiltration by mononuclear cells into the CNS of rats induced for EAE, thereby lessening the histological changes and clinical signs and thus ameliorating the disease. These observations indicate that Lovastatin treatment may be of therapeutic value against inflammatory disease process associated with infiltration of activated mononuclear cells into the tissue.     

T cell immunity and interferon-γ secretion during experimental allergic encephalomyelitis in Lewis rats

An immunospot assay that detects single secretory cells was used to enumerate interferon-γ secreting cells (IFN-γ-sc) in mononuclear cell suspensions from the central nervous system (CNS) and peripheral lymphoid organs after actively induced experimental allergic encephalomyelitis (EAE) in Lewis rats. In the CNS compartment there was a significant increase in the number of IFN-γ-sc preceding the onset of the clinical signs of EAE. Both in rats with EAE and rats immunized with Freund's complete adjuvant (FCA) the number of IFN-γ-sc increased in peripheral lymphoid organs, as compared to non-immunized controls. In view of the potent immunoregulatory effects of IFN-γ, its intra-CNS secretion may play a crucial role for clinicophatological events in EAE.To study the numbers of primed T cells that in response to myelin antigens produced IFN-γ, mononuclear cell suspensions from peripheral lymphoid organs were precultured to allow for antigen uptake, presentation and T cell triggering, followed by enumeration of IFN-γ-sc. T cells responding to a peptide of myelin basic protein (MBP) that previously have been shown encephalitogenic in Lewis rats, appeared initially and were quantitatively dominant over the course of EAE. Later, T cell reactivities to multiple regions of MBP appeared, showing that the concept of immunodominance in EAE is non-absolute and time dependent.Splenocyte cultures from EAE rats exposed to the different antigens showed a reduced number of IFN-γ-sc compared to cultures not exposed to antigen, suggesting an antigwn-induced suppression of T cell effector molecules.     

Chronic relapsing experimental allergic encephalomyelitis in the Lewis rat

Chronic relapsing experimental allergic encephalomyelitis (CR-EAE) was induced in rats with an emulsion of guinea-pig spinal cord tissue (GPSC) in complete Freund's adjuvant (CFA) enriched with Mycobacterium tuberculosis H37RA (Tbc). 78% of the sensitized rats developed a CR-EAE showing 2 to 3 clinical relapses during the first 40 days. After 60-80 days, approximately half of the rats with CR-EAE had a further relapse which was followed by complete recovery in only 35% of the cases. The remaining 65% of these animals showed a progressive state of the disease, characterized by paralysis or severe motor deficit, eventually leading to death. CR-EAE in rats showed some similarities to multiple sclerosis in man (MS) and it may be a useful model for the study of this disease.     

实验性自身免疫性脑脊髓炎大鼠中1,25二羟基维生素D3的免疫调节作用

目的探讨1,25二羟基维生素D3[1,25(OH)2D3]对实验性自身免疫性脑脊髓炎(EAE)的免疫调节作用。方法建立Lewis大鼠主动免疫EAE实验动物模型,分别于致敏当天(预防组)及EAE症状出现当天给药(治疗组),并设立相应对照组。动态观察各组大鼠的临床评分,于致敏第13天处死,测定其引流淋巴结中细胞总数,单个核细胞(MNC)中CD 4CD 25T细胞含量、CD 86干细胞含量及MNC培养上清中干扰素(IFN)γ和白细胞介素(IL)4含量。结果预防组1,25(OH)2D3使EAE发病高峰延迟,治疗组显示1,25(OH)2D3能减轻EAE的病情,使高峰期评分[(3.3±0.6)分]比对照组[(4.0±0.3)分]降低(P<0.05);1,25(OH)2D3干预后EAE大鼠高峰期发现淋巴结中CD 4CD 25T细胞、引流淋巴结的细胞总数与对照组差异无统计学意义;而预防组CD 4CD 25/CD 4T细胞比值(15.1±3.3)高于其对照组(12.3±2.6,P<0.05);预防组、治疗组CD 86细胞明显降低(P<0.05)。1,25(OH)2D3干预后预防组及治疗组中淋巴结MNC培养上清中IFNγ无明显改变,而IL4显著增多(P<0.01)。结论应用1,25(OH)2D3治疗EAE大鼠,可通过改变共刺激分子表达、调节性T细胞比例以及不同细胞因子的分泌能力使EAE症状缓解。     

Reactive oxygen species are involved in the pathogenesis of experimental allergic encephalomyelitis in Lewis rats

During experimental allergic encephalomyelitis (EAE), both blood-borne macrophages as well as activated, resident microglial cells are considered to be involved in inflammatory reactions in the central nervous system (CNS), resulting in the neurological deficits common to EAE. Both cell types can produce multiple mediators of tissue damage, among which are the reactive oxygen species (ROS). In this study we show that macrophages and microglial cells, isolated from the CNS of Lewis rats with clinical signs of EAE, exhibited significantly elevated spontaneous and phorbol myristate acetate (PMA)-inducible levels of ROS compired to similar cells isolated from healthy controls, sham (complete Freund's adjuvant, CFA)-immunized rats as well as rats sacrificed before the manifestation of clinical signs of EAE. However, during clinical EAE, peripheral blood mononuclear cells (PBMC) did not show increased spontaneous nor PMA-inducible ROS production compared to controls. In vivo treatment of EAE with catalase, which scavenges the ROS H₂O₂, markedly suppressed the severity of the disease as compared to sham (albumin)-treated controls. In contrast, Superoxide dismutase had no effect on clinical signs. Our studies point at a putative functional role for ROS, and in particular H₂O₂, in the pathogenesis of EAE.     

Down-regulation of experimental allergic encephalomyelitis in DA rats by tiazofurin

The immunomodulatory potential of tiazofurin (TR) on experimental autoimmune encephalomyelitis (EAE) was investigated. Given continuously, TR dose-dependently suppressed the development of EAE in Dark Agouti (DA) rats immunized with either rat spinal cord homogenate (SCH) or myelin oligodendrocyte glycoprotein (MOG). Amelioration of clinical signs was also obtained when the drug was administered during the inductive phase only (day 0 to 8), or during the effector phase (day 10 to 20) of the disease. Efficacy of TR was further evaluated by adoptive transfer of the disease with myelin basic protein (MBP)-sensitized draining lymph node cells (DLNC). Cells from TR-protected rats failed to transfer the disease into naive syngeneic recipients; in addition, TR treatment of recipient rats that had received MBP-sensitized lymphoid cells diminished the adoptively transferred EAE. A reduction of clinical EAE in TR-treated rats was accompanied with the absence of mononuclear infiltration in the spinal cord and defective adhesive cell-cell interactions. The anti-MOG autoAb production was also decreased. Importantly, no evidence for a generalized impairment of the T cell activity, nor decreased in vitro proliferative antigen specific response of LNC from TR-treated animals was found. These results suggest that TR exerts its EAE protective and suppressive effects by limiting adhesive interactions involved in the autoimmune pathogenic process, and due to the lack of general immunosuppressive activity, it should be considered as a candidate drug for the treatment of neuroinflammatory diseases like multiple sclerosis (MS).