Clinical Experimental Allergic Encephalomyelitis Research Articles

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), actively 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.

Induction of Experimental Allergic Encephalomyelitis by Myelin Proteolipid-Protein-Specific T Cell Clones and Synthetic Peptides

Proteolipid protein (PLP) is the major protein of central nervous system (CNS) myelin. SJL(H-2s) mice immunized with a synthetic peptide corresponding to PLP residues 139-151 (HSLGKWLGHPDKF) develop acute experimental allergic encephalomyelitis (EAE). In the present study a T cell line and 4 clones were derived from SJL/J mice following immunization with this synthetic peptide. Severe clinical and histological EAE could be induced by adoptive transfer of the peptide-specific T cell line and 3 of 4 T cell clones. The T cell line/clones all responded strongly to PLP peptide 139-151 in in vitro proliferative assays. However, two different reactivity patterns emerged when truncated PLP peptides 141-150 and 141-149 were tested, suggesting that more than 1 epitope may be present within the PLP 139-151 determinant. To evaluate the encephalitogenic potential of the truncated peptides, we compared the ability of 2 truncated PLP peptides to induce EAE in vivo and proliferative responses in vitro. Immunization with PLP peptide 141-150 induced acute EAE in about 70% of mice tested, but PLP peptide 141-149 induced a comparatively mild form of EAE in 4 out of 9 mice tested. Lymph node cells from mice immunized with these peptides showed in vitro proliferative responses to each of the peptides, but the response to peptide 139-151 was always strongest. These combined in vivo and in vitro data further define the epitopes involved in PLP-induced EAE in SJL mice. Furthermore, the availability of multiple PLP-specific T cell clones will enable us to study the diversity of the T cell repertoire to PLP.

Adoptive transfer of experimental allergic encephalomyelitis and localization of the encephalitogenic epitope in the SWR mouse

Adoptively-transferred experimental allergic encephalomyelitis (EAE) was induced in the SWR (H-2 q) strain of mouse using lymph node cells, spleen cells, or cell lines sensitized to whole myelin basic protein (MBP). With the aid of synthetic peptides, the minimal encephalitogenic epitope of MBP for the SWR strain was localized to amino acids 87–99 of the MBP molecule. The 87–99 sequence is also encephalitogenic for the SJL strain of mouse and the Lewis rat. EAE was induced with a protocol similar to that for the induction of EAE in the SJL strain with the exception that sublethal irradiation of recipients was necessary. Mice developed typical clinical and pathological EAE 6–14 days post-transfer of cells sensitized to either whole MBP or peptide 87–99, after which they remitted. No relapses were observed. Thus, adoptively transferred EAE can be induced in irradiated H-2 q mice for which the encephalitogenic epitope is one of the nested encephalitogenic epitopes for SJL (H-2 s) mice, namely residues 87–99.

Changes of experimental allergic encephalomyelitis by methionine-enkephalin injected into lateral ventricles of the rat brain

Our previous investigations have shown that the opioid peptide methionine-enkephalin (Met-Enk) modulates in vivo a variety of humoral and cell-mediated immune performances. In this study, rats bearing polyethylene cannulae permanently inserted into the lateral ventricles of the brain were used. Experimental allergic encephalomyelitis (EAE) was induced with guinea pig spinal cord in complete Freund's adjuvant injected into the left hind foot pad. The following groups of cannulated rats were tested: nontreated with Met-Enk or saline, intracerebroventricularly (i.c.v.) injected with saline, and i.c.v. treated with low (1 microgram/kg) dose of Met-Enk and high (1 mg/kg) dose of Met-Enk. Intact noncannulated rats sensitized for EAE served as an additional control. The results showed that i.c.v. treatment with 1 microgram/kg of Met-Enk significantly increased the incidence and severity of EAE. In contrast, injections of 1 mg/kg of Met-Enk produced a moderate decline of clinical EAE, but marked diminution of inflammatory lesions in the brain. Interestingly, histopathology of EAE was more pronounced in control rats treated i.c.v. with saline. On the other hand, control cannulated rats noninjected with saline exhibited a striking decrease of neurological and histopathological signs of the disease, thus indicating a suppressive effect of stress (surgical procedure) on EAE. In conclusion, the present study showing the central effect of Met-Enk on EAE when peptide was applied in the cerebral cavity, and earlier studies which revealed the peripheral effect on EAE when Met-Enk was administered intraperitoneally, suggests that Met-Enk exerts its immunomodulatory action both centrally and peripherally.

Adoptive transfer of experimental allergic encephalomyelitis: Conditions influencing memory and effector cell development

The cellular transfer of clinical experimental allergic encephalomyelitis (EAE) with immune spleen cells is only accomplished following lymphoid cell stimulation during an intervening in vitro culture activation period. Recipients of these cells recover from the ensuing adoptively transferred paralytic episode and subsequently respond to active challenge with myelin basic protein (BP)-CFA in an accelerated time frame consistent with the presence of memory cells in the initial cell transfer inoculum. We have found that the addition of anti-CD4 antibody or dexamethasone during the activation period inhibits the development of the transfer active EAE effector cell subpopulation, but does not alter the in vitro development and subsequent expression of the BP-specific memory cell subpopulation. Additional experiments also suggest the development of memory cells in the absence of effector cell activity. PMA + ionomycin when used as a stimulus during the culture activation period leads to effector and memory cell development. The stimulation response is dose dependent, in that a reduced concentration of PMA + ionomycin does not lead to EAE effector cell development; however, at these reduced levels of PMA + ionomycin, memory cell development still occurred. Additional evidence which supports the concept of independent development of memory cells and effector cells was obtained with a BP-specific cell line. Following recovery from cell line-mediated clinical EAE, as well as following adoptive transfer of the cell lines in the precursor stage, cell recipients did not develop an early onset of active EAE when subsequently immunized with BP-CFA. Thus the BP-specific T-cell line appears to contain the precursors of the effector cell subpopulation but does not appear to contain the BP memory cell subpopulation. Collectively these observations suggest the existence of distinct T-cell subsets or pathways of development that are followed during the response to BP as measured by the development of Clinical EAE.

Acute Experimental Allergic Encephalomyelitis in SJL/J Mice Induced by a Synthetic Peptide of Myelin Proteolipid Proteina

Clinical, histologic, and ultrastructural characteristics of acute experimental allergic encephalomyelitis (EAE) induced by sensitization with a synthetic peptide corresponding to mouse myelin proteolipid protein (PLP) residues 139-151 HCLGKWLGHPDKF were studied in SJL/J mice. Groups of mice were immunized with 20, 50, or 100 nmol of the peptide and were killed from seven to 28 days after sensitization or when they were moribund. Beginning on Day 9, the mice showed signs of EAE and the disease progressed rapidly to paralysis. Central nervous system (CNS) inflammation, edema, gliosis, and demyelination were found in all mice killed between Days 10 and 28 and white matter lesion areas correlated with clinical score at the time the mice were killed. Peripheral nerve roots and the cauda equina did not have lesions. Within the range studied, the severity of clinical or histologic disease was the same regardless of the PLP peptide dose. Two of ten mice immunized with 100 nmol and none of 14 mice given smaller doses of a synthetic peptide of mouse myelin basic protein (MBP) showed clinical EAE. These mice had small numbers of CNS lesions that were indistinguishable from those in PLP peptide-sensitized mice. These findings demonstrate that immunization of SJL/J mice with PLP peptide 139-151 produces a disease with the clinical and morphologic features of CNS tissue-, whole PLP-, whole MBP-, and MBP peptide-induced acute EAE. Thus, PLP is a major encephalitogen and immune reactions to epitopes of different myelin proteins may induce identical patterns of injury in the CNS.

Acute Experimental Allergic Encephalomyelitis in SJL/J Mice Induced by a Synthetic Peptide of Myelin Proteolipid Protein

Clinical, histologic, and ultrastructural characteristics of acute experimental allergic encephalomyelitis (EAE) induced by sensitization with a synthetic peptide corresponding to mouse myelin proteolipid protein (PLP) residues 139-151 HCLGKWLGHPDKF were studied in SJL/J mice. Groups of mice were immunized with 20, 50, or 100 nmol of the peptide and were killed from seven to 28 days after sensitization or when they were moribund. Beginning on Day 9, the mice showed signs of EAE and the disease progressed rapidly to paralysis. Central nervous system (CNS) inflammation, edema, gliosis, and demyelination were found in all mice killed between Days 10 and 28 and white matter lesion areas correlated with clinical score at the time the mice were killed. Peripheral nerve roots and the cauda equina did not have lesions. Within the range studied, the severity of clinical or histologic disease was the same regardless of the PLP peptide dose. Two of ten mice immunized with 100 nmol and none of 14 mice given smaller doses of a synthetic peptide of mouse myelin basic protein (MBP) showed clinical EAE. These mice had small numbers of CNS lesions that were indistinguishable from those in PLP peptide-sensitized mice. These findings demonstrate that immunization of SJL/J mice with PLP peptide 139-151 produces a disease with the clinical and morphologic features of CNS tissue-, whole PLP-, whole MBP-, and MBP peptide-induced acute EAE. Thus, PLP is a major encephalitogen and immune reactions to epitopes of different myelin proteins may induce identical patterns of injury in the CNS.

Genetic control of resistance to clinical EAE accompanied by histological symptoms

The susceptibility of rats to experimental allergic encephalomyelitis (EAE) induced by myelin basic protein (MBP) was studied in a variety of genetic crosses. Rats were evaluated according to weight loss, neurological symptoms, and histological criteria. The results demonstrate that three different types of genes are involved in susceptibility. An RT1-linked gene is necessary but not sufficient for full expression of EAE induced by MBP in complete Freund's adjuvant (CFA). Additional genes are required for the occurrence of histological EAE, but a full-blown inflammatory reaction is not sufficient for the expression of clinical EAE. A third type of gene, which can be demonstrated in appropriate crosses, is required for the consistent expression of clinical symptoms. Dominant genes for resistance to clinical symptoms were transferred to the Lewis (LEW) background from the BN.B1 strain through two generations of backcrossing. Thus, there are genetically controlled mechanisms involved in the neurological expression of EAE which are independent of the inflammatory reaction as observed in central nervous system (CNS) histology.

The adoptive transfer of chronic relapsing experimental allergic encephalomyelitis with lymph node cells sensitized to myelin proteolipid protein

In this report we describe the transfer of chronic relapsing experimental allergic encephalomyelitis (EAE) with in vitro-stimulated lymph node cells (LNC) from SJL/J mice immunized with human myelin proteolipid protein (PLP). No additional immune enhancing procedures were applied in the transfer recipients. Clinical and histological EAE was transferred with 10–30 × 10 6 LNC to 27/28 mice. The LNC proliferated in vitro to PLP, but not to myelin basic protein (MBP), and induced delayed-type hypersensitivity. Enrichment for lymphoblasts by Ficoll centrifugation was essential for the disease development. The clinical course usually showed an early episode of acute paralytic illness, followed by chronic relapsing disease, and resembled the transfer of EAE using MBP-specific cells, both clinically and histologically.

Successful treatment of EAE in rhesus monkeys with MHC class II specific monoclonal antibodies

Experimental Allergic Encephalomyelitis (EAE) was induced in rhesus monkeys by subcutaneous immunization with calfbrain homogenate in complete Freunds adjuvant. Monkeys were treated with major histocompatibility complex (MHC) Class II specific monoclonal antibodies (MoAb) as soon as the first clinical EAE signs became apparent. Two different treatments were tested. One consisted of 10 daily injections of a mixture of two MHC Class II specific MoAb, reactive with a monomorphic structure of rhesus monkey Class II molecules. The other consisted of 10 daily injections of Genox3.53, specific for HLA-DQW1. This MoAb crossreacts well with monkeys and also detects a polymorphism in this species and is presumably reactive with the RhLA-DQW1 antigen. Both MoAb treatments could modify the clinical course of EAE favourably. Untreated animals invariably died within 3 d of the onset of clinical EAE signs. Only one of the three monkeys treated with the monomorphic MHC Class II MoAb preparation died within 3 d, and the other two survived significantly longer than untreated animals. Both animals treated with Genox3.53 survived significantly longer than untreated control animals. Although only a few animals were tested, these results clearly show the possible beneficial influence of MHC Class II specific MoAb on a T-cell mediated autoimmune disease.

Replication of donor lymphocytes in recipients is not essential for the passive transfer of allergic encephalomyelitis

Gliotoxin is a fungal metabolite belonging to the class of epipolythiodioxopiperazines which possesses both immunomodulating and anti-phagocytic activities. We have examined the effect of gliotoxin on passively induced allergic encephalomyelitis and report here that pulse treating activated experimental allergic encephalomyelitis (EAE) effector lymphocytes with gliotoxin inhibits, in a dose-dependent manner, their ability to transfer disease. Cells treated with 300 ng/ml are unable to replicate in vitro in response to concanavalin A stimulation, nor did they produce interleukin-2 (IL-2) following stimulation. Furthermore this concentration of gliotoxin also causes complete fragmentation of genomic DNA in treated cells, yet these cells are still capable of transferring clinical EAE. These data suggest that replication of donor lymphocytes in the recipient is not essential for the development of EAE.

Effect of viral infection on experimental allergic encephalomyelitis in mice

BALB/c mice were irradiated with 350 R and injected with mouse spinal cord homogenate (MSCH) in complete Freund's adjuvant. Only 15–30% of these animals developed signs of experimental allergic encephalomyelitis (EAE) at 21–28 days after inoculation. Intraperitoneal infection with the non-lethal A7 strain of Semliki forest virus (SFV) 7 days after sensitization reduced the mean appearance time of the EAE symptoms to 14 days and the number of animals with clinical EAE increased up to 70%. In contrast, virus inoculation 10 days before induction of EAE decreased significantly the incidence of clinical EAE in both BALB/c and SJL mice. Demyelination with increased cellularity, presence of macrophages, stripping of myelin from the axons and sparing of oligodendrocytes was observed in spinal cords of animals at days 13–16 after induction of EAE and subsequent virus infection. No demyelination was seen in specimens taken at the same time from mice inoculated with MSCH or SFV alone. Combined MSCH and virus inoculations induced changes in the general immune response which may be one of the major reasons for the increase or decrease in demyelination in this model.

Inhibition of allergic encephalomyelitis by the iron chelating agent desferrioxamine: differential effect depending on type of sensitizing encephalitogen

Induction of experimental allergic encephalomyelitis (EAE) in Lewis rats by injection of guinea pig (GP) spinal cord homogenate (SCH) plus adjuvant (SCH-CFA) can be inhibited by treatment with the iron chelating agent desferrioxamine (DFOM). Interestingly, induction of EAE with purified myelin basic protein (BP-CFA) is not inhibited with DFOM. This dichotomy does not appear to be due to any quantitative differences in the two inocula since minimal clinical EAE produced by threshold levels of BP is not inhibited with DFOM. Passive EAE is not inhibited irrespective of the type of encephalitogen used to sensitize the donors. This suggests that the inhibitory effect of DFOM is acting on the afferent limb of the immune response to SCH-CFA. Injection of BP-CFA and SCH-CFA into the same site, mixing BP with central nervous system (CNS) lipids, or incorporating BP into liposomes, all induce EAE which can be partially inhibited by treatment with DFOM. These results support the hypothesis that the close association of lipids with the encephalitogen (i.e. BP) in SCH requires extensive lipid breakdown before adequate antigen presentation can occur, and it is at this level that DFOM exerts its inhibitory effect.

Treatment of clinical experimental allergic encephalomyelitis in the rat using fragments and combinations of monoclonal antibodies

We reported previously that two different anti-CD4 monoclonal antibodies (W3/25, MRC OX35) were effective in treating experimental allergic encephalomyelitis in the Lewis rat whereas anti-I-A antibody was ineffective. Further studies with other monoclonal antibodies and fragments have now been performed. Anti-I-E antibody was ineffective in shortening the disease duration even when used in combination with anti-I-A antibody. Anti-CD2 (T11) antibody was marginally effective, shortening the duration of disease by only one day on the average. Combination of anti-CD4 antibody with anti-CD2 antibody did not improve the recovery time over the use of anti-CD4 antibody alone. On the other hand, the F(ab′) 2 fragment of the anti-CD4 antibody was as effective in the treatment of disease as the intact antibody molecule, indicating that it was sufficient to block the CD4 molecules on the cell surface of the EAE effector cells in order to affect the disease course.

Limiting dilution analysis of the frequency of antigen-reactive lymphocytes isolated from the central nervous system of Lewis rats with experimental allergic encephalomyelitis

Lymphocytes were isolated from the spinal cord and draining lymph nodes of Lewis rats with acute experimental allergic encephalomyelitis (EAE) 12 days after immunization with myelin basic protein (MBP) and tetanus toxoid (TT). An average of 8.0 ± 2.0 × 10 6 cells was obtained from the spinal cord. Of these 71.1 ± 8.6% expressed the helper-T-cell marker W3/25 and 14.8 ± 6.2% expressed the killer/suppressor-T-cell marker O×8. By limiting dilution analysis of cells exhibiting an antigen-specific proliferative response, the average frequencies of cells reactive to MBP and TT were 3.36 ± 2.4 and 7.60 ± 4.1 per 10 4, respectively. In the draining lymph nodes, the frequencies of cells reactive to MBP and TT were 2.24 ± 1.7 and 2.69 ± 2.5 per 10 4. At a relatively early stage of clinical EAE, MBP-reactive T cells comprise only a small minority of the cells which can be isolated from the spinal cord; lymphocytes reactive to a protein antigen irrelevant to EAE pathogenesis are present in comparable numbers. This finding suggests that most of these cells accumulate as a result of mechanisms not specific for MBP-reactive lymphocytes.

The immunopathology of acute experimental allergic encephalomyelitis. IV. An ultrastructural immunocytochemical study of class II major histocompatibility complex molecule (Ia) expression.

Cell surface expression of Class II major histocompatibility complex (Ia) molecules is required for antigen recognition by T cells. To determine the ultrastructural cellular distribution of Ia molecules in the autoimmune disease model acute experimental allergic encephalomyelitis (EAE) we studied central nervous system (CNS) tissues from adult Strain 13 guinea pigs (GP). Experimental allergic encephalomyelitis was induced by sensitization with GP spinal cord homogenate in complete Freund's adjuvant (CFA). Nine of 11 sensitized GP had clinical and histologic EAE whereas unsensitized and CFA-sensitized controls were normal. Central nervous system tissues were reacted with monoclonal antibodies to either GP Ia or T cell surface antigen using an avidin-biotin immunoperoxidase technique and studied by electron microscopy; Ia was found on luminal but not abluminal surfaces of many meningeal and parenchymal vascular endothelial cells in GP with EAE. In EAE perivascular lymphocytes and macrophages and processes of unidentified cells in the parenchyma expressed surface Ia and Ia+ macrophages encircled and phagocytosed myelin. T cells were found predominantly in perivascular inflammatory cuffs. These observations indicate that following immunologic challenge Ia is expressed on luminal surfaces of vascular endothelium and on resident CNS cells, suggesting the possibility that these cells may have active antigen-presenting functions in CNS inflammatory reactions.

Experimental allergic encephalomyelitis in the Lewis rat: A model of predictable relapse by cyclophosphamide

Relapse of experimental allergic encephalomyelitis (EAE) was achieved in Lewis rats by cyclophosphamide (CY). All rats, immunized with an emulsion of guinea pig spinal cord homogenate in complete Freund's adjuvant and treated with 100 mg/kg of CY 21 days postimmunization (pi), developed moderate to severe paralysis 9–14 days following CY injection. A second relapse was observed in 4 of 11 rats reinjected with CY 49 days pi. Histologically, focal mononuclear cell infiltration with or without demyelination of the white matter of the central nervous system was observed. Cyclophosphamide administration caused transient leukopenia and T-cell defect, the resolution of which coincided with relapse of clinical EAE. Lymphocyte proliferative responses to myelin basic protein (BP) and concanavalin A (Con A) and antibody titers to BP were preserved in CY-treated rats. Adoptive transfer of EAE to naive recipients with Con A-activated spleen cells from donors with CY-induced relapse was unsuccessful.

Passive transfer of experimental allergic encephalomyelitis induced by proteolipid apoprotein

In an attempt to obtain insight into the pathogenesis of proteolipid apoprotein (PLP)-induced experimental allergic encephalomyelitis (EAE) in Lewis rats (Yamamura et al. 1986), PLP-sensitized lymph node cells or spleen cells were passively transferred into normal or irradiated (400 rad) recipients after incubation with concanavalin A or PLP. Clinical EAE manifested by paraparesis was successfully transferred into irradiated recipients with 2 − 2.5 × 10 8 of the primary cultured cells and histologic EAE could be transferred with as few as 5 × 10 7 cells into naive recipients. This is the first demonstration of passive EAE induced with PLP-sensitized lymphoid cells and suggests the pathogenetic importance of cell-mediated immunity to PLP.

The mechanism of inhibition of experimental allergic encephalomyelitis in the rat by monoclonal antibody against CD4

Lewis rats with actively induced or passively transferred experimental allergic encephalomyelitis (EAE) were treated with a monoclonal antibody (MAb) which binds to the CD4 antigen of rat helper/inducer T cells. Actively immunized animals treated at the first onset of clinical signs experienced only a mild form of the disease and rapidly recovered while the majority of those treated prophylactically never showed clinical signs of EAE. Passively transferred EAE was also completely inhibited with anti-CD4 MAb. In treated animals which exhibited only mild clinical signs of EAE, spinal cord and cerebellar leukocyte infiltrates were quite similar to those in untreated rats but where anti-CD4 MAb treatment completely prevented clinical EAE, histological signs were minimal or absent. Like Lewis rats which have recovered naturally from EAE, those treated with anti-CD4 MAb were both resistant to a secondary challenge with myelin basic protein and harboured potential encephalitogenic cells which were capable of transferring disease to recipient rats. Disease in these recipients was, however, of much greater severity than that experienced by animals receiving cells from naturally recovered (untreated) donors. These data demonstrate that administration of anti-CD4 MAb to rats can prevent EAE by a mechanism which does not ablate the encephalitogenic CD4 + cells or prevent the development of resistance to EAE but which may inhibit the disease by preventing the function of already activated effector cells.

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.