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Experimental allergic neuritis (EAN) is a T cell-mediated animal model of Guillain-Barré syndrome characterized by inflammation and demyelination of peripheral nerves. EAN can be induced by immunization of rats with bovine peripheral nerve myelin (BPM) or the myelin proteins P2 or P0, but the extent of T cell responses over the course of EAN is incompletely defined. We studied the T cell responses to these proteins and the glycolipid GM1 by enumerating T helper type 1 (Th1)-like cells secreting interferon-gamma (IFN-gamma) after short-term culture of mononuclear cells (MNC) in presence of antigen. Already 7 days post immunization (p.i.) with BPM and before onset of clinical EAN, lymph nodes contained elevated levels of P2 responsive T cells. At the height of EAN on day 14 p.i. and during recovery, T cell levels responding to BPM, P0 and GM1 were also elevated. The same temporal profiles and specificities were registered for antigen reactive spleen MNC. The results implicate that Th1-like cells with multiple specificities including the glycolipid GM1 occur at increased levels in lymphoid organs in EAN rats, and that IFN-gamma may be an important effector molecule in the induction of nerve damage.

Thirty patients with optic neuritis (ON), in which neither multiple sclerosis (MS) nor any other etiology could be discriminated, were reexamined after a mean observeation period of 5 years. Eleven patients revealed oligoclonal IgG in the CSF and in five of them a measles virus antibody response within the CNS was demonstrable. the remaining 19 patients did not display oligoclonal CSF IgG and no local antibody production was detectable. The occurrence of the HLA antigens A3 and B7 in ON did not correlate to the presence of oligoclonal IgG in CSF. the frequencies did not differ from those found in controls. the HLA-B7 linked lymphocyte defined antigen HLA-Dw2 occurred in ON at increased frequency, which was intermediate to that observed in MS and controls. an association was found in ON between oligoclonal IgG in CSF and Dw2. This association was of the same magnitude as in 22 MS patients who had ON as their first symptom of MS. In ON without oligoclonal CSF IgG the frequency of Dw2 was similar to that of controls. No association was observed between the occurrence of the HLA antigens A3, B7 and Dw2, and increased measles antibody titers in serum or a measles virus antibody response in the CNS. The occurrence of oligoclonal IgG in CSF in patients with ON may be assumed to increase the risk of developing MS.

Patients with aseptic meningitis (AM) were examined with the active T cell rosette test, which has been claimed to reflect cell-mediated immunocompetence more accurately than determination of total T cells. Higher percentages of active T cells were demonstrated in CSF compared to blood regardless if specimens were obtained on days 1-4, days 5-10, or later than 20 days after onset of symptoms, Active T cell percentages in CSF decreased when values for specimens obtained on days 5-10 were compared with those taken later than 20 days after onset, while no significant variations of active T cell percentages in blood were observed. The percentages of total T cells were higher in CSF than blood in specimens from days 5-10, and later than 20 days after onset, but no significant fluctuations of total T cells occurred in either CSF or blood over the course of AM.

Autoreactive T cells recognizing myelin basic protein (MBP), proteolipid protein (PLP) and MBP peptides have been described in multiple sclerosis (MS) and optic neuritis (ON), but their role in disease pathogenesis, if any, is unknown. A consistency of the T cell repertoire over the course of MS and ON should facilitate the development of specific immunotherapies. We have examined the T cell responses to autoantigens in two consecutive blood specimens taken from patients with ON and MS, and in two consecutive CSF specimens obtained from ON patients. As read-out numbers of T cells responding to antigen stimulation by the secretion of interferon-gamma were estimated. Pronounced differences in occurrence and numbers of T cells recognizing MBP, MBP peptides with the amino acid sequences 63-88, 110-128 and 148-165, and PLP were noticed in individual ON and MS patients over the course of disease. The MBP peptide among those three included, that was predominantly recognized by T cells in the individual patient, also varied over the course. The quantitative and qualitative changes of the myelin antigen-specific T cell response in MS and in ON, the latter to a certain extent reflecting the situation in early MS, do not favor the future useful development of specific immunotherapies in these diseases.

OBJECTIVES Multiple sclerosis (MS) is characterized by high levels of circulating mononuclear cells (MNC) that respond to myelin proteins like myelin basic protein (MBP) in vitro by expressing mRNA of both pro-inflammatory cytokines, e.g. interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha) and lymphotoxin (LT) that may make MS worse, and anti-inflammatory cytokines like IL-4, IL-10 and transforming growth factor-beta (TGF-beta) that may act beneficially. Substances that down-regulate cytokines such as TNF-alpha or promote IL-10 or TGF-beta can be anticipated to affect MS beneficially. MATERIAL AND METHODS In situ hybridization to detect and enumerate IFN-gamma, TNF-alpha, LT, IL-4, IL-10 and TGF-beta mRNA expressing blood MNC after stimulation with myelin basic protein (MBP), control antigens and without antigen in presence and absence of Linomide (roquinimex, LS-2616) was employed. In parallel, ELISPOT assay to detect MBP- and PHA-reactive IFN-gamma secreting blood MNC+/-Linomide was used. RESULTS Here we report that Linomide, a synthetic immunomodulator, at concentrations effective in vivo reduces the number of MBP-reactive TNF-alpha and increases MBP-reactive IL-10 and TGF-beta mRNA expressing MNC from MS patients' blood when analysed in vitro. Compared to dexamethasone, Linomide up-regulated levels of blood MNC expressing mRNA of TGF-beta after culture in presence of MBP. CONCLUSIONS Changes of cytokine balance towards a production of anti-inflammatory cytokines could be a desirable effect to be evaluated in future drug studies of Linomide-like substances. At present, Linomide is not evaluable in MS clinical trials due to side-effects.

Introduction Restricted usage of V alpha and beta genes has been found in several diseases, which exert autoreactive T cells. So far no information on T cell receptor (TCR) usage in degenerative diseases is available. Since T cells may be involved in pathogenesis of Parkinson's disease, the analysis of the TCR repertoire is of importance. Material and methods We have tested the frequency of 6 V beta-subtypes and the most common V alpha-subfamily on peripheral blood lymphocytes in 21 PD patients and 20 controls, separately on CD4+ and CD8+ T cells. For our study Diversi--T, the first available Mab set specific for TCR V regions, was used. Results As a results, no significant differences were found with one exception, i.e., lower frequency of V beta 8(a) expression on CD8+ T cells in PD patients than in controls. Conclusion The failure of preferential usage or lack of usage of the tested V-chain alleles by CD4+ T cells argues against an involvement of helper T cells in PD induction.

The mobility of the immunoglobulins G, A and M, beta-trace protein and gramma-trace protein on isoleectric focusing of serum and CSF was determined by immunofixation using specific antisera. Polyclonal IgG migrated as multiple bands between pH 4.7--8.6, polyclonal IgA as multiple bands between pH 4.9--6.1 in CSF and serum. IgM could not be identified in normal CSF or serum. beta-trace protein gave three bands at pH 8.0, 8.4 and 7.4--7.5, respectively, while gamma-trace protein gave one single band at pH 9.5--greater than 9.5. Oligoclonal IgG in CSF in multiple sclerosis and neurosyphilis migrated between pH 8.6--greater than 9.5 and was easily discriminated from other proteins.

Genetic factors influence the susceptibility to multiple sclerosis (MS). This disease is accompanied by augmented T cell responses to CNS myelin components such as myelin basic protein. To evaluate the familial occurrence of such T cell autoreactivity, we have studied 12 MS families including 37 healthy first-degree relatives for occurrence of numbers of interferon-gamma (IFN-gamma) secreting cells among blood mononuclear after culture in presence of myelin basic protein (MBP), eight synthetic MBP peptides and the control antigen acetylcholine receptor (AChR). There were no differences between MS patients and healthy family members regarding frequencies of autoreactive T cells recognizing MBP, the eight different MBP peptides or AChR. None of the MBP peptides predominated as T cell antigen among the MS patients or their unaffected family members. In some families the highest number of MBP peptide reactive T cells were found among unaffected family members. No correlation was observed between numbers of MBP or MBP peptide reactive T cells in various subjects and their HLA-DR-DQ phenotypes. In conclusion, this study has revealed the presence of MBP and MBP peptide reactive T cells of similar frequencies in MS patients and their healthy family members.

IFN-beta may modify the clinical course of multiple sclerosis (MS) but is not curative, and there are also patients whose disease does not respond to IFN-beta as currently administered. Tests are warranted with a capacity to early discriminate responders from non-responders, thereby altering treatment option for the individual patient. In vitro effects of IFN-beta on expression of activation-associated cell surface markers and cytokine production need to be explored in this context. Here we report on the influence in vitro of IFN-beta on blood mononuclear cells (MNC) prepared from MS patients and healthy controls. MNC were subjected to short-term culture in the presence of IFN-beta at concentrations of 100 U/ml and 1000 U/ml. Expression of cell surface molecules CD40, CD69, CD80, CD86, CD95 and HLA-DR was measured by flow cytometry. IL-10 and IL-12 p40 production in culture supernatants was measured by ELISA. MNC exposed to IFN-beta in vitro enhanced expression of the co-stimulatory CD80, CD86, the early activation antigen CD69 and the cell death receptor CD95. Expression of CD40 and HLA-DR was not influenced. IFN-beta increased IL-10 but suppressed IL-12 p40 production. In vitro effects of IFN-beta on MNC were similar in MS patients and in healthy subjects, except that IFN-beta-induced augmentation of CD86 and CD69 expression was less pronounced in MS, in particular in untreated MS patients. Individual MS patients clearly responded differently to IFN-beta in vitro in comparison with the majority of patients in this cross-sectional study. In conclusion, anti-inflammatory effects of IFN-beta on blood MNC include augmentation of IL-10 production and suppression of IL-12 p40 production, which are accompanied by enhancement of CD69, CD80, CD86 and CD95 expression. The less pronounced IFN-beta-induced effects on CD86 and CD69 expression in MS vs controls might reflect a defect in immunoregulation in MS. Larger groups should be evaluated, and follow-up studies performed in MS patients before/during IFN-beta treatment in relation to clinical outcome measures to evaluate the usefulness of these markers for possible differentiation between responders and non-responders to IFN-beta treatment.

Objectives To set up an epidemiological surveillance system of Guillain-Barre syndrome (GBS) in Sweden and to evaluate the pilot practice of the system for 2 years. Material and methods A network of neurologists reported incident patients with a GBS diagnosis among a general population of 4.5 million inhabitants in Sweden during 1996 1997. Historical GBS data from the national hospital in-patient registry were used for predicting incidences and controlling under-reporting. Results One hundred and seventeen cases were reported. No alarm signals were identified during the study period. A re-analysis of data in 1996 revealed: 1) a higher than expected incidence in the population aged below 40 years in January, and 2) 27% under-reporting mainly associated to one hospital and to GBS patients hospitalized outside neurological departments. Threshold values for GBS incidences among the general and selected populations were obtained from corrected data. Conclusion - The Swedish Network for GBS Surveillance can provide immediate support for epidemiological evaluation of suspected outbreaks or increased GBS incidence.

By enumerating cells secreting IgG antibodies of particular specificities using an enzyme-linked immunospot (ELISPOT) assay, the B-cell responses to Torpedo acetylcholine receptor (AChR) and its alpha-, beta-, gamma- and delta-subunits in peripheral blood from patients with myasthenia gravis (MG), and controls with other neurological diseases (OND) as well as healthy subjects were determined. Compared to controls, the patients with MG had elevated numbers of B cells secreting antibodies against AChR and its alpha-, beta-, gamma- and delta-subunits in peripheral blood in parallel. The mean numbers of anti-AChR antibody secreting cells were about 17 per 10(5) blood MNC, and for the subunits 10 to 15 in MG patients, compared to between 0.8 and 1.9 per 10(5) blood MNC in OND patients, and 0.1 to 0.3 in healthy controls. Such B cells detected in controls probably represent naturally occurring B cells responded to AChR and its subunits. The finding that most (60%) MG patients had B cells predominantly recognizing the alpha-subunit may be an indirect argument for the existence of a main immunogenic region (MIR). In the remaining 40% of patients with MG the predominant B-cell responses were directed to beta-, gamma- or delta-subunit. The data suggest that all four AChR subunits may function as strong immunogens in MG, though the alpha-subunit may be the major immune target in a substantial proportion of MG patients.

The CD30 molecule, a member of tumor necrosis factor superfamily, has been suggested to be preferentially expressed and released in soluble form by activated T cells that produce T helper 2 type (Th2) cytokines. To evaluate whether determination of soluble CD30 (sCD30) levels could have a diagnostic value in diseases associated with Th1 and Th2 cytokine involvement, we investigated sCD30 in plasma and cerebrospinal fluid (CSF) from patients with multiple sclerosis (MS), HIV infection and other nervous system diseases. There was no statistically significant difference for plasma sCD30 levels in these clinical groups. However, patients with HIV infection had higher levels of sCD30 in CSF than MS patients. The mean sCD30 values were 3 to 6 folds higher in plasma than in CSF in all patient groups. No relationships were found between sCD30 levels and different clinical variables of MS and HIV infection, except that higher plasma sCD30 levels in HIV-infected patients were found in those with higher CD4+ T cell counts (> 200 x 10(6)) compared to the group with lower cell counts. The findings indicate that determinations of plasma and CSF sCD30 levels in MS and HIV infection have limited or no value as diagnostic or prognostic indicator.

Cerebrospinal fluid (CSF) and serum from 15 patients with narcolepsy were examined regarding presence of oligoclonal bands on isoelectric focusing, IgG index elevation as indicator of intrathecal IgG production and CSF/serum albumin ratios. Two of 15 patients (13%) showed oligoclonal IgG bands in CSF, one of whom had increased IgG index. Slight disturbance of blood-brain-barrier function as reflected by elevated CSF/serum albumin ratios was present in 4 other patients. The frequency of oligoclonal IgG bands in CSF from patients with narcolepsy is within the range of what is found in other neurological, non-inflammatory diseases. These findings do not support the hypothesis of an immune-mediated pathogenesis of narcolepsy.

A sensitive sandwich ELISA was applied to the measurement of the terminal component of complement C9 in CSF and plasma from 40 tension headache patients (reference group), 33 affected by clinically definite MS and 10 by aseptic meningitis. The levels of C9 in plasma were increased in aseptic meningitis. The determinations of CSF/plasma C9 ratio and C9 index, equal to (CSF C9/plasma C9): (CSF albumin/plasma albumin), thus accounting for changes of plasma C9 levels as well as damaged blood brain barrier, documented the existence of local consumption of C9 in aseptic meningitis. In contrast, only borderline alterations were evident in MS. The results indicate that local consumption of total C9 in CSF is an additional variable reflecting an acute inflammation within the CNS, but not demonstrable in MS, a chronic inflammatory CNS disorder.

Beta-2-microglobulin (beta 2m) levels were studied in the cerebrospinal fluid (CSF) and plasma of 52 patients with clinically definite multiple sclerosis (MS) and of 14 with aseptic meningo-encephalitis (AM). Reference values for beta 2m were defined in 72 subjects of different age groups with tension headache (TH). Plasma levels of beta 2m increased with age in TH controls, particularly in the older age group, while no significant age variation could be detected for CSF beta 2m. Increased levels of beta 2m were found in the CSF and plasma of AM patients and in the CSF in MS. Calculations of CSF/plasma beta 2m ratios showed higher values in AM, reflecting intrathecal beta 2m synthesis, while only borderline alterations were evident in MS. Demonstration of intrathecal production of beta 2m is an additional CSF finding that may be useful in evaluating, among others, inflammatory nervous system disorders, but it has limited importance in the study of MS patients.

A patient is described having Borrelia burgdorferi spirochetal infection clinically affecting central motor neurons selectively and without any sensory impairment. Diagnosis was based on elevated B. burgdorferi IgG antibody titers in cerebrospinal fluid (CSF) and titer normalization at clinical recovery. This occurred promptly and was complete after penicillin treatment despite 14 months of progressive central nervous system (CNS) dysfunction, favouring the hypothesis of the presence of the organism within the CNS. CSF findings characteristic of neuroborreliosis were registered, including parallel occurrence of mononuclear plecytosis, severe blood-brain barrier damage and marked CSF IgM index elevation of prolonged duration. Some earlier reports of CNS manifestations related to B. burgdorferi are reviewed.

— The intrathecal production (ITP) of the immunoglobulins (Ig) G, A, M and D was examined in untreated patients with multiple sclerosis (MS) and controls. Sensitive sandwich enzyme-linked immunosorbent assay systems were applied to the determination of IgA, IgM and IgD levels in the cerebrospinal fluid and plasma from a group of 61 consecutive MS patients (41 relapsing-remitting and 20 secondary chronic progressive MS). Age-related reference limits for all Ig variables were defined in a group of 57 patients with tension headache. ITP of IgG was demonstrated in 85% of the MS patients, ITP of IgA in 41%, of IgM in 44% and of IgD in 18%. Among 9 MS patients with normal IgG index, 3 displayed ITP of IgA, 3 of IgM and 1 of IgD. Plasma IgA was elevated in 20% and plasma IgM in 24% of the MS cases. No significant variation of Ig ITP was demonstrated in a different group of 27 untreated MS patients examined during exacerbation and remission. Among control patients with other inflammatory nervous system diseases, ITP of IgA, IgM and IgD was found more frequently, and of IgG in a smaller percentage compared with MS patients.

Serum and cerebrospinal fluid (CSF) from 28 patients with multiple sclerosis (MS), 14 with other neurological diseases (OND) and 31 control subjects with tension headache were analysed for presence of IgM antibodies against measles, mumps and varicellae zoster by a specific enzyme-linked immunosorbent assay (ELISA). This technique excluded false positive reaction due to possible presence of rheumatoid factor. Twelve of the 28 patients with MS had IgM antibodies in serum and 4 in CSF, the latter always being accompanied by presence of corresponding IgM antibodies in serum. Six patients had mumps specific IgM, 5 had measles specific IgM and 3 varicellae specific IgM. In 2 patients, viral IgM antibodies were demonstrated in serum against 2 different viruses. Among the 14 OND patients, one with Wilson's disease had demonstrable serum IgM varicellae antibodies and one with radicultis had elevated serum and CSF measles and varicellae IgM antibodies. Among 31 controls, 2 had IgM antibodies in serum, one against varicellae and one against mumps. No correlations were found between viral IgM antibodies and CSF IgM index, serum IgM levels or blood-brain barrier state. Our data show that MS may be accompanied by a systemic IgM response against the 3 viruses tested, occasionally against 2 of the 3 different viruses simultaneously. The occurrence in MS of virus-specific IgM may be a reflection of viral reactivation and/or polyclonal B cell activation.

Female preponderance in many diseases suggested with autoimmune pathogenesis, multiple sclerosis (MS) being classified as one of them, indicates a role for hormonal factors such as estrogen in disease development. To bypass monthly hormonal fluctuations in females, we evaluated in male patients with MS and male blood donors the effect of 17-beta-estradiol on numbers of IgG, IgA and IgM producing cells in cultures of peripheral blood lymphocytes. While estradiol alone had no effect, estradiol in combination with poke-weed mitogen (PWM) yielded in both groups higher numbers of IgG and IgA producing cells when compared with numbers obtained by PWM stimulation alone, indicating an additory effect of estradiol to that of PWM on B cell maturation. This effect was less pronounced in MS than in blood donors, especially for IgG producing cells, probably reflecting higher B cell activation in vivo taking place in MS. On the contrary, cells producing IgG, IgA and IgM antibodies against myelin, myelin basic protein and measles virus were not detectable after stimulation with PWM, nor with PWM and estradiol. Estradiol can in many patients with MS and in blood donors be considered a potent co-activator of B cells in presence of B cell stimulating factor in the form of PWM.

Elevated concentrations in CSF of the immunoglobulins G, A, and M, when expressed as a percentage of the total protein concentration of CSF, were demonstrated during the initial phase of the course of the disease in a patient with Guillain-Barre syndrome (GBS). A slight elevation of the relative concentrations of IgG and IGM in CSF were also registered later in the course of the disease, at the time when the patient's neurological symptoms were in regression. Multiple discrete IgG bands were demonstrated by agarose electrophoresis in serum and in CSF during the first 2 weeks after onset of the neurological symptoms, and also after week 2 and still during week 12 after onset. The finding of transient oligoclonal IgG in a parient with GBS may suggest stimulation with e.g. viral antigen.

With the use of enzyme-linked immunosorbent assay (ELISA) for demonstration of specific viral antibodies in 21 patients with mumps meningitis (MM), 19 (91%) displayed evidence for synthesis within the central nervous system (CNS) of IgG antibodies and 11 (52%) of IgM antibodies. Only one MM patient did not show local antibody production, but he had high mumps IgM titers in serum. Elevated IgG and IgM indices reflecting local IgG and IgM production, were found in 4 (19%) and 17 (81%) of the patients, respectively, and frequently did not coincide with local synthesis of specific antibodies. None of 21 control patients with acute aseptic meningitis of other etiology showed evidence for a local antibody response against mumps. In one of the MM patients, local production of measles IgM antibodies was demonstrated, reflecting an IgM response within the CNS against an antigen unrelated to the disease. This study concludes that in less serious CNS infections, such as MM, highly sensitive methods as ELISA are needed to demonstrate intrathecal specific viral immune response.

The concentrations of two of the complement factors, beta1C/beta1A globulin (C3) and beta1E globulin (C4) have been determined in unconcentrated cerebrospinal fluid (CSF) and in serum obtained from patients with multiple sclerosis, from patients with other neurological disorders, and from healthy controls. A positive, statistically significant correlation was found in all three groups between the absolute concentrations in CSF of these two complement factors and the total protein concentrations. No statistically significant differences were found between the concentrations of the two complement factors in the specimens from the three groups, nor in the specimens obtained during an exacerbation of multiple sclerosis and those obtained during remission of this disease.

Genetic subtypes of group-specific component (Gc), a protein possibly influencing susceptibility to human immunodeficiency virus infection, were assessed by isoelectric focusing of plasma from 88 patients with multiple sclerosis (MS) and related to frequencies among 3394 control individuals subjected to paternity studies. We found no clear association between MS and frequencies of phenotypes or alleles of Gc protein.