Increased muscarinic cholinergic receptor density on CD4 lymphocytes in progressive multiple sclerosis

We measured the density and affinity of muscarinic cholinergic receptors (MR) in 29 chronic progressive and ten stable multiple sclerosis (MS) patients and 27 control subjects using [3H]N-methyl-scopolamine. The density of MR on CD4+ lymphocytes was significantly higher in chronic progressive MS (CPMS) than in controls (7.9 +/- 0.7 vs. 4.5 +/- 0.4 fmol/10(6) cells, p less than 0.001). Stable patients did not differ significantly from control subjects. Receptors of the M1 subtype were measured on CD4+ lymphocytes of nine patients and seven controls with the selective antagonist [3H]methylpirenzepine: M1/total receptor ratio was 64.1% in CPMS and 81.2% in controls, suggesting a selective increase of M2-type MR in CPMS. The findings may relate to parasympathetic denervation hypersensitivity of lymphocytes or to lymphocyte activation which is known to be associated with increased MR number.     

Sympathetic skin responses are decreased and lymphocyte beta-adrenergic receptors are increased in progressive multiple sclerosis

Immune abnormalities, including deficient CD8 lymphocyte-mediated suppression, have been implicated in the progression of multiple sclerosis (MS). The peripheral sympathetic branch of the autonomic nervous system innervates the lymphoid organs and affects immune function. Animals with an ablated sympathetic nervous system develop more severe experimental allergic encephalomyelitis than control animals and exhibit an increased density of beta-adrenergic receptors on their lymphocytes. Experimental allergic encephalomyelitis shares many features with MS. Accordingly, we investigated the psychogalvanic skin reflex in patients with rapidly progressive MS and found that 13 patients (57%) lacked this sympathetic-mediated response. The density of beta-adrenergic receptors on lymphocyte subsets was increased in progressive MS, most notably on the CD8 suppressor/cytotoxic subset. B lymphocytes had the greatest number of receptors with 12.1 +/- 1.8 fmol/10(6) cells in control subjects and 18.7 +/- 2.6 fmol/10(6) cells in patients with MS. CD8 lymphocytes possessed an intermediate number of receptors with 3.4 +/- 0.4 fmol/10(6) cells in control subjects and 9.1 +/- 1.6 fmol/10(6) cells in patients with MS. CD4 lymphocytes demonstrated the fewest receptors with 1.2 +/- 0.1 fmol/10(6) cells in control subjects and 1.8 +/- 0.4 fmol/10(6) cells in patients with MS. No differences in the affinity or function (cyclic adenosine monophosphate levels in response to 10(-5) M (-)isoproterenol) of the adrenergic receptor were found when patients with progressive MS and control subjects were compared. Autonomic abnormalities in progressive MS and the increased beta-adrenergic receptor density found on CD8 lymphocytes may be related.     

Increased CD8+ central memory T cells in patients with multiple sclerosis

A T-cell-mediated autoimmune process against central nervous system myelin is believed to underlie the pathogenesis of multiple sclerosis (MS). Formation of immunological memory is based on the differentiation of na?ve T cells to memory T cells after exposure to antigens and specific cytokines. The aim of this study was to analyse peripheral blood mononuclear cells in patients with MS for different T-cell subsets including na?ve and memory T cells. Flow cytometry and enzyme-linked immunosorbent assay were used to analyse memory T-cell subsets and plasma concentration of interleukin-15 (IL-15) in peripheral blood of MS patients, patients with other neurological disorders and healthy controls. MS patients had a skewed distribution of T cells with an increased level of CD8+/CCR7+/CD45RA - central memory T cells (TCM) compared to healthy controls. In addition, MS patients showed significantly higher levels of plasma IL-15 than healthy controls did. Upregulated CD8+ TCM in MS patients may reflect a persistent chronic inflammatory response that may have been induced during early stages of the disease. This derangement may be important for maintaining chronic inflammation in MS.     

Increased T cell expression of CD154 (CD40-ligand) in multiple sclerosis

CD154 (CD40-ligand, gp39), expressed on activated T cells, is crucial in T cell-dependent immune responses and may be involved in the pathogenesis of multiple sclerosis (MS). We studied cerebro-spinal fluid and peripheral blood T cell expression of CD154 in MS by flow cytometry. Patients with secondary progressive MS (SPMS) had constitutive CD154 expression on CD4 and CD8 T cells in blood. Constitutive CD154 expression was not observed in patients with relapsing-remitting MS (RRMS) or clinically isolated syndromes (CIS) suggestive of demyelinating disease. After ex vivo activation CD154 was, however, expressed on a higher percentage of T cells from patients with CIS or RRMS than from healthy control subjects. These results suggest involvement of CD154 in the pathogenesis of MS, and the shift from a relapsing-remitting to a secondary progressive disease course may be associated with constitutive, systemic CD154 expression.     

A longitudinal study of the T cell activation marker CD26 in chronic progressive multiple sclerosis

Activated T cells are implicated in the pathogenesis of multiple sclerosis (MS), an autoimmune demyelinating disease of the central nervous system. Serial measurements of T cell activation molecules and T cell subpopulations were performed over 12 months in patients with chronic progressive multiple sclerosis and healthy controls, and correlated with clinical indices of disease progression measured by standardized disability scores. Of the markers studied, the activation molecule CD26 appeared to exhibit a more consistent pattern and to be elevated in MS patients; therefore we concentrated our attention on this marker, especially in view of recent evidence of its role in T cell activation. In this small patient group, the elevation of CD26 in the MS patients did not reach statistical significance, when compared to the level in the controls. Interestingly, the percentage of CD26 but not CD25 or HLA-DR correlated with the MS patients' disability scores. However, the clinical significance of this observation as an indication of disease activity in chronic progressive MS remains to be demonstrated in studies of larger patient populations.     

Molecular analysis of the CD8 gene in multiple sclerosis

Analysis of DNA fragments of the CD8 gene following digestion with three restriction enzymes showed no differences between patients with chronic progressive multiple sclerosis (MS), familial MS and controls. These abnormalities suggest that any abnormalities in the expression of CD8 protein in MS are not likely to be due to a gross deletion or rearrangement of the CD8 gene.     

Increased circulating T cell reactivity to GM3 and GQ1b gangliosides in primary progressive multiple sclerosis.

We have previously shown that patients with primary progressive multiple sclerosis (MS) have significantly elevated plasma levels of antibody to GM3 ganglioside compared to patients with relapsing-remitting MS, healthy subjects and patients with other neurological diseases. Anti-GM3 antibody levels were elevated also in patients with secondary progressive MS but to a lesser extent than in primary progressive MS. As gangliosides are particularly enriched in the axonal membrane, these findings suggested that antiganglioside immune responses might contribute to the axonal damage in progressive forms of MS. The present study was performed to determine whether peripheral blood T cell responses to GM3 are also increased in progressive MS. Blood was collected from 98 untreated patients with MS (40 with relapsing-remitting, 27 with secondary progressive and 31 with primary progressive MS), 50 healthy subjects and 24 patients with other disorders of the CNS, and reactivity to GM1, GM3, GD1a, GD1b, GD3, GT1b, GQ1b and sulphatide was assessed by 6-day T cell proliferation assays. Increased T cell reactivity to GM3 and GQ1b occurred significantly more often in patients with primary progressive MS than in healthy subjects and patients with other CNS diseases. These findings suggest that ganglioside-specific T cells may contribute to the axonal damage in primary progressive MS.     

The germline repertoire of T cell receptor beta-chain genes in patients with chronic progressive multiple sclerosis

The T cell receptor (TcR) beta-chain germline gene repertoire of multiple sclerosis (MS) patients was compared to that of 100 normal individuals. No differences in the number of gene segments defined by probes representing 14 different human V beta subfamilies and the constant region genes were found. The distribution of haplotypes defined by restriction fragment length polymorphism (RFLP) alleles detected with V beta 8, V beta 11, and C beta probes in the MS patients was significantly different from that found in normal individuals. Because 84% of the MS patients were DR2+, the findings in these patients were compared to a second group of 43 normals who were DR2+. The distribution of TcR haplotypes in MS patients was also significantly different from that in the DR2+ normals. The data suggest that an MS susceptibility gene(s) may be located in the region of the TcR beta-chain gene complex.     

Dual labelling of circulating CD8 cells in patients with multiple sclerosis.

Peripheral blood T cell phenotypes have been analysed in serial samples from patients with multiple sclerosis, their unaffected relatives and controls using a panel of antibodies chosen to distinguish T suppressor and activated suppressor cells from other CD8 lymphocytes. Overall, the percentage of Leu 2a cells correlated with alterations in the Leu 2a/15 suppressor sub-population (r = 0.79, p less than 0.001). Fewer circulating Leu 2a and Leu 2a/15 positive cells were identified in multiple sclerosis patients than unaffected individuals but there was no alteration in percentage of activated (Leu 2a/DR) CD8 cells. These findings suggest that the fluctuations in CD8 cells, characteristic of patients with multiple sclerosis, are due to alterations in T suppressor phenotype; this may then lower the threshold for activation of other T cell subpopulations.     

Immunohistological analysis of T lymphocyte subsets in the central nervous system in chronic progressive multiple sclerosis

In an analysis of pooled data, we have found that cytotoxic-suppressor T cells outnumber the helper-inducer subset and also the population of cells bearing the pan-T cell marker in specimens of CNS from patients dying with MS (Booss et al. 1983). In the present study of individual data, we have reviewed the case histories to determine if these findings were consistent in various clinical settings. Variables examined included disease duration, tempo of evolution, immunosuppressive therapy, and other potentially immunomodulating features such as tumours. The predominance of the cytotoxic-suppressor subset was not found to be altered by any of these variables. We also present an individual data analysis of cases dying without known CNS disease and of cases with chronic non-inflammatory CNS disease. We found that the low but consistently observed number of T cells was apparently unrelated to the age of the individual or the site of the CNS sampled. Analysis of selected perivascular infiltrates showed, in contrast to the CNS parenchyma, that the pan-T cells and each of the subsets were approximately equal in proportion. Consideration of this observation and of the cytotoxic-suppressor subset preponderance in the parenchyma is compatible with the possibility of antigenic modulation of the T cell differentiation antigens. Finally, the potential contribution of perivascular infiltrates to the CSF pleocytosis is considered.     

Proteolytic shedding of the macrophage scavenger receptor CD163 in multiple sclerosis

The scavenger receptor CD163 is selectively expressed on tissue macrophages and human monocytes. CD163 has been implicated to play a role in the clearance of hemoglobin and in the regulation of cytokine production by macrophages. Membrane CD163 can be cleaved by matrix metalloproteinases (MMP) resulting in soluble CD163 (sCD163). In the present report the shedding of CD163 was investigated in multiple sclerosis (MS). An upregulation of plasma sCD163 and a down regulation of membrane CD163 in MS patients compared to healthy controls was observed. The levels of plasma sCD163 correlated with plasma MMP-9 levels in controls, but not in MS patients. Moreover, evidence was obtained for CD163-cleaving MMP activity in plasma of MS patients. Finally, the increased proteolytic shedding of CD163 correlated to reduced plasma levels of circulating inflammatory cytokines. Collectively, our results provide evidence for proteolytic shedding of CD163 in MS and suggest a possible link to cytokine production.     

The role of the Epstein-Barr virus receptor CD21 in multiple sclerosis

Multiple Sclerosis (MS) is characterised by a chronic inflammation and demyelination of brain and spinal cord with a yet unknown aetiology. Based on multiple epidemiological and immunological studies, which suggest a role of Epstein–Barr virus (EBV) infection in the pathogenesis of MS, we investigated CD21 (CR2, complement receptor type 2), which serves as the EBV receptor. Serum concentrations of soluble CD21 receptor (sCD21) were determined in MS patients and controls. In accordance with findings in other autoimmune disorders decreased sCD21 levels were found in MS patients. On ß-IFN treatment serum sCD21 concentrations further decreased. To explore the role of the CD21 gene for MS susceptibility and the altered CD21 levels in MS patients we performed exon sequencing of the CD21 gene. While we identified new single nucleotide polymorphism (SNP) and confirmed previously reported SNPs, none of the SNPs was associated with MS. Our findings demonstrate that sCD21 expression is altered in MS patients similar to other autoimmune diseases although no evidence was found for a specific role of the CD21 gene in MS.     

JC virus induces a vigorous CD8+ cytotoxic T cell response in multiple sclerosis patients

We sought to compare the ongoing CD8+ cytotoxic T lymphocytes (CTL) immune response of MS patients to self and viral antigens. Using 51Cr release and tetramer staining assays, we found that the CTL response against VP1, the major capsid protein of the polyomavirus JC (JCV), was significantly higher than the one against epitopes of MBP and PLP. The JCV-specific CTL response was also significantly stronger in MS patients than healthy control subjects. These findings may shed a new light on the recent events related to the development of progressive multifocal leukoencephalopathy in three natalizumab-treated patients.     

CD4 and CD8 lymphocyte subsets in cerebrospinal fluid and peripheral blood from patients with multiple sclerosis, meningitis and normal controls

Objectives - To study the distribution of CD4⁺ and CD8⁺ T-cell subsets in cerebrospinal fluid (CSF) and peripheral blood from patients with multiple sclerosis (MS), meningitis, other neurological diseases and healthy controls.
Material and methods - The expression of markers for naive and memory cells (CD45RA⁺ and CD45RO⁺), and helper/inducer cells (CD29⁺) on CD4⁺ cells as well as CD45RO⁺ and killer/effector (S6F1⁺) on CD8⁺ cells was investigated in cerebrospinal fluid (CSF) and peripheral blood from patients with multiple sclerosis (n=28), meningitis (n = 13), other neurological diseases (n = 16), and healthy controls (n = 16) by 2-color flow cytometry.
Results - The majority of T cells in the CSF of the 4 groups exhibited the phenotype of memory cells (CD45RO⁺) on both CD4⁺ and CD8⁺ cells. The proportion of helper/inducer (CD29TD4⁺ in CD4⁺) cells was also larger in the CSF compared to peripheral blood in the 3 patient groups and controls investigated. In contrast, CD8⁺ cells with killer/effector (S6F1+) phenotype were fewer in CSF compared to peripheral blood in all 4 groups. There were no significant differences between patients and controls regarding the distribution of these activation markers in the CSF or peripheral blood.
Conclusion - Our observations support the notion that activated T cells of both CD4⁺ and CD8⁺ phenotype selectively pass the blood-brain barrier under both pathological and normal conditions.     

Factors influencing circulating OKT8 cell phenotypes in patients with multiple sclerosis.

Peripheral blood OKT8 cell phenotypes were correlated with measurements of plasma cortisol and serological evidence for exposure to 15 infectious agents, in longitudinal studies involving 13 patients with multiple sclerosis, 13 of their siblings, nine spouses and 13 unrelated controls; 44/48 individuals were HLA typed. Neither circadian rhythms, nor exposure to any one infectious agent accounted for the serial changes in OKT8 cells but there was an association between the presence of HLA-DR2 and periodic reductions in OKT8 cells irrespective of clinical status. Taken with previously reported serial observations in patients and cohabiting relatives, this finding provides indirect evidence for an interplay between environmental and genetic factors in determining OKT8 cell phenotypes in multiple sclerosis.     

The CD4+ T-cell subset lacking expression of the CD28 costimulatory molecule is expanded and shows a higher activation state in multiple sclerosis

Multiple sclerosis (MS) is a chronic debilitating disease, in which T-cells are considered to play a pivotal role. CD28 is the quintessential costimulatory molecule on T-cells and its expression declines progressively with repeated stimulations, leading to the generation of CD28(-) T-cells. Our aim was to examine whether CD4(+)CD28(-) T-cells were enriched in MS patients, and characterize the phenotype of this subset in MS patients and healthy controls (HC). All these changes could provide these CD4(+)CD28(-) T-cell characteristics that might be involved in the pathogenesis of MS, turning this T-cell subset into a potential target for future therapeutic strategies.     

Expansion of regulatory CD8+ T-lymphocytes and fall of activated CD8+ T-lymphocytes after i.v. methyl-prednisolone for multiple sclerosis relapse

INTRODUCTION: Multiple sclerosis (MS) is a multifocal chronic inflammatory demyelinating disease of the central nervous system. Axonal damage correlates with the presence of macrophages and CD8+ T-lymphocytes at brain lesions. The gold standard of therapy at MS relapse are iv glucocorticoids (GC). The aim of the study was to assess the changes on the different subsets of circulating CD8+ T-lymphocytes at relapse and after iv GC therapy. PATIENTS AND METHODS: We consecutively studied 20 patients at MS relapse before and at day 5 after initiation of i.v. methyl-prednisolone (MP) therapy (1 g/day for 3-5 days). CD4+ and CD8+ T-lymphocytes subsets were studied by multiparametric flow-cytometry. As control group, 18 healthy subjects were studied. RESULTS: Treatment with i.v. MP suppressed activated (CD8+CD38+HLA-DR+, p=0.05) and effector memory (CD8+CD27-CD45RO+) T-lymphocytes (p=0.07). By contrast, an increase of na?ve (CD8+CD27+CD45RO-) (p=0.07) and regulatory CD8+CD25+ T-lymphocytes was observed (p<0.002). At MS relapse, there was an inverse correlation between regulatory CD8+CD25+CD28- T-lymphocytes and activated CD4+ (r = -0.6; p=0.012) and CD8+ (r = -0.66; p=0.004) T-lymphocytes. After i.v. MP treatment, positive correlation between regulatory CD4+CD25+high T-lymphocytes and CD8+CD25+ T-lymphocytes was observed (r=0.74; p<0.0001). CONCLUSIONS: Our data suggest that i.v. MP may contribute to changes observed on the differentiation of CD8+ T-lymphocytes, namely blocking their complete maturation, and expansion of regulatory CD8+ T-lymphocytes. We hypothesize an additional effect of i.v. MP in inhibiting axonal damage which may add a neuroprotective effect on MS relapse.     

Intense immunosuppression in chronic progressive multiple sclerosis: the Kaiser study.

The value of a short course of intensive immunosuppression with cyclophosphamide in stabilising chronic progressive multiple sclerosis (MS) was examined in a randomised single-blinded, placebo-controlled clinical trial. Forty two patients, from the Kaiser Permanente Medical Care Program, Northern California, were studied. Twenty two patients received a short course of cyclophosphamide in an outpatient neurology clinic until their leucocyte counts fell below 4000/mm3, and 20 patients received folic acid. Level of disability, impairment of functional systems, and performance of social roles were assessed before randomisation and reassessed 12, 18, and 24 months after therapy. In both the cyclophosphamide and folic acid groups, the mean level of disability increased from the baseline examination to the 12 month follow up examination (the primary endpoint) by 0.5 on Kurtzke's Expanded Disability Status Scale, indicating similar disease progression in the two groups. Although immunosuppression therapy can be safely administered to MS patients in an outpatient clinic, evidence of substantial benefits was not found.     

多发性硬化患者CD8+记忆性T细胞亚群穿孔素和颗粒酶-B表达的研究

目的 测定多发性硬化(MS)患者外周血中CD8+记忆性T细胞亚群效应细胞因子的表达,并将其与MS病情严重程度进行相关分析.方法 利用四色-流式细胞术检测未经治疗的MS患者、其他神经系统疾病(OND)患者和正常对照(NC)成员组外周血表达穿孔素和颗粒酶-B的CD8+记忆性T细胞亚群数量,并利用扩展的残疾状况量表(EDSS)对MS患者病情严重程度做评分.结果 与NC组比较,MS患者外周血表达颗粒酶-B的CD8+效应记忆性T细胞(TEM)和终末效应记忆性T细胞(TerTEM)均明显减少,比较差异有统计学意义(P＜0.05);表达穿孔素和颗粒酶-B的TEM数量与EDSS呈负相关(r=-0.493,P=0.027;r=-0.594,P=0.009).结论 CD8+TEM参与MS相关的CNS内炎性免疫应答,外周血穿孔素和颗粒酶-B表达阳性的CD8+TEM数量可在一定程度上反映MS患者CNS的病损程度.