Expansion of CD4+CD28- T cells producing high levels of interferon-{gamma} in peripheral blood of patients with multiple sclerosis

CD4(+) T cells that lack surface expression of the CD28 co-stimulatory molecule (CD4(+)CD28(-) T cells) were expanded in peripheral blood of patients with multiple sclerosis (MS) [5.20 +/- 1.67% vs 13.00 +/- 2.68% (healthy controls (HC) versus patients with MS)]. Both the CD4(+)CD28(+) and CD4(+)CD28(-) T-cell populations of patients with MS produced higher levels of interferon (IFN)-gamma compared with those in HC. In particular, the proportion of IFN-gamma(+) cells among CD4(+)CD28(-) T cells from patients with MS was considerably high. However, expression of co-stimulatory molecules including inducible costimulator (ICOS), activating natural killer receptors, or members of tumor necrosis factor receptor family that replace CD28 in CD4(+)CD28(-) T cells of patients with MS could not be identified. A unique subpopulation bearing the CD45RA(high)CCR7(-) phenotype was identified among the CD4(+)CD28(-) T cells of some patients with MS. Because only MS samples contained this CD45RA(high)CCR7(-) population attributed to terminally differentiated effector memory cells and lacked naive CD45RA(high)CCR7(+) cells, we suggest that CD4(+)CD28(-) T cells of patients with MS represent a cell population which is in more differentiated state than healthy subjects. In patients treated with IFN-beta-1b, IFN-gamma production from CD4(+)CD28(+) T cells was suppressed compared with that in untreated patients. On the contrary, in the CD4(+)CD28(-) population, production of IFN-gamma in IFN-beta-1b-treated patients was not significantly suppressed compared with that in untreated patients with MS. Thus, an additional treatment strategy that specifically targets this cell population may enhance the beneficial effect of IFN-beta on MS.     

Expression of the B7-related molecule ICOSL by human glioma cells in vitro and in vivo

Human glioblastoma is a highly lethal tumor known for its capability of interfering with effective antitumor immune responses. Costimulatory signals are of critical relevance in both the inductive and effector phases of immune responses. Inducible costimulator-ligand (ICOSL), a member of the B7 family of costimulatory molecules related to CD80/CD86, regulates CD4 as well as CD8 T-cell responses via interaction with its receptor, ICOS, on activated T cells. We report the expression of ICOSL by glioma cells in vitro and in vivo. In contrast to CD80 (B7.1) and CD86 (B7.2), ICOSL protein and mRNA was expressed in 7 of 12 glioma cell lines. ICOSL expression is upregulated by the inflammatory cytokine, tumor necrosis factor-alpha (TNF-alpha), whereas interferon-gamma (IFN-gamma) has no such effect. Further, immunohistochemical analysis of human brain tumors demonstrates the expression of ICOSL in three of four tissue samples. ICOSL expression is functional in that a neutralizing ICOSL antibody (HIL-131) reduces Th1 and Th2 cytokine levels in cocultures of peripheral blood lymphocytes or T-cell subsets (CD4 and CD8) with glioma cells. However, ICOSL gene transfer into glioma cells does not alter their immunogenicity under primary or secondary alloreactive coculture assays.     

Beta blocker and angiotensin-converting enzyme inhibitor therapy is associated with decreased Th1/Th2 cytokine ratios and inflammatory cytokine production in patients with chronic heart failure

OBJECTIVE: To examine the potential impact of beta-blockers and angiotensin-converting enzyme (ACE) inhibitors, medications which modulate beta-adrenergic signaling, on immune function in patients with chronic heart failure (HF). METHODS: 118 patients attending an HF center were tested for circulating levels of norepinephrine (NE), T cells and the inflammation-associated cytokine interleukin 6 (IL-6). Levels of the cytokines interferon-gamma (IFNgamma), IL-10, and tumor necrosis factor-alpha (TNFalpha) produced by cultured peripheral blood mononuclear cells (PBMC) were measured in culture supernatants following T cell stimulation in vitro. RESULTS: NE levels were significantly lower in patients receiving ACE inhibitors (p = 0.0263), with a trend toward lower NE in patients receiving beta-blockers. All patients exhibited relatively normal levels of T cells, and there was a trend toward higher levels of total (CD3+) and helper (CD4+) T cells (p = 0.0578 and 0.0932, respectively) in patients receiving either type of medication. The ratios of Th1 (IFNgamma) to Th2 (IL-10) cytokines were lower in patients receiving a combination of beta-blocker and ACE inhibitor therapy (p = 0.0373). NYHA class was a significant predictor of serum IL-6 (p < 0.0001). There was a trend toward lower levels of serum IL-6 in patients receiving both types of medications (p = 0.0606). TNFalpha production by CD3/CD28-stimulated PBMC was significantly lower in patients receiving ACE inhibitor medications (p = 0.0223). CONCLUSIONS: These results suggest that high sympathetic tone associated with chronic HF affects Th1/Th2 and inflammatory cytokine production, and that these effects can be modulated by medications. In addition to improvement in clinical parameters relating to cardiovascular function, beta-blocker and ACE inhibitor medications also appear to have a beneficial effect on the immune system in HF.     

The role of costimulation in autoimmune demyelination

Experimental allergic encephalomyelitis (EAE) is a T cell-mediated, autoimmune disorder characterized by central nervous system (CNS) inflammation and demyelination, features reminiscent of the human disease, multiple sclerosis (MS). In addition to the signal the encephalitogenic T cell receives through the T cell receptor (TCR), a second signal, termed costimulation, is required for complete T cell activation. The B7 family of cell surface molecules expressed on antigen presenting cells (APC) is capable of providing this second signal to T cells via two receptors, CD28 and CTLA-4. Our studies have shown that costimulation provided by B7 molecules to its ligand CD28 is important in the initiation of the autoimmune response in EAE. Further, it appears the costimulation provided by B7-1 is important in disease development, while B7-2 may play an important regulatory role. We and others later showed that B7/CTLA-4 interaction plays a critical role in down-regulating the immune response. Previous work has shown that activated T cells and T cells of a memory phenotype are less dependent on costimulation than naive T cells. T cells reactive with myelin components that are involved in the pathogenesis of EAE and possibly MS would be expected to have been activated as part of the disease process. Building upon our prior work in the EAE model, we have tested the hypothesis that myelin-reactive T cells, which are relevant to the pathogenesis of CNS inflammatory demyelination, can be distinguished from naive myelin-reactive T cells by a lack of dependence upon costimulation for activation and that the costimulatory requirements of these myelin-reactive T cells change during the course of disease. Our studies in the EAE model have also addressed the mechanisms of extrathymic (peripheral) T cell tolerance following intravenous (i.v. ) administration of high dose antigen. It is believed that TCR signaling in the absence of costimulation is a vital component of peripheral tolerance mechanisms. However, recent evidence suggests that peripheral tolerance of antigen-specific T cells induced in vivo may require CTLA-4 engagement of the tolerized T cells. We have begun to examine the molecular mechanisms of tolerance induction following intravenous and intraperitoneal administration of myelin antigens in the EAE model and test the hypothesis that tolerance induction is dependent on the B7:CD28/CTLA-4 pathway. The results from our studies will enhance our understanding of the role that myelin-reactive T cells may play in the pathogenesis of MS. We have determined that MBP-reactive T cells in MS patients are less dependent upon CD28 costimulation than in normal controls, suggesting that these T cells were previously primed in vivo. Characterization of these CD28-independent myelin-specific T cells will have broad implications for a variety of immunologically based therapies in diseases such as MS.     

2q33区段CD28/ICOS基因多态性与中国南方汉族人群多发性硬化疾病易感性及严重程度关系的研究

目的 探讨2q33区段CD28及ICOS基因多态性与多发性硬化(MS)遗传易感性及病情严重程度的关系.方法 以来自中国南方汉族人群的83名确诊MS患者(MS组)和110名非自身免疫性疾病患者及健康志愿者(对照组)为研究对象,外周静脉血提取DNA,利用PCR-RFLP技术、琼脂糖凝胶电泳技术分析检测扩增产物CD28及ICOS基因的多态性.免疫荧光双标记流式细胞学法检测2组患者外周静脉血淋巴细胞表面CD28及ICOS表达水平.结果 MS组ICOS-2394位点TT基因型频率明显高于对照组(33.7%vs10.9%),T等位基因频率明显高于对照组(56.0%vs30.9%),差异有统计学意义(P＜0.05).3个位点单核苷酸多态性间无明显联合效应.ICOS-2394多态位点TT基因型与原发进展型(PP)-MS发病相关,而CT基因型与继发进展型(SP)-MS发病无关.3个位点基因型及等位基因频率与MS急性期患病严重程度均无明显相关(P＜0.05).MS组外周血淋巴细胞表面CD28及ICOS的表达均明显高于对照组,差异有统计学意义(P＜0.05).结论 中国南方汉族人群ICOS-2394位点多态性与MS发病相关,其中TT基因型与PP-MS发病相关,而CT基因型与SP-MS发病无关,提示该基因为MS的易感基因.CD28基因多态性与MS患病无关.

Abstract：

Objective To investigate the polymorphisms of CD28 and COS genes in chromosome 2q33 region and susceptibility to and severity of multiple sclerosis (MS).Methods Eighty-three patients diagnosed as having MS from Han population of South China were studied; one hundred and ten patients with non-autoimmune diseases or healthy volunteers were selected as controls.DNA was obtained from peripheral venous blood; the polymorphisms of amplified productions (CD28 and ICOS genes) were detected by PCR-RFLP and analyzed by agarose gel electrophoresis.Immunofluorescent two-color flow cytometry was used to study the expressions of CD28 and ICOS genes of lymphocytes in the peripheral blood of these 2 groups. Results The genotype frequency of TT at ICOS-2394 site in patients with MS (33.7%) was significantly higher than that in controls (10.9%,P＜ 0.05), and the allele frequency of T in patient group (56.0%) was obviously higher than that in the controls (30.9%, P＜0.05). No marked combined effects were noted in the 3 SNPs (CD28-372, ICOS-2349 and ICOS-2119). TT genotype in the polymorphic site of ICOS-2394 was correlated to primary progressive MS, while CT genotype in the polymorphic site of ICOS-2394 was not correlated to secondary progressive MS. The genotype and allele frequencies of the 3 SNPs had no marked association with the severity of MS (P＞0.05). The levels of CD28 and ICOS gene expressions in lymphocytes of peripheral blood of patients with MS were significantly higher in patients than that in control group (P ＜ 0.05). Conclusion ICOS polymorphisms might be related to MS in Han population of South China,which suggests that ICOS might be one of genes having susceptibility to MS. No association between CD28 gene polymorphisms and susceptibility to MS is noted.     

Bystander modulation of chemokine receptor expression on peripheral blood T lymphocytes mediated by glatiramer therapy

BACKGROUND: Glatiramer acetate therapy is thought to be effective for multiple sclerosis (MS) by promoting T(H)2 cytokine deviation, possibly in the brain, but the exact mechanism and site of action are incompletely understood. Determining the site of action and effect of glatiramer on cell trafficking is of major importance in designing rational combination therapy clinical trials. OBJECTIVE: To determine whether glatiramer therapy will also act in the peripheral blood through bystander modulation of chemokine receptor (CKR) expression and cytokine production on T lymphocytes. DESIGN: Before-and-after trial. SETTING: A university MS specialty center. PATIENTS: Ten patients with relapsing-remitting MS. INTERVENTIONS: Treatment with glatiramer for 12 months and serial phlebotomy. MAIN OUTCOME MEASURES: Cytokine production, CKR expression, and cell migration. RESULTS: The glatiramer-reactive T cells were T(H)2 cytokine biased, consistent with previous studies. We found a significant reduction in the expression of the T(H)1 inflammation associated with the CKRs CXCR3, CXCR6, and CCR5 on glatiramer- and myelin-reactive T cells generated from patients with MS receiving glatiramer therapy vs baseline. Conversely, expression of the lymph node-homing CKR, CCR7, was markedly enhanced on the glatiramer-reactive T cells derived from patients with MS undergoing glatiramer therapy. There was a reduction in the percentage of CD4+ glatiramer-reactive T cells and an increase in the number of CD8+ glatiramer-reactive T cells. CONCLUSIONS: Glatiramer may suppress autoreactive CD4+ effector memory T cells and enhance CD8+ regulatory responses, and bystander modulation of CKRs may occur in the periphery.     

Interleukin-10 and tumor necrosis factor-alpha: model of immunomodulation in multiple sclerosis

OBJECTIVES: The mechanisms involved in the pathogenesis of relapsing-remitting multiple sclerosis are still unclear. The aim of the present study was to evaluate both cerebrospinal fluid (CSF) CD4+ CD7+ T cells and peripheral blood (PB) interleukin-10 (IL-10) as well as tumor necrosis-alpha (TNF-alpha) levels in patients with definite multiple sclerosis of the relapsing-remitting type.METHODS: To assess the above-mentioned cytokine levels we performed our test by the means of ELI-spot assay; the T-helper cell subset was assayed using flow cytometry.RESULTS: PB IL-10 levels of multiple sclerosis (MS) patients in remission were significantly (p<0.001) higher than in MS patients in the active phase. There was significant and increased evidence of TNF-alpha levels only in the MS patients in the active phase. CD4+ CD7+ T cells, characterized by a preferential Th1-like cytokine profile, were detectable only in seven patients in the active phase without evidence of a statistical significance with respect to cytokine levels.CONCLUSION: The data indicate that the production of different cytokines characterized the expression of relapsing-remitting MS. The data also suggest that is it possible to control MS using the regulatory cytokine balance.     

Cytokine secretion profile of myelin basic protein-specific T cells in multiple sclerosis

Multiple sclerosis (MS) is a demyelinating disease of the central nervous system with a presumed autoimmune pathogenesis involving autoantigen-specific CD4+ T cells and cytokines. A similar frequency of T cells responding to myelin basic protein (MBP), a putative target in MS, has been observed in MS patients and controls. To dissect the differences between MBP-specific T cells in patients and controls, we have analyzed the cytokine secretion profile of such autoreactive T cells. MBP-specific T cell clones (TCC) were isolated from the peripheral blood of MS patients and controls by limiting dilution. Expression of mRNA for interferon-gamma (IFN-gamma), interleukin (IL)-4, IL-10, tumor necrosis factor-alpha (TNF-alpha) and transforming growth factor-beta (TGF-beta) was assessed by polymerase chain reaction whereas secretion of cytokine protein was measured by ELISA. MBP-specific TCC exhibited a heterogeneous cytokine secretion profile with clones displaying Th1, Th2 and Th0 phenotypes. A significant difference in the distribution of the cytokine profile was noted between MS patients and controls. Although the frequency of Th1 secreting MBP-reactive TCC was similar between MS patients and controls, stable MS patients had a significant association with the Th0 phenotype whereas healthy individuals were associated with the Th2 phenotype. In comparison to control TCC, MBP-specific TCC from MS patients secreted increased amounts of IFN-gamma, IL-4 and IL-10 and decreased quantities of TGF-beta. Thus, these studies suggest that there is a dysregulation in the balance between pro-inflammatory Th1 and anti-inflammatory Th2 cytokines in MS. It appears that the presence of Th1 secreting autoreactive T cells in healthy individuals may be counterbalanced by the presence of cells secreting Th2 cytokines and by the augmented production of the immunosuppressive cytokine TGF-beta, whereas in MS there is a decrease in these anti-inflammatory agents.     

Flow cytometric differentiation of Asian and Western types of multiple sclerosis, HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) and hyperIgEaemic myelitis by analyses of memory CD4 positive T cell subsets and NK cell subsets

We examined the alterations of memory CD4(+) T cell subsets bearing surface receptors linked to either Th1 or Th2 cytokine production as well as natural killer (NK) cell subsets by three color flow cytometry in the peripheral blood from 36 patients with clinically definite multiple sclerosis (MS), 27 patients with HAM/TSP, 13 patients with hyperIgEaemic myelitis who had mite antigen-specific IgE and 25 healthy controls (HC). The patients with MS were clinically classified into an optico-spinal form of MS (Asian type, MS-A) and the conventional form of MS (Western type, MS-W). MS-A showed a significant increase of CD4(+)CD45RA(-)CCR5(+) cells (Th1 cells) through the relapse and remission phases in comparison to HC, while MS-W showed a significant increase of CD4(+)CD45RO(+)CD62L(-) cells (Th1 cells) only at the relapse phase. HAM/TSP showed a significant increase of CCR5(+) and CD62L(-) memory CD4(+) T cells as well as CD30(+) memory CD4(+) T cells (Th2 cells) in comparison to HC. On the other hand, a selective increase of CD4(+)CD45RO(+)CD30(+) cells was found in hyperIgEaemic myelitis. The percentage of mature NK cells (CD3(-)CD16(+)CD56(+) cells) as well as double negative T cells (CD3(+)CD4(-)CD8(-) cells) decreased significantly in HAM/TSP in comparison to HC. Our findings therefore suggest a flow cytometric analysis of Th1/Th2-associated markers on memory CD4(+) T cells as well as NK cell subsets to be useful for differentiating various inflammatory neurologic conditions.     

多发性硬化患者外周血CD4+CD25+ T细胞变化及其机制探讨

目的探讨多发性硬化(MS)患者外周血CD4 CD25 T细胞数量及叉头样转录因子(FOXP3)表达水平与MS病情的关系。方法选择温州地区MS患者44例(男12例、女32例),均按Poser诊断标准诊断,结合头颅MRI增强扫描排除合并其他神经系统和免疫系统疾病,并统一行EDSS评分;对照组43例(男13例、女30例)为健康查体者。具体方法:流式细胞仪检测外周血CD4 CD25 T细胞数量;免疫磁珠法分离CD4 CD25 T细胞;RT-PCR法检测CD4 CD25 T细胞FOXP3 mRNA表达并进行半定量分析。结果MS患者外周血中CD4 CD25 调节性T细胞数量与对照组比较无明显变化(P>0.05);活化的效应性T细胞数量增加(P<0.05)且活动期增加更为显著(P<0.01)。同一个体疾病活动期外周血CD4 CD25 调节性T细胞数量较非活动期减少(P<0.05)。MS患者外周血中CD4 CD25 T细胞的FOXP3 mRNA表达降低(P<0.05),且活动期降低更明显(P<0.01)。结论此组MS患者外周血CD4 CD25 调节性T细胞抑制活性降低,FOXP3 mRNA表达减少,活化的效应性T细胞数量增加,且与MS疾病活动性有关。     

Glatiramer acetate and IFN-beta act on dendritic cells in multiple sclerosis.

Glatiramer acetate (GA; Copolymer 1; Copaxone) and interferon-beta (IFN-beta) modulate the course of multiple sclerosis (MS), but probably by different mechanisms. GA, a mixture of synthetic peptides, is believed to act as an altered peptide ligand with inhibitory effects on autoreactive T cells and promoting Th2 cells. It is unknown whether GA affects dendritic cells (DCs), which, besides strong antigen presenting capacity, orchestrate Th1 and Th2 responses. IFN-beta inhibits IL-12 production by DCs over unknown mechanisms. This study was designed to investigate in vitro effects of GA and IFN-beta on the development and function of DCs from MS patients and healthy controls, and to explore their possible synergistic effects on DCs. DCs were generated from adherent blood mononuclear cells (MNCs). GA or IFN-beta or both, when added at initiation of DC cultures, rapidly promoted the development of adherent MNCs into floating, activated DCs as reflected by up-regulation of HLA-DR and CD86 expression. IFN-beta, but not GA, induced IL-3R expression on DCs. Compared to DCs from healthy controls, MS patients' DCs expressed higher levels of the myeloid DC marker CD1a and produced lower amounts of IL-10. GA reduced IL-12 production by DCs. IFN-beta also reduced IL-12, but increased IL-10 production by DCs from both MS patients and healthy controls. GA and IFN-beta synergistically suppressed CD1a and enhanced CD86 expression on MS DCs. These findings document novel mechanisms of action of GA and IFN-beta at the DC level in MS.     

Expression of costimulatory molecules on peripheral blood mononuclear cells in multiple sclerosis

OBJECTIVE: To determine whether there was aberrant expression of costimulatory molecules and their receptors on peripheral blood mononuclear cells of MS patients and healthy individuals and whether expression correlated with disease status. METHODS: Forty-six patients with MS and 29 healthy individuals were analyzed by direct 2-color immunofluorescence flow cytometry for expression of CD80, CD86, CD28 and CTLA-4 on T and B cells and monocytes. RESULTS: Expression of CD80 on CD4+ T cells was upregulated in progressing MS patients compared to stable MS patients and controls. Marked increase in the expression of CD80 and CD86 was seen on both CD4+ and CD8+ T cells from an MS patient with rapidly progressing disease. No difference in the expression of the costimulatory molecules or their ligands was seen between IFN-beta treated and non-treated MS patients. CONCLUSIONS: Aberrant expression of costimulatory molecules occurs on T lymphocytes in MS patients with progressing disease.     

Effects of sex hormones on costimulatory molecule expression in multiple sclerosis

Sex hormones play a central role as modulators of immune responses and autoimmune diseases. We hypothesized that suppression of MS disease during pregnancy may be mediated by sex steroid hormones via regulation of costimulatory molecules such as CD40L or CD80/CD86 (B7-1/B7-2). We tested two sex hormones that are implicated in immune suppression during pregnancy: estriol and progesterone. We also examined whether this regulation is gender-specific or disease-related. PBMC from untreated relapsing remitting multiple sclerosis (RR MS) patients and controls were examined for expression of T cell and monocyte costimulatory molecules following mitogen stimulation in the presence or absence of sex hormones. In the absence of hormones, we confirmed that mitogen stimulation induced significantly more CD40L on the surface of CD4(+)T cells in MS patients compared to controls, and we extend these findings by showing there were no gender differences in induction of CD40L. Although supra-physiologic doses of hormones mildly suppressed CD40L expression on activated T cells, in vitro exposure to typical pregnancy-related physiologic doses of estriol or progesterone showed very little or no suppression of CD40L. On monocytes, neither estriol nor progesterone significantly altered the expression of CD80/CD86. These results suggest that physiologic doses of estriol or progesterone cannot alter CD40L on T cells or CD80/CD86 on monocytes sufficiently to explain the improvement observed in MS during pregnancy. Thus, although amelioration of MS and other autoimmune diseases during pregnancy is thought to be due to increased sex hormones, the present results do not support a role for suppression of costimulation via estriol or progesterone.     

Monocyte-derived HLA-G acts as a strong inhibitor of autologous CD4 T cell activation and is upregulated by interferon-beta in vitro and in vivo: rationale for the therapy of multiple sclerosis

Peripheral antigen presenting cells (APCs) contribute to the maintenance of immune tolerance and are considered to play a critical role in promoting the (re)activation of autoreactive T cells in multiple sclerosis (MS). Interferon-beta (IFN-beta) is the principle immune-modulatory agent used in the treatment of MS, but its mechanism of action remains elusive. HLA-G is a non-classical MHC molecule (MHC class Ib) attributed chiefly immune-regulatory functions. We here investigated the role of monocyte-derived HLA-G in the immune-regulatory processes of MS and its implications for current immune-modulatory therapies. Monocytes constitutively express cell surface HLA-G1 and soluble HLA-G5. Comparison of monocytic HLA-G expression between patients with relapsing-remitting MS (n=17) and healthy donors (n=20) revealed significantly lower levels of HLA-G1 protein in MS patients. However, both groups showed a significant upregulation of HLA-G in response to IFN-beta in vitro. Serial measurements of HLA-G mRNA levels in MS patients before and during IFN-beta therapy corroborated the relevance of these results in vivo: 1 month after initiation of IFN-beta1b therapy (n=9), HLA-G1 and HLA-G5 were significantly increased compared to baseline levels and remained elevated during treatment for 6 months (n=3). Importantly, functional experiments demonstrated that monocyte-derived HLA-G inhibits both Th1 (IFN-gamma, IL-2) and Th2 (IL-10) cytokine production by antigen-stimulated autologous CD4 T cells. Soluble HLA-G added to antigen-specific T cell lines (TCLs) has similar effects on the release of cytokines and reduces T cell proliferation. Although both IFN-beta and IFN-gamma strongly enhance HLA-G1 and HLA-G5 expression by monocytes in vitro, IFN-beta leads to a stronger relative upregulation of HLA-G compared to classical MHC class I molecules than stimulation with IFN-gamma. Taken together, monocyte-derived HLA-G mediates the inhibition of autologous CD4 T cell activation and might be involved in immune-regulatory pathways in the pathogenesis of MS. We conclude that some desirable immune-modulatory effects of INF-beta might be accomplished via the upregulation of the immune-tolerogenic molecule HLA-G.     

Analysis of CD27 surface expression on T cell subsets in MS patients and control individuals

Within the peripheral blood, CD4⁺CD27⁻ T cells only reside within the CD45RAT ⁻(memory or primed) T cell subset. Cells with this phenotype have characteristics of specialized effector T cells according to their cytokine secretion profiles and the expression of tissue-specific adhesion molecules. This subset was previously found to be increased in certain diseases that are associated with immune activation. Therefore we analyzed CD27 expression of peripheral blood and CSF T cells in MS patients. Within the CD4⁺ T cell subset no differences were seen between MS patients and controls in proportions of CD45RA⁻CD27⁻ cells. However, when the CD3⁺ T cell compartment was analyzed, CD27⁻ cells were also found within the CD45RA⁺ subset. These cells, most likely CD8⁺, are significantly reduced in PBL and CSF of MS patients as compared with OND patients. In MS and OND groups the level of CD27⁻ cells in peripheral blood correlated significantly with that in CSF, indicating a balanced migration of CD27⁻ cells between the two compartments. In OIND patients, however, this equilibrium was lost. The correlation of the level of CD27⁺ cells with the amount of intrathecally produced IgG in MS patients may suggest that CD27⁺ cells are responsible for B cell help in this disease.     

Membrane bound IL-15 is increased on CD14 monocytes in early stages of MS

IL-15 is a pro-inflammatory cytokine whose three-dimensional structure is similar to that of IL-2. IL-2 and IL-15 have similar as well as distinct biological functions. An active form of IL-15 that is membrane bound has also been described. Furthermore, IL-15 is known to play a role in autoimmune diseases. We thus investigated the expression of membrane bound IL-15 on monocytes (CD14+ cells) and studied its effect on T cell activation in MS patients. We found that unstimulated CD14+ cells from relapsing remitting MS patients had increased membrane bound IL-15. Those with high surface levels of IL-15 on monocytes were in the early stages of the disease. In addition, we found that T cells of MS patients had enhanced responsiveness to IL-15 and there was increased expression of IL-15 receptor on CD4+ T cells. Thus, IL-15 may be an important cytokine that drives Th1 responses early in the course of the disease and could serve as a target for immunotherapy and as an early marker in the immunologic staging of MS.     

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.     

Cytokine production by naive and primary effector CD4+ T cells exposed to norepinephrine

We recently showed that clones of Th1 cells, but not Th2 cells, expressed a functional beta-2-adrenergic receptor (beta2AR) and that either norepinephrine or the beta2AR agonist terbutaline stimulated this receptor to modulate the level of Th1 cytokines produced. In the present study, we show that norepinephrine and terbutaline stimulate the beta2AR to decrease the level of IL-2 produced by freshly isolated murine splenic naive CD4+ T cells from either Balb/C or DO11.10 transgenic mice and activated polyclonally with anti-CD3 and anti-CD28 mAbs. In contrast, the level of cytokines produced by primary effector Th1 and Th2 cells were unaffected when norepinephrine, terbutaline, or cAMP analogs were added at the time of restimulation. These results suggest that a diversity exists among CD4+ T-cell subsets with respect to the level of adrenergic receptor expression, responsiveness to cAMP, stage of cell differentiation, or a combination of the above.     

Altered neuroantigen-specific cytokine secretion in a Th2 environment reduces experimental autoimmune encephalomyelitis

Activation of Th2 cells suppresses clinical experimental autoimmune encephalitis (EAE), demyelination and expression of genes associated with Th1-mediated inflammation. Despite both reduced central nervous system inflammation and IFN-gamma induced MHC class II expression by microglia, the composition of CNS infiltrates in Th2-protected mice were similar to mice with EAE. Analysis of the CNS infiltrating cells by flow cytometry suggests that protection did not correlate with abrogation of CD4+ T cell recruitment, preferential recruitment of donor Th2 cells or an increased frequency of CD25+ CD4+ T cells. By contrast, protection correlated with an increased frequency of neuroantigen-specific Th2 cells infiltrating the CNS. These data suggest that a peripheral Th2 cytokine environment influences both potential antigen presenting cells as well as recruitment and/or retention of neuroAg-specific Th2 CD4+ T cells.