CD226 expression deficiency causes high sensitivity to apoptosis in NK T cells from patients with systemic lupus erythematosus

Humans and mice with systemic lupus erythematosus (SLE) and related autoimmune diseases have reduced numbers of NK T cells. An association between NK T cell deficiency and autoimmune disease has been identified. However, the mechanisms for reduction of NK T cell number in patients with SLE are unknown. In the present study we report that NK T cells from active SLE patients are highly sensitive to anti-CD95-induced apoptosis compared with those from normal subjects and inactive SLE patients. CD226 expression is deficient on NK T cells from active SLE patients. The expression of one antiapoptotic member protein, survivin, is found to be selectively deficient in freshly isolated NK T cells from active SLE patients. CD226 preactivation significantly up-regulates survivin expression and activation, which can rescue active SLE NK T cells from anti-CD95-induced apoptosis. In transfected COS7 cells, we confirm that anti-CD95-mediated death signals are inhibited by activation of the CD226 pathway through stabilization of caspase-8 and caspase-3 and through activation of survivin. We therefore conclude that deficient expression of CD226 and survivin in NK T cells from active SLE is a molecular base of high sensitivity of the cells to anti-CD95-induced apoptosis. These observations offer a potential explanation for high apoptotic sensitivity of NK T cells from active SLE, and provide a new insight into the mechanism of reduction of NK T cell number in SLE and understanding the association between NK T cell deficiency and autoimmune diseases.     

Hypergravity-induced immunomodulation in a rodent model: lymphocytes and lymphoid organs

The major goal of this study was to quantify changes in lymphoid organs and cells over time due to centrifugation-induced hypergravity. C57BL/6 mice were exposed to 1, 2 and 3 G and the following assays were performed on days 1, 4, 7, 10, and 21: spleen, thymus, lung, and liver masses; total leukocyte, lymphocyte, monocyte/macrophage, and granulocyte counts; level of splenocyte apoptosis; enumeration of CD3+ T, CD3+/CD4+ T helper, CD3+/CD8+ T cytotoxic, B220+ B, and NK1.1+ natural killer cells; and quantification of cells expressing CD25, CD69, and CD71 activation markers. The data show that increased gravity resulted in decreased body, spleen, thymus, and liver, but not lung, mass. Significant reductions were noted in all three major leukocyte populations (lymphocytes, granulocytes, monocyte/macrophages) [correction of macrphages] with increased gravity; persistent depletion was noted in blood but not spleen. Among the various lymphocyte populations, the CD3+/CD8+ T cells and B220+ B cells were the most affected and NK1.1+ NK cells the least affected. Overall, the changes were most evident during the first week, with a greater influence noted for cells in the spleen. A linear relationship was found between some of the measurements and the level of gravity, especially on day 4. These findings indicate that hypergravity profoundly alters leukocyte number and distribution in a mammalian model and that some aberrations persisted throughout the three weeks of the study. In certain cases, the detected changes were similar to those observed after whole-body irradiation. In future investigations we hope to combine hypergravity with low-dose rate irradiation and immune challenge.     

Circulating TCR gammadelta cells in the patients with systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is a disorder with a wide range of immunological abnormalities. The results of the studies undertaken in the last decade indicated that SLE pathogenesis was mainly connected with the breakdown of the activation control of B and T cells, generating humoral or cell-mediated responses against several self-antigens of affected cells. The last studies demonstrate that the role of gammadelta T lymphocytes in autoimmune diseases can be especially important. Flow cytometry techniques were used to investigate the number and percentage of TCR gammadelta T cells and their most frequent subtypes in peripheral blood of 32 patients with SLE and 16 healthy volunteers. We also correlated TCR gammadelta cells number with the level of T CD3+, T CD4+, T CD8+, and NK (CD16) cells (cytometric measurements) and SLE activity (on the basis of clinical investigations). Our studies were preliminary attempts to evaluate the role of that minor T cell subpopulation in SLE. Absolute numbers of cells expressing gammadelta TCR in most SLE blood specimens were significantly lower than in the control group (P<0.006). However, since the level of total T cell population was also decreased in the case of SLE, the mean values of the percentage gammadelta T cells of pan T lymphocytes were almost the same in both analysed populations (7.1% vs 6.3%, respectively). In contrast to Vdelta2+ and Vgamma9+ subtypes of pan gammadelta T cells, Vdelta3+ T cells number was higher in SLE patients (20 x 10 cells/microl) than in healthy control group (2 x 2 cells/microl) (P=0.001). However, we found no differences between the numbers of pan gammadelta T lymphocytes and studied their subtypes in the patients with active and inactive disease. These cell subpopulations were doubled in the treated patients with immunosuppressive agents in comparison with untreated ones; however, data were not statistically significant. Our study indicated that Vdelta3+ subtype of gammadelta T cells seems to be involved in SLE pathogenesis; however, we accept the idea that the autoimmunity does not develop from a single abnormality, but rather from a number of different events.     

Reciprocal regulation between natural killer cells and autoreactive T cells

The initiation and the progression of autoimmune diseases stem from complex interactions that involve cells of both the innate and the adaptive immune system. As we discuss here, natural killer (NK) cells, which are components of the innate immune system, can inhibit or promote the activation of autoreactive T cells during the initiation of autoimmunity. After they have been activated, autoreactive T cells contribute to the homeostatic contraction of NK-cell populations. The dynamic interaction between NK cells and autoreactive T cells might indicate the transition from the innate immune triggering of autoimmunity to the progressive phase of the disease. Understanding the mechanisms and signals that control the reciprocal regulation of NK cells and autoreactive T cells could have important implications for treatment in the clinic.     

A defect in the suppressor circuits among OKT4+ cell populations in patients with systemic lupus erythematosus occurs independently of a defect in the OKT8+ suppressor T cell function

The autologous mixed lymphocyte reaction (MLR) is thought to be part of a regulatory role of T cells on B cell function. OKT4+, but not OKT8+, cells can proliferate in response to autologous non-T cells. Moreover, the OKT4+ cell population activated early in the course of autologous MLR functioned as inducer cells for the differentiation of B cells, whereas later in the response, the activated OKT4+ cells were particularly enriched in suppressor cells. A part of the autologous MLR appears to be an important pathway for the activation of feedback suppression mechanisms among cells contained within the OKT4+ populations. Patients with systemic lupus erythematosus (SLE) were studied with regard to the following OKT4+ cell functions in vitro after activation in the autologous MLR: a) proliferative response, and b) helper and suppressor activities for differentiation of B cells. A marked reduction in the proliferative response of OKT4+ cells was observed in SLE patients. SLE OKT4+ cells activated in the autologous MLR could function as helper cells but could not exert any suppressor activity. This OKT4+ cell abnormality was present regardless of the disease activity, and occurred in the absence of autoantibodies including anti-T cell antibodies. Instead, SLE anti-T cell antibodies could preferentially eliminate cells bearing the OKT8+ phenotype characteristic of suppressor cells in populations of normal T cells. These results suggest that the defect in the suppressor circuits among OKT4+ cell populations is intrinsic to SLE lymphocytes and that the OKT8+ suppressor T cell defect is caused by antibodies produced by the B cells of SLE patients.     

Effect of medical castration on CD4+ CD25+ T cells, CD8+ T cell IFN-gamma expression, and NK cells: a physiological role for testosterone and/or its metabolites

The higher prevalence of autoimmune disease among women compared with men suggests that steroids impact immune regulation. To investigate how sex steroids modulate cellular immune function, we conducted a randomized trial in 12 healthy men aged 35-55 yr treated for 28 days with placebo, a GnRH antagonist, acyline to induce medical castration, or acyline plus daily testosterone (T) gel to replace serum T, followed by a 28-day recovery period. Serum hormones were measured weekly and peripheral blood lymphocytes (PBLs) were collected biweekly for analyses of thymus-derived lymphocyte (T cell) subtypes and natural killer (NK) cells. Compared with the other groups and to baseline throughout the drug exposure period, men receiving acyline alone had significant reductions in serum T (near or below castrate levels), dihydrotestosterone, and estradiol (P < 0.05). Medical castration significantly reduced the percentage of CD4+ CD25+ T cells (P < 0.05), decreased mitogen-induced CD8+ T cell IFN-gamma expression, and increased the percentage of NK cells without affecting the ratio of CD4+ to CD8+ T cells and the expression of NK cell-activating receptor NKG2D or homing receptor CXCR1. No changes in immune composition were observed in subjects receiving placebo or acyline with replacement T. These data suggest that T and/or its metabolites may help maintain the physiological balance of autoimmunity and protective immunity by preserving the number of regulatory T cells and the activation of CD8+ T cells. In addition, sex steroids suppress NK cell proliferation. This study supports a complex physiological role for T and/or its metabolites in immune regulation.     

Suppressive effect of human natural killer cells on pokeweed mitogen-induced B cell differentiation

The suppressive effect of human natural killer (NK) cells on B cell differentiation induced by pokeweed mitogen (PWM) was investigated. By using Percoll discontinuous density gradient centrifugation, peripheral blood nonphagocytic and nonadherent mononuclear cells were divided into low and high density fractions for which NK cells (Large granular lymphocytes, LGL) and T cells were enriched, respectively. These fractionated mononuclear cells were co-cultured with purified autologous B cells in the presence of PWM, and were examined for their helper and suppressor activities on differentiation of B cells to immunoglobulin-(IgM and IgG) producing cells by a highly sensitive reversed hemolytic plaque assay. The T cell-enriched high density fractions provided help for B cell differentiation to levels higher than that of unfractionated mononuclear cells. On the other hand, the NK-enriched low density fractions did not show helper activity, and when added to the culture of B cells plus helper T cells, they markedly suppressed B cell differentiation. This suppressive activity, as well as the NK cytotoxicity of the NK-enriched fractions, was abrogated by treatment of the cells with monoclonal antibody against human NK cells (HNK-1), but not against T cells (OKT3) in the presence of complement. NK cells also suppressed PWM-driven B cell differentiation in the presence of T4+ (helper/inducer T) but not T8+ (cytotoxic/suppressor T) cells; however, they showed no inhibition of soluble factor-induced B cell differentiation assayed in the absence of helper T cells. It is thus concluded that human peripheral blood NK cells exhibit an ability to suppress PWM-driven B cell differentiation, possibly by acting through the effect on helper T cells but not directly on B cells.     

Immunoregulatory role of CD1d in the hydrocarbon oil-induced model of lupus nephritis

Systemic lupus erythematosus (SLE) is a systemic autoimmune disease that is accompanied by the emergence of autoreactive T cells and a reduction in regulatory T cells. Humans and mice with SLE have reduced numbers of CD1d-restricted NK T cells, suggesting a role for these cells in the regulation of SLE. In this study, we show that CD1d deficiency exacerbates lupus nephritis induced by the hydrocarbon oil pristane. This exacerbation in disease is associated with: 1) reduced TNF-alpha and IL-4 production by T cells, especially during the disease induction phase; and 2) expansion of marginal zone B cells. Strikingly, inoculation of pristane in wild-type mice resulted in reduced numbers and/or functions of NK T cells and CD1d-expressing dendritic cells. These findings suggest that CD1d may play an immunoregulatory role in the development of lupus in the pristane-induced model.     

Evaluation of the circulating and splenic lymphocyte subpopulations in patients with non-Hodgkin lymphomas and Hodgkin's disease using monoclonal antibodies

The number of B lymphocytes, T lymphocytes and their helper/inducer, cytotoxic/suppressor and NK/K subpopulations was measured in peripheral blood and spleen cell suspensions from patients with Hodgkin's disease (HD) in the active stage of the disease and in remission status, as well as in Non-Hodgkin lymphomas (NHL) in active stage of the disease. B lymphocytes were determined by direct immunofluorescence and T lymphocytes with the E rosette technique. Helper/inducer, cytotoxic/suppressor, and NK/K T lymphocytes were determined by indirect immunofluorescence with the monoclonal antibodies OKT4, OKT8 and Leu 7 (HNK1). In the same way, Lyt3 was used for determination of the total T lymphocytes. Whereas in peripheral blood of the NHL group an increase of B lymphocytes and a slight reduction of T lymphocytes could be observed, with normal distribution of the subpopulations, in patients with active HD as well as in those in remission, a marked absolute and relative decrease of T helper/inducer cells was found with normal cytotoxic/suppressor and NK/K proportion. In contrast to this, a significant increase of helper/inducer T lymphocytes with decreased cytotoxic/suppressor T proportion was found in spleen cell suspensions of patients with HD.     

Deficient CD4+CD25high T regulatory cell function in patients with active systemic lupus erythematosus

CD4(+)CD25(+) T regulatory cells (Tregs) play an essential role in maintaining immunologic homeostasis and preventing autoimmunity. Systemic lupus erythematosus (SLE) is a systemic autoimmune disease characterized by a loss of tolerance to nuclear components. We hypothesized that altered function of CD4(+)CD25(high) Tregs might play a role in the breakdown of immunologic self-tolerance in patients with SLE. In this study, we report a significant decrease in the suppressive function of CD4(+)CD25(high) Tregs from peripheral blood of patients with active SLE as compared with normal donors and patients with inactive SLE. Notably, CD4(+)CD25(high) Tregs isolated from patients with active SLE expressed reduced levels of FoxP3 mRNA and protein and poorly suppressed the proliferation and cytokine secretion of CD4(+) effector T cells in vitro. In contrast, the expression of FoxP3 mRNA and protein and in vitro suppression of the proliferation of CD4(+) effector T cells by Tregs isolated from inactive SLE patients, was comparable to that of normal individuals. In vitro activation of CD4(+)CD25(high) Tregs from patients with active SLE increased FoxP3 mRNA and protein expression and restored their suppressive function. These data are the first to demonstrate a reversible defect in CD4(+)CD25(high) Treg function in patients with active SLE, and suggest that strategies to enhance the function of these cells might benefit patients with this autoimmune disease.     

Molecular Characterization of Circulating Plasma Cells in Patients with Active Systemic Lupus Erythematosus

Systemic lupus erythematosus (SLE) is a generalized autoimmune disease characterized by abnormal B cell activation and the occurrence of increased frequencies of circulating plasma cells (PC). The molecular characteristics and nature of circulating PC and B cells in SLE have not been completely characterized. Microarray analysis of gene expression was used to characterize circulating PC in subjects with active SLE. Flow cytometry was used to sort PC and comparator B cell populations from active SLE blood, normal blood and normal tonsil. The gene expression profiles of the sorted B cell populations were then compared.SLE PC exhibited a similar gene expression signature as tonsil PC. The differences in gene expression between SLE PC and normal tonsil PC and tonsil plasmablasts (PB) suggest a mature Ig secreting cell phenotype in the former population. Despite this, SLE PC differed in expression of about half the genes from previously published gene expression profiles of normal bone marrow PC, indicating that these cells had not achieved a fully mature status. Abnormal expression of several genes, including CXCR4 and S1P₁, suggests a mechanism for the persistence of SLE PC in the circulation. All SLE B cell populations revealed an interferon (IFN) gene signature previously only reported in unseparated SLE peripheral blood mononuclear cells. These data indicate that SLE PC are a unique population of Ig secreting cells with a gene expression profile indicative of a mature, but not fully differentiated phenotype.     

Differential expression of NK T cell V alpha 24J alpha Q invariant TCR chain in the lesions of multiple sclerosis and chronic inflammatory demyelinating polyneuropathy

Human V alpha 24+ NK T cells are a unique subset of lymphocytes expressing the V alpha 24J alpha Q invariant TCR chain. Because they can rapidly produce large amounts of regulatory cytokines, a reduction of NK T cells may lead to the development of certain autoimmune diseases. Using a single-strand conformation polymorphism method, we demonstrate that a great reduction of V alpha 24J alpha Q NK T cells in the peripheral blood is an immunological hallmark of multiple sclerosis, whereas it is not appreciable in other autoimmune/inflammatory diseases such as chronic inflammatory demyelinating polyneuropathy. The chronic inflammatory demyelinating polyneuropathy lesions were often found to be infiltrated with V alpha 24J alpha Q NK T cells, but multiple sclerosis lesions only rarely expressed the V alpha 24J alpha Q TCR. It is therefore possible that the extent of NK T cell alteration may be a critical factor which would define the clinical and pathological features of autoimmune disease. Although the mechanism underlying the NK T cell deletion remains largely unclear, a remarkable contrast between the CNS and peripheral nervous system diseases allows us to speculate a role of tissue-specific elements such as the level of CD1d expression or differences in the CD1d-bound glycolipid.     

Expression and function of inducible co-stimulator in patients with systemic lupus erythematosus: possible involvement in excessive interferon-gamma and anti-double-stranded DNA antibody production

Inducible co-stimulator (ICOS) is the third member of the CD28/cytotoxic T-lymphocyte associated antigen-4 family and is involved in the proliferation and activation of T cells. A detailed functional analysis of ICOS on peripheral blood T cells from patients with systemic lupus erythematosus (SLE) has not yet been reported. In the present study we developed a fully human anti-human ICOS mAb (JTA009) with high avidity and investigated the immunopathological roles of ICOS in SLE. JTA009 exhibited higher avidity for ICOS than a previously reported mAb, namely SA12. Using JTA009, ICOS was detected in a substantial proportion of unstimulated peripheral blood T cells from both normal control individuals and patients with SLE. In CD4+CD45RO+ T cells from peripheral blood, the percentage of ICOS+ cells and mean fluorescence intensity with JTA009 were significantly higher in active SLE than in inactive SLE or in normal control individuals. JTA009 co-stimulated peripheral blood T cells in the presence of suboptimal concentrations of anti-CD3 mAb. Median values of [3H]thymidine incorporation were higher in SLE T cells with ICOS co-stimulation than in normal T cells, and the difference between inactive SLE patients and normal control individuals achieved statistical significance. ICOS co-stimulation significantly increased the production of IFN-gamma, IL-4 and IL-10 in both SLE and normal T cells. IFN-gamma in the culture supernatants of both active and inactive SLE T cells with ICOS co-stimulation was significantly higher than in normal control T cells. Finally, SLE T cells with ICOS co-stimulation selectively and significantly enhanced the production of IgG anti-double-stranded DNA antibodies by autologous B cells. These findings suggest that ICOS is involved in abnormal T cell activation in SLE, and that blockade of the interaction between ICOS and its receptor may have therapeutic value in the treatment of this intractable disease.     

Patterns of peripheral cellular immune disorders in severe rheumatoid arthritis

The study is focused on the correlated peripheral cellular immune disorders registered in a group of 23 patients with severe, progressive rheumatoid arthritis, on methotrexate therapy. We investigated a panel of peripheral immune parameters: leukocyte counts, the proportions of lymphocyte populations (T, Thelper, Tcytotoxic/suppressor, B lymphocytes and NK cells) and the polyclonal activation of lymphocytes. Results show that leukocytosis is due to simultaneously elevated values of monocytes, granulocytes and, to a lesser extent, lymphocytes. The registered high values of the Th to Tc/s ratio are mainly attributed to the abnormal low proportions of the Tc/s subpopulation. Inverse correlations were emphasized between B, Tc/s lymphocytes and NK cells or granulocytes. The unbalance of the lymphocyte to monocyte ratio or of the Th to Tc/s ratio does not impair the polyclonal activation of lymphocytes. In conclusion, we have characterized different patterns of correlated cellular peripheral immune disorders in rheumatoid arthritis, associated to pathological processes in conjunction with the immunsuppressive and anti-inflammatory action of methotrexate that might be relevant for further investigation of disease and further therapy outcome. We emphasize the special relation between the adaptive and innate immune system at the level of cell counts and proportions. The correlations between the peripheral abnormalities in the rheumatoid arthritis group are better highlighted by analyzing subgroups of patients characterized by particular values of the investigated parameters.     

Aberrant phenotype and function of myeloid dendritic cells in systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is characterized by a systemic autoimmune response with profound and diverse T cell changes. Dendritic cells (DCs) are important orchestrators of immune responses and have an important role in the regulation of T cell function. The objective of this study was to determine whether myeloid DCs from individuals with SLE display abnormalities in phenotype and promote abnormal T cell function. Monocyte-derived DCs and freshly isolated peripheral blood myeloid DCs from lupus patients displayed an abnormal phenotype characterized by accelerated differentiation, maturation, and secretion of proinflammatory cytokines. These abnormalities were characterized by higher expression of the DC differentiation marker CD1a, the maturation markers CD86, CD80, and HLA-DR, and the proinflammatory cytokine IL-8. In addition, SLE patients displayed selective down-regulation of the maturation marker CD83 and had abnormal responses to maturation stimuli. These abnormalities have functional relevance, as SLE DCs were able to significantly increase proliferation and activation of allogeneic T cells when compared with control DCs. We conclude that myeloid DCs from SLE patients display significant changes in phenotype which promote aberrant T cell function and could contribute to the pathogenesis of SLE and organ damage.     

MicroRNAs hsa-miR-99b,hsa-miR-330, hsa-miR-126 and hsa-miR-30c: Potential Diagnostic Biomarkers in Natural Killer (NK) Cells of Patients with Chronic Fatigue Syndrome (CFS)/ Myalgic Encephalomyelitis (ME)

Background

Chronic Fatigue Syndrome (CFS/ME) is a complex multisystem disease of unknown aetiology which causes debilitating symptoms in up to 1% of the global population. Although a large cohort of genes have been shown to exhibit altered expression in CFS/ME patients, it is currently unknown whether microRNA (miRNA) molecules which regulate gene translation contribute to disease pathogenesis. We hypothesized that changes in microRNA expression in patient leukocytes contribute to CFS/ME pathology, and may therefore represent useful diagnostic biomarkers that can be detected in the peripheral blood of CFS/ME patients.

Methods

miRNA expression in peripheral blood mononuclear cells (PBMC) from CFS/ME patients and healthy controls was analysed using the Ambion Bioarray V1. miRNA demonstrating differential expression were validated by qRT-PCR and then replicated in fractionated blood leukocyte subsets from an independent patient cohort. The CFS/ME associated miRNA identified by these experiments were then transfected into primary NK cells and gene expression analyses conducted to identify their gene targets.

Results

Microarray analysis identified differential expression of 34 miRNA, all of which were up-regulated. Four of the 34 miRNA had confirmed expression changes by qRT-PCR. Fractionating PBMC samples by cell type from an independent patient cohort identified changes in miRNA expression in NK-cells, B-cells and monocytes with the most significant abnormalities occurring in NK cells. Transfecting primary NK cells with hsa-miR-99b or hsa-miR-330-3p, resulted in gene expression changes consistent with NK cell activation but diminished cytotoxicity, suggesting that defective NK cell function contributes to CFS/ME pathology.

Conclusion

This study demonstrates altered microRNA expression in the peripheral blood mononuclear cells of CFS/ME patients, which are potential diagnostic biomarkers. The greatest degree of miRNA deregulation was identified in NK cells with targets consistent with cellular activation and altered effector function.     

Involvement of CD226+ NK cells in immunopathogenesis of systemic lupus erythematosus

Dysfunction of immune systems, including innate and adaptive immunity, is responsible for the immunopathogenesis of systemic lupus erythematosus (SLE). NK cells are a major part of the innate immune system, and diminished populations of NK cells have been reported in SLE patients. However, the mechanisms behind this decrease and the role of NK cells in SLE pathogenesis remain poorly understood. In this study, we found that a deficiency of NK cells, especially CD226(+) NK cells, is prominent in patients with active SLE. Meanwhile, expression of the CD226 ligands CD112 and CD155 on plasmacytoid dendritic cells is observed in SLE patients; thus, activation of CD226(+) NK cells may be induced by CD226-ligand interactions. Furthermore, IFN-α, which is mainly produced by plasmacytoid dendritic cells, can mediate the activation-induced cell death of NK cells. Therefore, these processes likely contribute to the loss of NK cells in patients with active SLE. Despite the impaired cytotoxicity of peripheral NK cells in human SLE patients and mouse SLE models, we provide evidence that CD226(+) NK cells infiltrate the kidneys of predisease MRL-lpr/lpr mice. Kidney-infiltrating NK cells displayed an activated phenotype and a marked ability to produce cytotoxic granules. These results suggest that, before apoptosis, activated NK cells can infiltrate tissues and, to some extent, mediate tissue injury by producing cytotoxic granules and immunoregulatory cytokines.     

Antibodies to common leukocyte antigen p220 influence human T cell proliferation by modifying IL 2 receptor expression

The p220 antigen is found on the high m.w. form of the T200 common leukocyte antigen family. Although T200 monoclonal antibodies (MAb) react with all hematopoietic cells, p220 MAb react only with B cells, NK cells, about 50% of CD4+ T helper cells, and about 90% of CD8+ T suppressor cells. The p220 antigen appears to play an important role in T cell activation, because anti-p220 MAb at doses as low as 5 ng/ml accelerate the proliferation kinetics of PHA-stimulated T cells and augment anti-CD3-driven proliferation when IL 2 is in excess. Fab fragments have no effect on PHA-stimulated cells but partially block proliferation in response to anti-CD3. Our data suggest that p220 is functionally related to expression of the IL 2 receptor. Anti-p220 MAb cause an increase in the number of T cells that express the IL 2 receptor early after activation. In addition, T cells begin to turn off expression of p220 after activation, and two-color immunofluorescence shows that by day 3 after activation the cells expressing the most IL 2 receptor have the least p220. The loss of p220 on T cells may reflect a post-thymic differentiation process related to cell activation. Our data are consistent with a model where the p220- T cells in peripheral blood are a more activated population of T cells that have lost p220 and its ability to regulate their IL 2 receptor expression.     

Expression and function of inducible co-stimulator in patients with systemic lupus erythematosus: possible involvement in excessive interferon-γ and anti-double-stranded DNA antibody production

Inducible co-stimulator (ICOS) is the third member of the CD28/cytotoxic T-lymphocyte associated antigen-4 family and is involved in the proliferation and activation of T cells. A detailed functional analysis of ICOS on peripheral blood T cells from patients with systemic lupus erythematosus (SLE) has not yet been reported. In the present study we developed a fully human anti-human ICOS mAb (JTA009) with high avidity and investigated the immunopathological roles of ICOS in SLE. JTA009 exhibited higher avidity for ICOS than a previously reported mAb, namely SA12. Using JTA009, ICOS was detected in a substantial proportion of unstimulated peripheral blood T cells from both normal control individuals and patients with SLE. In CD4⁺CD45RO⁺ T cells from peripheral blood, the percentage of ICOS⁺ cells and mean fluorescence intensity with JTA009 were significantly higher in active SLE than in inactive SLE or in normal control individuals. JTA009 co-stimulated peripheral blood T cells in the presence of suboptimal concentrations of anti-CD3 mAb. Median values of [³H]thymidine incorporation were higher in SLE T cells with ICOS co-stimulation than in normal T cells, and the difference between inactive SLE patients and normal control individuals achieved statistical significance. ICOS co-stimulation significantly increased the production of IFN-γ, IL-4 and IL-10 in both SLE and normal T cells. IFN-γ in the culture supernatants of both active and inactive SLE T cells with ICOS co-stimulation was significantly higher than in normal control T cells. Finally, SLE T cells with ICOS co-stimulation selectively and significantly enhanced the production of IgG anti-double-stranded DNA antibodies by autologous B cells. These findings suggest that ICOS is involved in abnormal T cell activation in SLE, and that blockade of the interaction between ICOS and its receptor may have therapeutic value in the treatment of this intractable disease.