CD3zeta Chain Expression Research Articles

Women with cervical cancer have high levels of citrulline that are inversely correlated with percentages of circulating IFN-γ+/perforin+ E6 HPV-16-specific CD8+ T-cells

Women with cervical cancer have high levels of citrulline that are inversely correlated with percentages of circulating IFN-γ+/perforin+ E6 HPV-16-specific CD8+ T-cells

Arginase activity and CD3-zeta chain expression after major surgery: a role for postoperative inmunosupression

Palomero-Rodríguez, M. A.1; Sánchez-Conde, M. P.1; Laporta-Baez, Y.1; Garcia-Navas, R.2; Mollinedo, F.2; De Vicente-Sánchez, J.2 Author Information

SAT0078 Decreased peripheral CD3 zeta chain expression in patients with active rheumatoid arthritis can be restored by various biologic DMARDs and methotrexate

BackgroundCD3 zeta chain (CD3 zeta) is a member of T cell antigen receptor CD3 complex and has crucial role in proximal signal transduction after peptide antigen recognition. CD3 deficiency and...

Vaccination‐induced functional competence of circulating human tumor‐specific CD8 T‐cells

T-cells specific for foreign (e.g., viral) antigens can give rise to strong protective immune responses, whereas self/tumor antigen-specific T-cells are thought to be less powerful. However, synthetic T-cell vaccines composed of Melan-A/MART-1 peptide, CpG and IFA can induce high frequencies of tumor-specific CD8 T-cells in PBMC of melanoma patients. Here we analyzed the functionality of these T-cells directly ex vivo, by multiparameter flow cytometry. The production of multiple cytokines (IFNγ, TNFα, IL-2) and upregulation of LAMP-1 (CD107a) by tumor (Melan-A/MART-1) specific T-cells was comparable to virus (EBV-BMLF1) specific CD8 T-cells. Furthermore, phosphorylation of STAT1, STAT5 and ERK1/2, and expression of CD3 zeta chain were similar in tumor- and virus-specific T-cells, demonstrating functional signaling pathways. Interestingly, high frequencies of functionally competent T-cells were induced irrespective of patient's age or gender. Finally, CD8 T-cell function correlated with disease-free survival. However, this result is preliminary since the study was a Phase I clinical trial. We conclude that human tumor-specific CD8 T-cells can reach functional competence in vivo, encouraging further development and Phase III trials assessing the clinical efficacy of robust vaccination strategies.

Effect of L‐arginine supplementation on immune responsiveness in patients with sickle cell disease

L-arginine (L-Arg) is deficient in sickle cell disease (SSD) during vasoocclusion. We investigated possible causal relationship between L-Arg deficiency and immune dysfunction in SSD in steady-state. Fifteen patients with SSD in steady-state and 13 controls were studied. Plasma L-Arg levels were measured using liquid chromatography. T cell subsets and CD3zeta (CD3zeta) chain expression were analyzed using flow cytometry. Lymphocyte proliferative response to phytohemagglutinin (PHA) and production of IL-6 and interferon-gamma (IFN-gamma) were evaluated with and without L-Arg. SSD patients had significantly lower L-Arg levels than controls. CD3 and CD19 cell populations were comparable for both groups, but SSD patients had above normal numbers of natural killer cells (P = 0.06). Patients and controls exhibited significantly increased lymphocyte blastogenesis to PHA after introduction of L-Arg to cultures; response of patients was significantly greater than values for control individuals. Proliferative response to candida in SSD patients was significantly lower than in controls; L-Arg supplementation did not increase this response. L-Arg had no effect on blastogenic response to PPD and candida albicans. No effect was likewise seen in production of IL-6 and IFN-gamma after addition of L-Arg. CD3zeta chain expression increased after addition of L-Arg in both groups; differences were insignificant. L-Arg levels in steady-state SSD are significantly lower than in controls. L-Arg supplementation enhanced lymphocyte blastogenesis to PHA for both controls and patients, but not in response to antigen. There were no significant differences in CD3zeta chain expression although upregulation of expression occurred after L-Arg supplementation for both groups.

Activated CD11b+CD15+Granulocytes Increase in the Blood of Patients with Uveal Melanoma

To determine whether activated CD11b(+) CD15(+) granulocytes increase in the blood of patients with uveal melanoma. Peripheral blood mononuclear cells (PBMCs) were isolated by density gradient centrifugation from the blood of patients with primary choroidal/ciliochoroidal uveal melanomas (six women, four men; age range, 46-91 years) and healthy control donors (14 women, 10 men; age range, 50-81 years). The expression of CD15 and CD68 on CD11b(+) myeloid cells within PBMCs and primary uveal melanomas was evaluated by flow cytometry. CD3zeta chain expression by CD3epsilon(+) T cells in PBMCs and within primary uveal melanomas was measured as an indirect indication of T-cell function. The percentage of CD11b(+) cells in PBMCs of patients with uveal melanoma increased 1.8-fold in comparison to healthy donors and comprised three subsets: CD68 negative CD15(+) granulocytes, which increased 4.1-fold; CD68(-) CD15(-) cells, which increased threefold; and CD68(+) CD15(low) cells, which were unchanged. A significant (2.7-fold) reduction in CD3zeta chain expression on CD3epsilon(+) T cells, a marker of T-cell dysfunction, was observed in PBMCs of patients with uveal melanoma in comparison with healthy control subjects and correlated significantly with the percentage of CD11b(+) cells in PBMCs. CD3zeta chain expression on T cells within primary tumors was equivalent to CD3zeta expression in PBMCs of the same patient in four of five patients analyzed. Activated CD11b(+) CD15(+) granulocytes expand in the blood of patients with uveal melanoma and may contribute to immune evasion by ocular tumors by inhibiting T-cell function via decreasing CD3zeta chain expression.

Alterations in the expression of signal-transducing CD3ζ chain in T cells from patients with chronic inflammatory/autoimmune diseases

The CD3 zeta chain, a component of the T cell receptor (TCR)/CD3 complex, is considered to be a limiting factor in the assembly and transport of the TCR/CD3 complex to the cell surface and is crucial to receptor signaling function. Recent studies have demonstrated altered expression and function of this signal transduction molecule in T and natural killer cells in patients with chronic inflammatory/autoimmune diseases. In this review, current knowledge concerning the expression of CD3 zeta chain as well as the mechanisms responsible for abnormal expression of this molecule in systemic lupus erythematosus, rheumatoid arthritis, and childhood idiopathic nephrotic syndrome are summarized.

Lung carcinomas do not induce T-cell apoptosis via the Fas/Fas ligand pathway but down-regulate CD3 epsilon expression.

Non-small cell lung carcinoma (NSCLC) patients have impaired cellular immune responses. It has been hypothesized that tumor cells expressing Fas Ligand (FasL) induce in T lymphocytes: (a) apoptosis (tumor counterattack) and (b) down-regulation of CD3zeta expression. However, the hypothesis of tumor counterattack is still controversial. We analyzed FasL expression on NSCLC cell lines and on tumor cells from lung adenocarcinoma patients by flow cytometry and immunocytochemistry. FasL mRNA expression was detected in NSCLC cell lines using RT-PCR, and functional FasL was evaluated on Fas-expressing Jurkat T-cells by annexin-V-FITC staining and by SubG(1) peak detection. Also, the proapoptotic effect of microvesicles released from NSCLC cell lines in Jurkat T-cells was studied. Alterations in the expression levels of CD3zeta, CD3epsilon, and CD28 [measured as mean fluorescence intensity (MFI)] were determined in Jurkat T-cells after co-culture with NSCLC cell lines or tumor-derived microvesicles. Furthermore, the expression levels of CD3zeta and CD3epsilon in CD4+T and CD8+T lymphocytes from lung adenocarcinoma patients was studied. Our results indicate that NSCLC cells neither FasL expressed nor induced apoptosis in Jurkat T-cells. Tumor-derived microvesicles did not induce apoptosis in Jurkat T-cells. In contrast, NSCLC cell lines down-regulated CD3epsilon but not CD3zeta chain expression in Jurkat T-cells; this effect was induced by soluble factors but not by microvesicles. In lung adenocarcinoma patients, significant decreases of MFI values for CD3epsilon, but not CD3zeta, were found in CD4+T and CD8+T cells from pleural effusion compared to peripheral blood and in peripheral blood of patients compared to healthy donors. Our data do not support the tumor counterattack hypothesis for NSCLC. Nonetheless, down-regulation of CD3epsilon in T-cells induced by NSCLC cells might lead to T-cell dysfunction.

Tumor-associated macrophages and CD3-ζ expression of tumor-infiltrating lymphocytes in human esophageal squamous-cell carcinoma

The clinical significance of tumor-associated macrophages (TAMs) and CD3-zeta chain expression of tumor-infiltrating lymphocytes (TILs), and their correlation in human esophageal squamous cell carcinoma (SCC) are not very clear. Serial histological slides from 137 esophageal SCC patients who had undergone tumor resection were immunohistochemically studied with anti-CD68, anti-CD3-zeta and anti-CD3-epsilon antibodies. TAMs infiltration (expressed as macrophage index, M(phi)I) and CD3-zeta expression (judged by Z/E = CD3-zeta+ cells/CD3-epsilon+ cells ratio) in different tissue compartments were observed. We found that the total tumor tissue region had significantly higher macrophage density and lower CD3-zeta expression (mean +/- SD: M(phi)I(normal): 225.3 +/- 85.9; Z/E(total): 0.52 +/- 0.25; n = 137) relative to adjacent histologically normal esophageal squamous epithelium (M(phi)I(normal): 60.5 +/- 31.7, P < 0.001; Z/E(normal): 0.79 +/- 0.35, P = 0.001; n = 70). Significantly higher M(phi)I(stroma) (P = 0.006) and lower Z/E(total) (P = 0.016) were detected in patients with lymph node metastasis than in those without. Patients with high M(phi)I(total) and M(phi)I(cancer) but low Z/E(total) had poorer surgical outcomes. Univariate analysis of M(phi)I(total) and multivariate analysis of M(phi)I(total) with specific clinico-pathological parameters demonstrated M(phi)I(total) to be an independent prognostic factor for survival in esophageal SCC patients (Cox proportional hazard model, P = 0.029 and P = 0.031, respectively).

PD.100 Loss of CD3-zeta chain expression and immune dysfunctions in oral cancer patients

PD.100 Loss of CD3-zeta chain expression and immune dysfunctions in oral cancer patients

L-arginine consumption by macrophages modulates the expression of CD3 zeta chain in T lymphocytes.

L-Arginine plays a central role in the normal function of several organs including the immune system. It is metabolized in macrophages by inducible nitric oxide synthase to produce nitric oxide, important in the cytotoxic mechanisms, and by arginase I (ASE I) and arginase II (ASE II) to synthesize L-ornithine and urea, the first being the precursor for the production of polyamines needed for cell proliferation. L-Arginine availability can modulate T cell function. Human T cells stimulated and cultured in the absence of L-arginine lose the expression of the TCR zeta-chain (CD3zeta) and have an impaired proliferation and a decreased cytokine production. The aim of this work was to test whether activated macrophages could modulate extracellular levels of L-arginine and alter T cell function, and to determine which metabolic pathway was responsible for this event. The results show that macrophages stimulated with IL-4 + IL-13 up-regulate ASE I and cationic amino acid transporter 2B, causing a rapid reduction of extracellular levels of L-arginine and inducing decreased expression of CD3zeta and diminished proliferation in normal T lymphocytes. Competitive inhibitors of ASE I or the addition of excess L-arginine lead to the re-expression of CD3zeta and recovery of T cell proliferation. In contrast, inducible nitric oxide synthase or ASE II failed to significantly reduce the extracellular levels of L-arginine and modulate CD3zeta expression. These results may provide new insights into the mechanisms leading to T cell dysfunction and the down-regulation of CD3zeta in cancer and chronic infectious diseases.

Absent or low expression of T-cell receptor zeta-chain in T cells infiltrating human pathological skin conditions.

The T-cell receptor (TCR) zeta-chain is involved in signal transduction necessary for T-cell activation and subsequent proliferation. Expression of the TCR zeta-chain in vivo has been studied by a variety of technical approaches on different cell and tissue specimen. However, the in situ situation concerning the expression of the TCR zeta-chain has not yet been investigated on infiltrating T lymphocytes in neoplastic and inflammatory cutaneous diseases. In this study, we analysed the expression of the TCR zeta-chain in a number of skin tissues affected by established inflammatory and neoplastic conditions. Serial sections of different tissue specimens were stained immunoenzymatically for CD3 and TCR zeta-chain expression. No or at most scarce expression of TCR zeta-chain was detectable in the inflammatory and neoplastic skin conditions investigated as compared to CD3-positive cells. It is possible that this TCR zeta-chain deletion is induced by the skin microenvironment as an effect of local immunoregulatory influences. Alternatively, lymphocytes located in the skin may generally not express this molecule. In our study, tumour-infiltrating T lymphocytes of CTCL were negative for TCR zeta-chain expression. It has been hypothesized that downregulation of the TCR zeta-chain on tumour-infiltrating T lymphocytes is a mechanism by which neoplastic cells escape the cellular immune response. Our findings showing the absence or reduction of TCR zeta-chain expression also in inflammatory skin lymphocytic infiltrates is not consistent with a pivotal role of the TCR zeta-chain in the process of immune escape of tumour cells.

Shed membrane fragment modulation of CD3-zeta during pregnancy: link with induction of apoptosis

Our objective was to identify shed placental plasma membrane fragments in the maternal circulation and determine whether these fragments are capable of down-regulating CD3-zeta chain expression and inducing apoptosis in T lymphocytes. Sera, isolated from the blood of pregnant women at 26–29 weeks gestation that subsequently had uncomplicated term deliveries, were subjected to high exclusion-limit gel chromatography to isolate placental membrane fragments. The placental origin of the fragments was confirmed by the presence of placental-type alkaline phosphates. These shed membrane fragments were further analyzed for the presence of Fas ligand (FasL) and modulation of CD3-zeta expression on cultured T-lymphocytes (Jurkat cells). The ability of the shed membrane fragments to induce apoptosis was assayed using a cell death ELISA. Components associated with Fas-dependent apoptosis (caspase-3, bcl-2 and bax) were characterized using western immunoblot following exposure to serum-derived membrane fragments. Placental membrane fragments were identified in all pregnancy sera, but not in non-pregnant controls. The 41 kDa FasL was identified in membrane fragment isolates and all samples were capable of inducing apoptosis as determined by the ELISA assay. Exposure of T lymphocytes to isolated membrane fragments suppressed the expression of CD3-zeta. The induction of apoptosis correlated with the induction and activation of caspase 3 and the induction of bax. Placenta-derived membrane fragments are detectable in the maternal circulation. These membrane fragment isolates are capable of inducing FasL-mediated apoptosis and down-regulating CD3-zeta expression, which may contribute to the immune tolerance of the fetus.

Modulation of T-cell CD3-zeta chain expression in early pregnancy.

To assess the modulation of T-cell CD3-zeta chain expression by a factor in the sera of women, prior to egg retrieval and 14 days after an in vitro fertilization (IVF) and to delineate the mechanism of this modulation. In this prospective study, blood samples were obtained from 17 patients during an IVF cycle, prior to human chorionic gonadotropin (hCG), and 14 days after embryo return. Serum was incubated with cultured T-lymphocytes (Jurkat cells) for 96 hr and expression of CD3-zeta chain was evaluated. Eight patients had a positive serum hCG titer 14 days after retrieval, while nine patients had a negative hCG titer. Serum from pregnant patients significantly suppressed CD3-zeta chain expression as compared with their sample prior to retrieval (85.6 +/- 6.2%), while subjects not becoming pregnant failed to suppress zeta expression (99.1 +/- 0.9%, P < 0.0001). A factor capable of suppressing TcR/CD3-zeta expression can be detected in the sera of pregnant women 14 days after embryo retrieval. Loss of zeta chain was associated with the induction of apoptosis.

CD3-zetachain expression of intratumoral lymphocytes is closely related to survival in gastric carcinoma patients.

Impaired or reduced CD3 zeta chain (CD3-zeta) expression in T cells has been identified in various cancers and may be associated with an ineffective immune response. The clinical significance of CD3-zeta chain expression in tumor-infiltrating lymphocytes (TILs) in gastric carcinoma remains unclear. The authors immunohistochemically investigated CD3-zeta expression in TILs in 185 patients who had undergone curative gastrectomy. CD3-zeta/CD3 epsilon (CD3-epsilon) ratios were calculated. Patients were divided into two groups: a normal CD3-zeta group (n = 121) and a reduced CD3-zeta group (n = 64). Patients with a zeta per epsilon ratio of greater than 66% were placed in the normal CD3-zeta group. Patients in the normal CD3-zeta group had fewer lymph node metastasis (P < 0.01) and a shallower depth of invasion (P < 0.05) than those in the reduced CD3-zeta group. The 5-year survival rate was 72% in the normal CD3-zeta group, which was significantly better than that in the reduced CD3-zeta group (55%; P < 0.01). When stratified according to clinical stage, the prognostic value was significantly different only in Stage IV patients. Multivariate analysis showed that CD3-zeta expression was an independent prognostic factor (P = 0.03) next to depth of invasion and lymph node involvement. Reduced CD3-zeta expression in TILs was strongly correlated with progressive disease in gastric carcinomas. CD3-zeta expression in TILs is considered an immunologic, independent prognostic marker in gastric carcinoma patients. CD3-zeta normalization with cytokine treatment may lead to prolonged survival in advanced gastric carcinoma patients.

Modulation of T-cell CD3-zeta chain expression during normal pregnancy

The objectives of this study were to assess the modulation of T-cell CD3-zeta expression by factor(s) present in sera of pregnant women, to correlate this activity with markers of T-cell function associated with pregnancy, and to identify the presence of a circulating pregnancy-associated factor responsible for the suppression of CD3-zeta chain. The suppression of TcR/CD3-zeta expression on cultured T-lymphocytes (Jurkat cells) by sera and amniotic fluids from pregnant women was examined by Western immunoblots and quantitated by densitometry. This suppression was correlated with the induction of T-cell apoptosis and reduced production of IL-2. The serum component suppressing zeta expression was characterized by ultrafiltration and protease sensitivity. Incubation of Jurkat cells with sera obtained from women in the first trimester produced a slight, but not statistically significant, suppression of zeta expression; however, sera from pregnant women in the second and third trimesters and amniotic fluids significantly suppressed zeta levels in a dose-dependent manner. The loss of zeta chain correlated with both reduced secretion of IL-2 and induction of lymphocyte apoptosis. Fractionation of sera by ultrafiltration demonstrated that the zeta chain suppressive factor was <5 kDa, and its trypsin-sensitivity suggests a proteinaceous moiety. Pregnancy is associated with a progressive suppression of cell-mediated immunity. These suppressed T-cell functions have been linked to Fas/Fas ligand-induced apoptosis and suppression of Th1 cytokines, including IL-2. We demonstrate that these pregnancy-associated events are mimicked by a factor(s) present in patient-derived fluids. Suppression of zeta expression appears to be due to a circulating low-molecular-weight protein that suppresses CD3-zeta in a concentration-dependent manner.

Reduced T-cell receptor CD3zeta-chain protein and sustained CD3epsilon expression at the site of mycobacterial infection.

Control of mycobacterial infection by the cellular immune system relies both on antigen-presenting cells and on T lymphocytes. The quality of an effective cellular immune response is dependent on functional signal transduction residing in the cytoplasmic tails of the T-cell receptor CD3 components. In order to investigate potential effects of mycobacteria on T-cell receptor signalling, we examined the protein expression of T-cell signal transduction molecules (CD3zeta, ZAP-70, p59fyn, p56lck). In Western blots of peripheral blood mononuclear cells of Mycobacterium tuberculosis infected patients, only the CD3zeta-chain showed a marked reduction in protein expression. To investigate the situation in situ, immunoenzymatic and immunofluorescence stainings for CD3epsilon and CD3zeta expression were performed on sections of normal lymphoid tissue, M. leprae infected and sarcoid tissue. CD3epsilon and CD3zeta expression were similar with respect to intensity, localization and the number of cells stained in normal lymphoid tissue and in sarcoid granulomas. In contrast, the granulomas of M. leprae infected tissues showed a significantly reduced expression of CD3zeta compared to CD3epsilon. Using double immunofluorescence analysis, virtually no CD3zeta expression could be detected in comparison to the CD3epsilon expression in the lesions. Apparently, mycobacteria are capable of significantly reducing CD3zeta-chain expression, which may be restored by cytokines. IL-2-enhanced zeta-chain expression and T-cell effector functions, defined by interferon-gamma release, in M. tuberculosis-specific and human leucocyte antigen-DR restricted CD4+ T cells isolated from granuloma lesions from patients with pulmonary tuberculosis. Because CD3zeta is essential for CD3 signalling and for eliciting T-cell effector functions, reduced CD3zeta protein expression could result in altered signal transduction and inefficient T-cell effector functions. Alternatively, reduced CD3zeta-chain expression may protect T cells from repetitive TCR stimulation associated with anergy or apoptosis.

Induction of systemic CTL responses in melanoma patients by dendritic cell vaccination: cessation of CTL responses is associated with disease progression.

Two HLA-A2-positive patients with advanced stage IV melanoma were treated with monocyte-derived dendritic cells (DC) pulsed with either tumor peptide antigens from gp100, MART-1 and MAGE-3 alone or in combination with autologous oncolysates. Clinically, the rapid progression of disease was substantially stalled and both patients were alive for more than 15 months after initiation of therapy. Specific CTL reactivity against several tumor antigens was detectable in peripheral blood, which declined just before reactivation of disease progression. Furthermore, CD3 zeta-chain expression detected by Western lotting was decreased in PBL at this time. In summary, our data confirm that DC-based vaccinations induce peptide-specific T cells in the peripheral blood of advanced-stage melanoma patients. Although successful induction of systemic tumor antigen-specific CTL may not lead to objective clinical tumor regression, their presence are indicative of a prolonged survival.

Alteration of signal-transducing molecules in tumor-infiltrating lymphocytes and peripheral blood T lymphocytes from human colorectal carcinoma patients.

Tumor development or growth is accompanied by impaired immune responses, such as a poor proliferative response or down-regulated cytolytic T lymphocyte activity. Although recent reports have suggested that modification of the signal-transducing molecule is responsible for impaired immune responses in tumor-bearing hosts, the causes of defective immune function are not yet completely understood. Furthermore, the clinical significance of the findings is not yet clear. In this study, we investigated the alteration of several signal-transducing molecules in peripheral blood T lymphocytes (T-PBL) as well as in tumor-infiltrating lymphocytes (TIL) from human colorectal carcinoma patients and their relationship with the impaired host immune responses. A greater reduction in CD3zeta chain level was observed in TIL than in T-PBL from tumor-bearing hosts. CD3zeta chain reduction in T-PBL correlated with the clinicopathological stage of a tumor, especially with the status of lymph node metastasis. The levels of p56lck and p59fyn protein tyrosine kinase in T-PBL were also compared between tumor-bearing hosts and normal healthy volunteers. In T-PBL from tumor-bearing hosts, expression of protein tyrosine kinase p59fyn was significantly lower than that of p56lck. However, the level of CD3zeta chain expression did not correlate with T lymphocyte functions such as T lymphocyte proliferative response or allogeneic target cell lysis.