Modulation of T-Cell Adhesion Markers, and the CD45R and CD57 Antigens in Human Alcoholics

Direct and indirect evidence indicates that T cells are altered in alcoholics. The most commonly reported changes under direct examination have been consistent with an increased level of activation as reflected by shifts in the ratio of common leukocyte antigen isoforms expressed at the cell surface, by increases in the expression of class II antigen, or by alterations in the expression of various adhesion molecules. Functional evidence for T-cell abnormality includes loss of delayed hypersensitivity and a number of findings attributed to dysregulation of B cells by alcoholic T cells; these include the widely reported disturbances of immunoglobulin production in vivo and a range of abnormal responses when T and B cells are combined in vitro. Detailed flow cytometric examination of T cells from alcoholics with or without active liver disease reveals a significant loss of l -selectin CD8⁺ T cells, but not usually of CD4⁺ T cells. There is an inverse increase in the expression of CD11b on the CD8⁺ cells that have decreased L-selectin⁺ percentages. Both CD8⁺ and CD4⁺ T cells in alcoholics display a significant loss of the CD45RA isoform and a gain of cells exhibiting the CD45RO isoform. Other surface alterations include increased expression of CD57, a marker most commonly associated on T cells with conditions of chronic increased antigenic exposure. It is argued that these and other T-cell alterations in alcoholics are cytokine-driven in part and result in T-cell differentiation states that are functionally inappropriate. The results of these alterations may include reductions in normal lymphocyte traffic, an increase in cell-mediated cytotoxicity, and intermittent loss of normal suppressor functions for immunoglobulin production permitting increased autoantibody formation. Chronic excessive antigen exposure may contribute at other times to the development of abnormal regulatory suppression of the immune response.     

Identification of activated T cells and the suppressor/inducer subset in patients suffering from severe aplastic anemia

Summary In patients with severe aplastic anemia (SAA), lymphocyte subpopulations were examined for the presence of HLA-DR and 2H4 (suppressor/inducer subset) antigen-expressing cells by flow cytometric analysis. Investigations were performed on peripheral blood lymphocytes before and after therapy with antithymocyte globulin (ATG) and methylprednisolone (MP), as well as on bonemarrow lymphocytes before therapy. Before treatment, only the absolute numbers of CD4⁺ T cells and the CD4⁺HLA^–DR⁺/CD8⁺HLA-DR⁺ activated T cell ratio were significantly decreased (p<0.01 and p<0.001, respectively). Following successful ATG/MP treatment, a decrease in the CD4⁺/CD8⁺T cell ratio was found. Regarding the suppressor/inducer subset, only absolute numbers of CD4⁺/2H4⁺ cells were somewhat higher in treated patients; the percentages were the same in all groups of patients. Studies performed on bonemarrow lymphocytes showed significantly decreased percentages of CD4⁺ and CD8⁺ T lymphocytes, which also express HLA-DR antigen. No significant changes in the distribution of activated T cells following ATG/MP therapy were found, suggesting that these cells play no major role in the pathogenesis of the disease.     

Effect of Acute Stress on Immune Cell Counts and the Expression of Tight Junction Proteins in the Duodenal Mucosa of Rats

Background/AimsDuodenal immune alterations have been reported in a subset of patients with functional dyspepsia (FD). The aim of this study was to investigate the effect of acute stress on immune cell counts and the expression of tight junction proteins in the duodenal mucosa.MethodsTwenty-one male rats were divided into the following three experimental groups: 1) the nonstressed, control group, 2) the 2-hour-stressed group, and 3) the 4-hour-stressed group. Eosinophils, mast cells and CD4⁺ and CD8⁺ T lymphocytes in the duodenal mucosa were counted. The protein and mRNA expressions of occludin and zonula occludens-1 (ZO-1) were examined.ResultsEosinophils, mast cells and CD8⁺ T lymphocyte counts did not differ between the stressed and control groups. The number of CD4⁺ T lymphocytes and the protein and mRNA expressions of occludin and ZO-1 were significantly lower in the 4-hour-stressed group compared with the control group. The plasma adrenocorticotrophic hormone and cortisol levels of the 4-hour-stressed group were significantly higher than those of the control group.ConclusionsAcute stress reduces the number of CD4⁺ T lymphocytes and the expression of tight junction proteins in the duodenal mucosa, which might be associated with the duodenal immune alterations found in a subset of FD patients.     

Blood B,T, CD4+ and CD8+ lymphocytes in female Wistar rats

Summary We have established reference values of peripheral blood lymphocyte subsets in healthy female Wistar rats under highly standardized conditions. Using monoclonal antibodies and flow cytometry, T lymphocytes (OX19⁺), B lymphocytes (OX6⁺ and antiIg⁺), T-helper/inducer (W3/25⁺), and T-suppressor/cytotoxic subsets (OX8⁺) were determined, from week 11 to week 21 after birth. The mean percentages of T and B lymphocytes with respect to total lymphocytes were 78.5% and 18%, respectively; the mean percentages of T-helper/inducer and T-suppressor/cytotoxic cells in relation to T lymphocytes were 59% and 25%, respectively (n=48). No difference in total leukocyte count, differential leukocyte analysis, or lymphocyte subsets was observed during the 10 weeks the rats were studied under standard housing conditions. Therefore, the period considered seems the most appropriate in which to carry out experiments that could involve lymphocyte subset disturbances.     

Colonic mucosal T lymphocytes in ulcerative colitis: Expression of CD7 antigen in relation to MHC class II (HLA-D) antigens

T-cell subsets and their activation state were examined by double-label immunofluorescence of cryostat tissue sections of the colon from 21 patients with ulcerative colitis (UC) and 30 histologically normal controls. Expression of MHC class I (HLA-A, B, C) and class II (HLA-D) antigens was studied in parallel. In the normal colonic mucosa, the CD4CD8 ratio in the epithelial compartment approximated 11, and in the lamina propria, 2.551. Of the CD8⁺ (cytotoxic/suppressor) subset, approximately half did not express the CD5 pan-T marker in either compartment. Virtually no Leu 8⁺ cells were observed, implying that the CD4⁺ subset consisted of helper, rather than suppressor-inducer cells. Classical markers of T-cell activation (CD25, HLA-D) and proliferation were absent, and strong expression of the CD7 immunostimulation marker was approximately equal in both CD4 and CD8 subsets. The epithelium was uniformly negative for class II antigens, but positive for class I. In UC, there were no significant alterations in CD4CD8 ratios in either compartment, and there were no changes with respect to phenotype of the subsets. In 11 of 19 patients (mainly with total colitis), enterocytes were HLA-D⁺. In this HLA-D⁺ group, there was an increase in the percentage of CD4⁺ cells coexpressing CD7; this difference was significant (P<0.02) in the lamina propria. Increased expression of CD7 was also found by the CD6⁺ T cell subset (P<0.05). These results suggest that class II expression is mediated by immunostimulated T helper cells in UC, with consequences for antigen presentation and maintenance of the chronic inflammatory state.HLA-D is used as a generic term for class II major histocompatibility complex (MHC) gene products (HLA-DR, DP, DQ) unless specified otherwise.     

A low blood lymphocyte count is associated with an expansion of activated cytotoxic lymphocytes in patients with B-cell chronic lymphocytic leukaemia

Abstract: In order to determine the relationships between CD2+ lymphocyte subpopulations and tumour mass, the immunophenotype of natural killer (NK) cells and T lymphocyte subsets was studied in 56 B-chronic lymphocytic leukaemia (B-CLL) patients and 38 healthy subjects. The patients were classified according to their blood lymphocyte count (BLC). Forty patients had BLC<30×10⁹/l (low BLC, less tumour mass) and 16 patients had BLC>30×10⁹/l (high BLC, larger tumour mass). The percentage of CD3^– CD56+ cells, as well as of CD8+, CD8+CD45RO+ and CD3+CD57+ T subsets in low BLC patients, were higher than those found in high BLC patients. Conversely, the percentages of CD3+HLA DR+, CD4+ and CD4+CD45RO+ lymphocytes were higher in high BLC patients than in low BLC patients. The CD4/CD8 ratio was decreased in low BLC patients while it was increased in high BLC patients and a significant positive correlation was found between their CD4/CD8 ratio and their BLC. We conclude that in low BLC B-CLL patients there is a decreased percentage of activated helper lymphocytes and an increased percentage of NK cells and activated cytotoxic T lymphocytes. These results suggest a role for NK cells, and helper and cytotoxic T lymphocytes in the control of tumour burden in B-CLL patients.     

The Prognostic Role of Peripheral Lymphocyte Subsets in Patients With Acute Pancreatitis

Background

The prognostic role of peripheral lymphocyte subsets in early stage of acute pancreatitis (AP) is unknown.

Impaired Mitogenic Response of Peripheral Blood T Cells in Ulcerative Colitis Is Not Due to Apoptosis

An abnormal immune response may play apathogenic role in ulcerative colitis (UC). Animalmodels suggest that T-cell regulation may be of centralimportance in the inflammatory process. Our aims werethe characterization of the phenotype andfunctional status of circulating T-cells in ulcerativecolitis patients and to determine if activation-inducedcell death in CD4⁺ and CD8⁺lymphocytes in patients differs from healthy controls.Forty-eight patients (24 women and 24 men) fulfillingthe histopathological, clinical, and immunologicalcriteria for UC were studied. T-cell phenotype andfunction were studied in blood lymphocytes from patientswith ulcerative colitis and healthy donors by flowcytometric analysis, as well as [³H]thymidineincorporation. There were no significant differences in the percentage of T-cell subpopulations(CD3, CD4, CD8) and NK cells in the different groups.The percentage of cells in growth phase S+G₂Mat two and three days of phytohemagglutinin (PHA) stimulation was significantly decreased in UCpatients, but the percentage of CD4⁺ andCD8⁺ cells in UC patients that showedapoptosis was not significantly different than that inthe control group. Proliferative responses to IL-4 also suggested that a reducedresponsiveness to this cytokine may be involved in UC.In conclusion, the impaired proliferative response toPHA of T lymphocytes from UC patients is not associated with an in vitro increase in theapoptotic response in CD4⁺ or CD8⁺cells. A reduced IL-4 response may be involved in thispeculiar mitogenic response. These changes may be pathogenic or a favorable adaptivemechanism.     

Effect of Surrogate Fostering on Splenic Lymphocytes in Fetal Ethanol Exposed Rats

This study evaluated the effects of surrogate fostering as a procedure to control for postnatal effects of ethanol on the maternal female that may indirectly affect the offspring. Effects of fostering on the development of splenic lymphocytes, as well as possible differential effects of fostering on female and male offspring were examined. Litters from prenatal ethanol exposed (E), pair-fed (PF), and ad libitum-fed control (C) condRions were fostered at birth to surrogate untreated dams who had given birth within the same 12-hr period, or were reared by their biological mothers. At 15 and 60 days of age, offspring from each of the conditions were sacrificed and splenic leukocytes were enumerated and analyzed for expression of differentiation antigens, using flow cytometry. At 15 days of age, fostering reduced the percentages of CD45RA⁺ and CD5⁺ cells in E compared with PF and in PF compared with C offspring, and reduced the percentage of CD4⁺ cells in E compared with C offspring. Fostering also had differential effects on E and C offspring, resulting in reduced percentages of CD45RA⁺ and CD5⁺ cells in fostered E compared with nonfostered E offspring, but increased percentages of CD45RA⁺, CD5⁺, and CD8⁺ cells in fostered C compared with non-fostered C offspring. Fostering also down-regulated CD5 antigen expression in E compared with C offspring and up-regulated CD4 antigen expression in C offspring compared with their nonfostered counterparts. At 60 days of age, E females overall had higher percentages of CD45RA⁺ cells compared with C females and higher percentages of CD4⁺ cells compared with PF and C females. Nonfostered E females had higher percentages of CD5⁺ cells than nonfostered C females. In contrast, E males overall had greater percentages of CD4⁺ cells compared with PF and C males. Among males, the percentage of CD5⁺ cells was increased in nonfostered E compared with nonfostered C, whereas the percentages of CD45RA⁺ and CD5⁺ cells were decreased in fostered E males compared with nonfostered E. For both females and males in the nonfostered condition there were no effects of prenatal ethanol treatment on differentiation antigen expression. However, after fostering, E females had higher CD45RA and CD5 antigen expression compared with PF and C females, whereas E males had increased CD4 antigen expression and C males had decreased CD5 antigen expression compared with their nonfostwed Counterparts. These data demonstrate that fostering at birth has differential effects on splenic lymphocyte populations in E, PF, and C offspring. Moreover, the effect of fostering varies with age and has differential long-term effects on female and male offspring. Thus, rather than serving as a control for indirect maternal effects of ethanol on offspring, fostering appears to be a treatment in itself and may actually confound the effects of prenatal ethanol exposure by differentially affecting E, PF, and C females and males.     

Interferon-γ+ and interleukin-2+ T cells in peripheral blood from multiple myeloma patients in relation to disease status and maintenance therapy with interferon-α2b (intron A)

Peripheral blood T cells from 83 patients with multiple myeloma (MM) were examined for the production of interferon-γ (INFγ) and interleukin-2 (IL-2) using three-colour flow cytometry. Comparisons were made between the percentage of cytokine-positive lymphocytes in normal donors and in patients during remission or relapse. Patients were divided into those who were on maintenance therapy with interferon-α2b (intron A) and those who had no further treatment after high-dose melphalan (HDM) with or without autologous bone marrow (ABMR) or peripheral blood stem cell rescue (PBSCR). The percentage of INFγ⁺/CD3⁺, INFγ⁺/CD45R0⁺/CD3⁺ and IL-2⁺/CD8⁺ was higher in patients on INFα2b during remission and relapse compared with normal donors (P < 0.005). During remission INFγ⁺/CD45R0⁺/CD3⁺ and IL-2⁺/CD8⁺ lymphocytes were higher in patients not on INFα2b (P < 0.05 and P < 0.005, respectively). In relapsed patients INFγ⁺/CD3⁺ and INFγ⁺/CD45R0⁺/CD3⁺ were increased in patients not taking INFα2b (P < 0.005). There was no significant difference between the percentages of cytokine-positive lymphocytes in patients taking or not taking INFα2b either during remission or relapse. Plasma IL-6 levels were similar in both groups of patients during remission. The data suggest that if maintenance therapy with INFα2b induces the synthesis of INFγ and IL-2 in vivo, the magnitude of the effect is small and may be unimportant in providing an anti-tumour effect in the majority of patients.     

Activated CD4+ and CD8+ T-cell subsets in Wegener's granulomatosis

Several lines of evidence argue in favour of an involvement of T cells in the pathogenesis of Wegener's granulomatosis (WG). These include the presence of highly specific IgG autoantibodies to proteinase 3, perivascular T-cell infiltrates and elevated amounts of soluble interleukin-2 (IL-2) receptors in patient's serum. In order to further address this question we evaluated by double immunoflourescence and flow cytometry the expression of several cell surface molecules associated with T-cell activation. As compared to healthy controls (n=15), the CD4⁺ subset was significantly diminished, while the percentage of CD8⁺ T cells was elevated in WG patients (n=24). Within the CD4⁺ T-cell subset we found a highly significant increase in activation/memory markers (CD25, CD29, HLA-DR). Within the CD8⁺ T-cell subset the expression of CD11b, CD29 and CD57 was significantly elevated, while the expression of VD28 was reduced. The use of 10 V-, 1 V-and 1 V-specific monoclonal reagents failed to reveal any significant bias in the peripheral T-cell receptor V-gene repertoire of WG patients. There was also no correlation between T-cell activation markers and laboratory parameters [C-reactive protein (CRP), ESR], disease duration or therapy. A significant correlation was found only for the degree of organ involvement and the increase in CD4⁺ T cells coexpressing HLA-DR, as well as the increase in CD57 expression on CD8⁺ T cells. In conclusion, both CD4⁺ and CD8⁺ T-cell subsets were activated in WG. Cytotoxic CD8⁺ CD57⁺ CD11b⁺ CD28^– T cells may directly contribute to damage of vascular endothelium.     

Total parenteral nutrition-associated changes in mouse intestinal intraepithelial lymphocytes

Intraepithelial lymphocytes (IEL) play a major role in mucosal defense mechanisms against intraluminal foreign antigens. To address the role luminal nutrients have on the phenotype and function of the IEL, we administered total parenteral nutrition (TPN) to mice, with the absence of enteral intake. We hypothesized that administration of TPN would result in changes in the phenotype and function of the IEL. For this, we utilized a mouse model of TPN. A significant decline in the CD4⁺ IEL population occurred with TPN. Additionally, the CD8⁺,CD44⁺ IEL subset showed a 65% decline (P < 0.05), and the CD4⁺,CD44⁺ subset declined by 55% with TPN (P < 0.05). The CD8⁺ population (a marker of thymic-dependence) also declined by 92% (P < 0.01) with TPN. IEL in the TPN group showed a significantly lower degree of in vitro proliferation. In conclusion, the IEL showed significant phenotypic changes with TPN including the loss of the thymic-derived population. Functionally, the IEL showed a significant decline in proliferation. Such changes demonstrate the important role luminal nutrients have on IEL phenotype and function.     

Expression of decay-accelerating factor and CD59 in lymphocyte subsets of healthy individuals and paroxysmal nocturnal hemoglobinuria patients

The expression of phosphatidylinositol (PI)-anchored complement-regulatory membrane proteins on circulating blood cells has been well clarified; however, the PI proteins on lymphocyte subsets have not been fully analyzed yet. We examined the expression of decay-accelerating factor (DAF) and CD59 on the T lymphocytes (CD2^?, CD3⁺, CD4^?, and CD8^?) and CD20⁺ B lymphocytes in ten healthy volunteers and 12 paroxysmal nocturnal hemoglobinuria (PNH) patients by cytofluorometry. In healthy controls, each subset of lymphocytes showed a small population of cells weakly positive and a large population of cells strongly positive for DAF and CD59, while erythrocytes showed a single population of cells positive for the PI proteins. The two-population expression of DAF was most distinctive in CD8^? T cells among the subsets. In PNH, each subset of lymphocytes showed a moderately higher population of cells weakly positive and a smaller population of cells strongly positive for the membrane proteins compared with those in the healthy controls. Moreover, in some PNH cases, a negative population for the proteins was found in all subsets. Thus the analysis of PI-anchored proteins on lymphocyte subsets (especially CD8^? T cells) was considered to be of diagnostic value in PNH patients who receive blood transfusion after hemolytic attack of affected erythrocytes. Furthermore, the two-population expression of PI proteins in normal lymphocytes suggests that membrane PI protein would be a new subset marker of lymphocytes.     

Increased frequency of CD4+PD-1+HLA-DR+ T cells is associated with disease progression in CLL

Chronic lymphocytic leukaemia (CLL) patients often have abnormal expansions of CD4⁺ and CD8⁺ T cells and this can be associated with progressive disease. To characterise the key T-cell populations involved in this phenomenon, we used flow cytometry and 11 phenotypic markers to study 74 CLL patients and 14 controls. T cells of CLL patients were more phenotypically complex than those of healthy controls with significant increases in the frequencies of CD4 and CD8 memory T cells expressing exhaustion-, activation- and senescence-associated markers. Multivariate analysis of 111 different T-cell subsets showed that high frequencies of four subsets (three CD8 and one CD4) were associated with shorter progression-free survival. The most significant association was with CD4⁺HLA-DR⁺PD-1⁺ T cells, and patients could be stratified into high- and low-risk groups based on the frequency of these T cells. The expansion of this CD4⁺ subset could not be accounted for by age, cytomegalovirus infection or increases in Treg cells. Overall, these results highlight two relatively simple biomarkers, percentage CD8⁺ and percentage CD4⁺PD-1⁺HLA-DR⁺ T cells, which can be used to risk-stratify CLL patients, independent of other tumour-associated markers. They also provide further evidence for the pivotal role of T cells in modulating the pathology of CLL.     

Highly Active Antiretroviral Therapy (HAART)-Related Hypertriglyceridemia Is Associated With Failure of Recovery of CD14lowCD16+ Monocyte Subsets in AIDS Patients

As cellular reservoirs, CD16⁺ monocyte subsets play important roles in the progression of HIV infection. Previous studies have shown that highly active antiretroviral therapy (HAART) reduced the percentages of CD14^highCD16⁺ monocyte subsets, but did not recover the percentages of CD14^lowCD16⁺ subsets. Eighty-four chronic HIV-infected, HAART-naïve individuals and 55 HIV-negative subjects (31 without hyperlipidemia and 24 with hypertriglyceridemia) were enrolled. Plasma HIV-1 RNA levels, CD4⁺ T-cell counts, triglycerides, total cholesterol, high-density lipoprotein, and low-density lipoprotein were followed up for 48 weeks during HAART treatment in the longitudinal study. We found that mild hypertriglyceridemia in HIV-negative subjects and HIV-infected patients, naïve to HAART, did not affect the percentage of monocyte subsets. However, a failure of CD14^lowCD16⁺ subset recovery was observed in patients with HAART-related hypertriglyceridemia at 48 weeks. Thus, HAART-related hypertriglyceridemia altered homeostasis of monocyte subsets to antiviral therapy, which might further affect immune reconstitution.     

Expression of adhesion molecules on T lymphocytes in young children and infants – a comparative study using whole blood lysis or density gradient separation

We investigated the use of two sample preparation techniques (whole blood lysis [WBL], and Ficoll‐Hypaque density separation [FDS]) for flow cytometric estimation of adhesion molecule (CD54, CD62L and CD11b) expression on T lymphocytes from children less than 2 years old, comparing the results with normal adult controls. Using WBL methods, young children had lower percentages of CD3⁺/CD54⁺ and CD3⁺/CD11b⁺ lymphocytes, but not of CD3⁺/CD62L⁺ lymphocytes than adult controls. FDS was associated with significantly higher percentages of CD3⁺/CD62L⁺ and CD3⁺/CD11b⁺ lymphocytes in young children and adults alike, while the percentages of CD3⁺/CD54⁺ cells from adults was not affected by FDS. The percent expression of CD54, CD62L, and CD11b on T cells from both children and adults were significantly higher following FDS, with a greater increase in CD11b expression on T cells from young children, reaching adult levels. The mean fluorescence intensity (MFI) of CD62L on T cells from young children, which was lower than that of adults using WBL preparation, was significantly higher following FDS, reaching adult levels. The higher levels of adhesion molecule expression associated with FDS did not result from T‐cell activation, as assessed by CD69, CD25, and HLA‐DR expression. Thus, adhesion molecule expression on T lymphocytes from young children is more sensitive to modification by isolation procedures than that on adult T cells. Caution should therefore be exercised in interpreting adhesion molecule expression data on T cells from children.     

Immunoregulatory actions of melatonin and zinc during chronic Trypanosoma cruzi infection

After one century of the discovery of Chagas' disease and the development of an efficient drug with amplitude of actions both in the acute and chronic phase is still a challenge. Alternative immune modulators have been exhaustively used. For that purpose, melatonin and zinc were administered during chronic Trypanosoma cruzi‐infected Wistar rats and several endpoints were assessed. Melatonin has a remarkable functional versatility, being associated with important antioxidant, anti‐inflammatory, and anti‐apoptotic effects. The cross‐talk between zinc and the immune system includes its ability to influence the production and signaling of numerous inflammatory cytokines in a variety of cell types. Our study showed that zinc triggered a decrease in the generation of IFN‐γ for TCD4⁺ cells. Reduced percentage of CD4⁺T cells producing TNF‐α was observed in control melatonin or zinc‐and‐melatonin‐treated animals as compared with untreated rats. On the other hand, a significant increase in the percentage of IL‐4 from CD4⁺ and CD8⁺ T lymphocytes producers was observed 60 days after infection, for all zinc‐treated animals, whether infected or not. Melatonin and zinc therapies increased the percentages of CD4⁺ and CD8⁺ T lymphocytes IL‐10 producers. CD4⁺CD25^highFoxp3⁺ T cells were also elevated in zinc‐ and melatonin‐treated animals. The modulation of the immune system influenced by these molecules affected cytokine production and the inflammatory process during chronic T. cruzi infection. Elucidation of the interplay between cytokine balance and the pathogenesis of Chagas’ disease is extremely relevant not only for the comprehension of the immune mechanisms and clinical forms but, most importantly, also for the implementation of efficient and adequate therapies.     

The Correlation Between the Subsets of Tumor Infiltrating Memory T Cells and the Expression of Indoleamine 2,3-Dioxygenase in Gastric Cancer

Backgrounds

Although many researchers have concentrated on the mechanism of T cell anergy resulting from over-expression of Indoleamine 2,3-dioxygenase (IDO), it remains unclear what alterations will developed in memory T cells (Tm) under over-expression of IDO.

Methods

Immunohistochemical staining for IDO expression in gastric cancer tissues (n = 50) was carried out. Tumor-infiltrating memory Tm cells were counted by flow cytometry. The association between IDO expression and the subsets of tumor infiltrating Tm are discussed.

Results

The higher IDO expressions were significantly associated with deeper invasion (P = 0.016) and higher frequencies (P = 0.038) of lymph node metastasis. The lower tumor-infiltrating CD4⁺Tm were significantly associated with the advanced clinical stage (P = 0.026) and lymph node metastasis (P = 0.016). The lower percentages of CD8⁺Tm were significantly related to undifferentiated histological type (P = 0.042) and lymph node metastasis (P = 0.037). However, the lower percentage of CD8⁺Tem was significantly correlated to differentiated histological type (P = 0.017), lower frequencies of lymph node metastasis (P = 0.014), and earlier clinical stage (P = 0.008). The higher IDO expression patients had significantly lower percentages of CD4⁺Tm (P = 0.012) and CD8⁺Tm (P = 0.033). Nevertheless, it was confirmed that the higher level of IDO expression correlated with higher percentages of CD8⁺Tm cells in univariate and multivariate analysis (P = 0.011).

Conclusion

IDO over-expression and Tm in tumor microenvironments were correlated with the disease stage and histological type of gastric cancer. Higher IDO expression was related to the lower percentage of CD4⁺Tm and CD8⁺Tm, whereas the higher level of IDO expression related with a higher percentage of CD8⁺Tem.     

Morphologic,flow cytometric,functional, and molecular analyses of S100B positive lymphocytes,unique cytotoxic lymphocytes containing S100B protein

Little is known about the S100B⁺ lymphocytes, which are unique human peripheral blood lymphocytes (PBL) containing the S100B protein. It has recently been shown that S100B is released from various types of S100B⁺ cells and exhibits varied cytokine‐like activities. In this study, we precisely characterized the S100B⁺ lymphocytes of healthy adults with respect to the proportion in the whole PBL, immunophenotypes, function, and their S100B mRNA expression and also evaluated their S100B‐releasing activity upon stimulation. S100B⁺ lymphocytes were detected in all individuals examined, and the proportion of S100B⁺ lymphocytes in the whole PBL ranged from 0.42% to 16.15% (mean, 4.21%). In addition, two subtypes of S100B⁺ lymphocytes, a CTL subtype (CD3⁺ CD8⁺ CD16^‐) and a NK subtype (CD3^‐ CD8^‐ CD16⁺), were detected. The majority of the CTL subtype of S100B⁺ lymphocytes expressed the αβ‐T‐cell receptor. Surprisingly, S100B mRNA was detected not only in S100B⁺ lymphocytes, but also in every S100B^‐ lymphocytes, although the expression levels of S100B mRNA in S100B^‐ lymphocytes were much lower than those of S100B⁺ lymphocytes. The CTL subtype of S100B⁺ lymphocytes exhibited blastic morphological changes, proliferated and released S100B upon stimulation with phytohemagglutinin. The NK subtype of S100B⁺ lymphocytes exhibited morphological NK activity when cocultivated with NK‐sensitive target, K‐562 cells. Thus, the CTL subtype of S100B⁺ lymphocytes exhibit the biological characteristics of T cells, while the NK subtype of S100B⁺ lymphocytes exhibit the characteristics of NK cells. These results suggest that S100B⁺ lymphocytes are a particular subtype of cytotoxic lymphocytes that play a unique role in antitumor immunity.     

Lymphocyte Subsets in Peripheral Blood and Smoking Habits

The investigation of peripheral blood lymphocyte (PBL) subpopulations is of interest in a wide variety of inflammatory diseases. Since the number of circulating lymphocytes has been shown to be affected by smoking habits, it seems useful to know how PBL subpopulations are influenced. We therefore determined percentages and absolute numbers of a wide range of PBL subpopulations in smokers (n= 14) and nonsmokers (n= 14). PBLs were obtained from healthy volunteers and analyzed by flow cytometry using antibodies for the detection of CD3, CD4, CD8, CD19, CD56, CD57, CD45RO, CD45RA, α/β and γ/δ T cell receptor epitopes. With the exception of CD3⁺ cells, no differences between smokers and nonsmokers were found regarding percentages of PBL subpopulations. Smokers were found to have higher absolute numbers of PBLs in the following subpopulations compared with nonsmokers: CD3⁺, CD4⁺, CD3⁺α/β⁺, CD45RO⁺/CD4⁺, and CD45RA⁺/CD4⁺. Cytotoxic lymphocytes, natural killer cells, and B cells did not differ in number between smokers and nonsmokers. There was likewise no difference in the number of the CD8⁺α/β⁺ and all cells bearing the γ/δ T cell receptor. Smoking increased the number of T cells and mainly CD4⁺ PBLs. The smoking habits of healthy control groups should therefore be taken into account when comparing lymphocyte subpopulations in different diseases. Accepted for publication: 13 February 1997