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Herpes simplex virus-induced stromal keratitis: role of T-lymphocyte subsets in immunopathology.

Herpetic stromal keratitis (SK), a frequent cause of visual impairment, is considered to represent an immune-mediated inflammatory response to persistent herpes simplex virus virions or subcomponents within the corneal stroma. The experimental disease in mice involves the essential participation of T lymphocytes, but the role of T-lymphocyte subsets in either mediating or controlling the disease is uncertain. In this report, rat monoclonal antibodies were used to selectively deplete mice in vivo of CD4+ (helper-inducer) and CD8+ (cytotoxic-suppressor) T-cell populations and the effect on herpetic SK was evaluated. As measured by flow cytometry, mice treated with anti-CD4 monoclonal antibody (GK 1.5) were greater than 95% depleted of CD4+ T lymphocytes and mice treated with anti-CD8 monoclonal antibody (2.43) were 90% depleted of CD8+ T lymphocytes. Depleted and nonspecific mouse ascites-treated control mice were infected topically on the corneas with herpes simplex virus type 1, and the induction of various immune parameters during the acute infection was evaluated. CD4+-depleted mice failed to produce either a significant antiviral antibody or delayed-type hypersensitivity response but were capable of producing normal cytotoxic T-lymphocyte responses. In contrast, CD8+-depleted mice produced antiviral antibody and delayed-type hypersensitivity responses comparable with those in control animals, but cytotoxic T-lymphocyte responses were markedly reduced. Clinical observations of the corneas revealed that SK in CD4+-depleted mice was significantly reduced, whereas in CD8+-depleted mice SK developed more rapidly, was more severe, and involved a greater percentage of mice. These observations implicate the CD4+ T-lymphocyte subset as the principal mediators of SK and CD8+ T lymphocytes as possible regulators that control the severity of SK.

Animals↗

The relationship between T-lymphocyte subset infiltration and survival in patients with prostate cancer.

The relationship between tumour stage, T-lymphocyte subset infiltration and survival was examined in patients with prostate cancer (n=80). On multivariate analysis PSA (HR 2.47, 95% CI 1.27-4.83, P=0.008) and CD4+ T-lymphocyte count (HR 2.29, 95% CI 1.25-4.22, P=0.008) had independent significance. Increased CD4+ T-lymphocyte infiltration within the tumour was stage independent and associated with poor outcome in patients with prostate cancer.

Aged↗

Lymphocyte subsets of the peripheral blood in myasthenia gravis determined by two-color flow cytometry.

Lymphocyte subsets of the peripheral blood in 43 patients with myasthenia gravis (MG) were determined by two-color flow cytometry using a number of monoclonal antibodies. In the MG patients without thymectomy (Tx) and prednisolone (PSL) treatment, lymphocyte counts, B-cells, CD4+ cells and their subsets were normal, but numbers of T-cells, CD8+ cells and CD8+ CD 11-subsets were significantly decreased. Furthermore, proportions of activated cells in T-cells, CD 16+ Leu7- and CD16+ Leu7+ NK subsets were significantly high in the patients. The changes in T-cells, CD8+ cells and activated T-cells were less marked in the MG patients than Sjögren's syndrome (SS) used as a disease control. Contrary to MG patients, lymphocyte counts, CD4+ cells and their subsets were decreased, and the proportions of B-cells were high in SS patients. These results suggest altered immunologic conditions, immunologically active and deficient conditions, in both diseases, although the alterations were more prominent in SS than MG. PSL treatments and Tx significantly altered the lymphocyte profiles: PSL decreased lymphocytes, B-cells, T-cells, CD4+ cells and their subsets, while the proportions of CD8+ cells were increased. The changes were compatible with the known immunosuppressive effects of PSL. After Tx, lymphocytes and B-cells decreased, but the proportions of T-cells, CD8+ cells and their subsets, and NK cells subsets returned toward normal. CD4+ CD8+ cells were not increased in MG patients, and the cells did not decrease after Tx. Some of these observations might be relevant to clinical effects of Tx, although the mechanism responsible for these changes is still unknown.

Adolescent↗

T lymphocyte subsets and platelet-associated IgG in idiopathic thrombocytopenic purpura: effect of splenectomy.

The modifications of cell-mediated immunity in idiopathic thrombocytopenic purpura (ITP) were investigated using the technique of Moretta et al. [J. exp. Med. 151: 969-974, 1980] to study T lymphocyte subsets in 35 ITP subjects at various clinical stages. In all these patients, platelet-associated IgG (PAIgG) were measured by a complement lysis inhibition technique. Untreated ITP patients did not show significant modifications of T lymphocyte subsets. Patients with thrombocytopenia not responsive to corticoids or splenectomy and increased PAIgG levels showed significant reduction of Fc-gamma-bearing T lymphocytes (T-gamma). Patients 'cured' by splenectomy (12 out of 17) showed a normal PAIgG level, a marked increase of T-gamma cells and a reduction of T-mu cells; patients 'not cured' by splenectomy showed an increased PAIgG level and a reduction of T-gamma cells. An increase of PAIgG levels and a reduction of T-gamma cells were found in 2 splenectomized patients after relapse. An association between chronic nonresponsive ITP and decrease of T-gamma cells was established.

Adolescent↗

Lifespan of human lymphocyte subsets defined by CD45 isoforms.

The lifespan of thymic-derived or T lymphocytes is of particular interest because of their central role in immunological memory. Is the recall of a vaccination or early infection, which may be demonstrated clinically up to 50 years after antigen exposure, retained by a long-lived cell, or by its progeny? Using the observation that T lymphocyte expression of isoforms of CD45 corresponds with their ability to respond to recall antigens, we have investigated the lifespan of both CD45R0 (the subset containing responders, or 'memory' cells) and CD45RA (the unresponsive, or 'naive' subset) lymphocytes in a group of patients after radiotherapy. Here we report rapid loss of unstable chromosomes from the CD45R0 but not the CD45RA pool. Immunological memory therefore apparently resides in a population with a more rapid rate of division. Differing survival curves for the two subsets are best described by a model in which there is also reversion in vivo from the CD45R0 to the CD45RA phenotype. Expression of CD45R0 in T cells may therefore be reversible.

Chromosome Aberrations↗

Lymphocytes subset in hyperandrogenic women with polycystic ovarian disease.

The correlation between the sex steroids and the immune system is already known but the relationships between testosterone, progesterone, estradiol, cortisol and the immune system are still controversial. The Authors have studied lymphocytes subset in ten women with polycystic ovarian disease (PCOD); it was observed: 1) a decrease of T8 in 64% and of natural killer activity (NK) in 60% of patients, and an increase of T4/T8 ratio in 55%; 2) a significant negative correlation between androstenedione and testosterone (P less than 0.05); 3) a significant positive correlation between the decreased T8 lymphocytes and NK cells and the increase of testosterone and androstenedione serum values. The data suggest that in PCOD a correlation between endocrine and immune system exists and the androgens can affect the lymphocyte subset.

Adolescent↗

Quantitative bone histomorphometry and circulating T lymphocyte subsets in postmenopausal osteoporosis.

To explore the influence of the immune system on the development of osteoporosis, 19 untreated postmenopausal women with osteoporosis were studied by means of quantitative histomorphometry of the ilium and an analysis of T lymphocyte subsets in the peripheral blood. Osteoporotic women had lower OKT3+ and OKT8+ counts and a higher OKT4+/OKT8+ ratio than nonosteoporotic control subjects. Linear regression analyses disclosed that the age of subjects correlated with bone mineral density (BMD; r = -0.634, p less than 0.01) and some of the histomorphometric parameters for bone formation (r = -0.694 to -0.467, p less than 0.01-0.05). The number of OKT4+ cells showed weak but significant negative correlation with the parameters for bone resorption (r = -0.549 to -0.462, p less than 0.05). In a multiple regression analysis, the advanced age, the increase in OKT3+, and the decrease in OKT4+ and OKT8+ counts were shown to be significant predictors for the decrease in BMD (R = 0.882, p less than 0.01). According to the regression formula obtained from the analysis, the parameters for bone formation were related only to the age of subjects whereas those for bone resorption were tightly associated with the number of OKT4+ and OKT8+ cells but not with the age of subjects. These results indicated that, in addition to the age factor, abnormalities of the peripheral T lymphocyte subsets, especially those of OKT4+ and OKT8+ cells, are closely associated with the decrease in bone mass in postmenopausal osteoporosis, supporting the causal relationship between T lymphocyte functions and the development of postmenopausal osteoporosis.

Aged↗

Quantitative monitoring of serum hepatitis B virus DNA and blood lymphocyte subsets during combined prednisolone and interferon-alpha therapy in patients with chronic hepatitis B.

Several investigators have reported a significantly reduced CD4/CD8 ratio, as defined by monoclonal antibodies, in the peripheral blood of Caucasian patients with chronic active hepatitis B (CAHB). In Asian patients with chronic hepatitis B, quantitative analyses of subpopulations of peripheral blood lymphocytes have not been able to confirm these findings. In this work, we analysed the frequency of peripheral blood lymphocyte subsets in 10 Chinese patients with histologically proven CAHB and seven healthy Chinese individuals. Four of the 10 CAHB patients received combined prednisolone/interferon-alpha2b (IFN-alpha2b) therapy. Peripheral blood samples were consecutively collected for analysis of lymphocyte subpopulations using an indirect immunofluorescence (IF) method, and hepatitis B virus (HBV) DNA was quantified by a chemiluminescent, molecular-hybridization assay. Peripheral blood mononuclear cells from seven Chinese control individuals comprised 63 +/- 3% CD3+ cells, of which 41 +/- 4% were of CD4+ and 23 +/- 2% of CD8+ subsets. The mean CD4/CD8 ratio in the healthy controls was 1.9 (95% confidence interval = 1.1-2.7). The CD4/CD8 ratios were significantly reduced (P < 0.01) in the 10 patients with chronic hepatitis B, compared with those of the controls, owing to a significant increase in the number of CD8+ cells (P < 0.005). During the treatment with prednisolone, a significant increase in the CD4/CD8 ratio was observed in all treated patients. This increase was mainly caused by a decrease in the number of CD8+ cells and was accompanied by an increase in serum HBV DNA levels, which peaked during the latter part of the prednisolone cycle. During the treatment with IFN-alpha2b, a second increase in the CD4/CD8 ratio was observed, which was caused by an increase in CD4+ cells. A marked decrease in viral load was observed, during treatment with IFN-alpha2b, in patients with HBV DNA levels below 10 000 pg ml-1. Our data indicate that the CD4/CD8 ratios in Chinese CAHB patients do not differ from those of Caucasian patients with CAHB, when analysed using similar methods for the enumeration of lymphocyte subsets. Profound effects on cellular distribution and viral replication were noted during the combined prednisolone/IFN-alpha2b therapy. Additional studies of the modulatory effect of the combined therapy on the distribution of lymphocyte subsets and cytokine profiles in relation to the therapeutic outcome of HBV infection are warranted.

Adult↗

Leucocytes in human milk and lymphocyte subsets in cow's milk-allergic infants.

The breast-fed infant ingests an average of 108 leucocytes per day, with breast-feeding often continuing for several months. The precise role of human milk leucocytes is still unresolved. Breast-feeding has been recommended for infants at high risk of allergy to prevent or delay the development of food allergies and atopic eczema. However, studies dealing with distinct immunologic factors in the mother's milk, and their effect on health status or development of allergies in the infant, are scarce. We evaluated the relationship between the cellular composition of human milk and the development of cow's milk allergy (CMA) in the breast-fed infant. Leucocyte subsets in the breast-fed infants were also measured. The study population comprised 61 breast-feeding mothers and their infants. Thirty-nine mothers each had a cow's milk-allergic infant, 10 had an infant with atopic dermatitis without CMA, and 12 mothers had a healthy infant. Leucocyte subsets in mothers' milk were counted using a light microscope and confirmed by flow cytometry. In infants, peripheral blood lymphocyte subsets were determined by flow cytometry and were correlated with the health status of the breast-fed infant and leucocyte composition of the mother's milk. Human milk was found to be a non-homogenous morphological entity. In the milk of mothers of infants with CMA, the proportion of macrophages was significantly smaller than in the mothers with infants without CMA (p = 0.036, t-test). Mothers with high proportions of neutrophils in their milk (> 20%) had significantly more often infants with CMA than did those with low proportions of neutrophils (p = 0.02; Fischer's exact test). Eosinophils comprising > 1% of milk cells were only detected in the mothers who had infants with CMA. Furthermore, the proportions of CD4+ T cells were positively correlated with the proportion of milk macrophages and negatively with the percentage of milk neutrophils and eosinophils. The proportions of total B cells and those expressing CD23, a low-affinity immunoglobulin E (IgE) receptor, were positively correlated with the proportions of neutrophils and eosinophils in mother's milk and negatively with the percentage of milk macrophages. To conclude, the composition of breast milk in some mothers is abnormal and correlates with a diagnosis of CMA in a breast-fed infant. This may provide a new and interesting insight into the development of food allergies.

B-Lymphocyte Subsets↗

[Effect of thymostimulin on chemotherapy-induced changes in lymphocyte subset distribution. A longitudinal study in patients with primary inoperable oropharyngeal cancers].

20 patients with primarily inoperable squamous cell carcinoma of the head and neck were treated with 2 cycles of chemotherapy prior to surgical treatment. The lymphocyte subsets were determined prior to chemotherapy, as well as directly after and 3 weeks after start of each chemotherapy cycle. For this, 2 different flow cytometric techniques were used in parallel. After the second cycle of chemotherapy 10 patients were additionally treated with thymostimulin for 2 weeks. The remaining ten patients received no additional immunotherapy and served as control group. Three patients (one in the control group and two in the verum group) had to be excluded from the study because the immunological examination could not be performed in accordance with the protocol. In addition another patient had a local skin reaction so that thymostimulin therapy could not be started as allergy to bovine proteins was suspected. Prior to the first chemotherapy treatment, there is no detectable defect in the distribution of lymphocyte subsets. Under chemotherapy treatment the absolute number of Pan-T cells, T-helper cells, T-suppressor cells and Leu-10 positive B-cells decreased. In contrast, additional treatment with thymostimulin directly after the chemotherapeutic regimen resulted in a marked increase of all T-lymphocytes as well as the Leu-10-positive cells. The results measured with two different flow cytometric techniques were comparable.

Adjuvants, Immunologic↗

Changes in peripheral blood lymphocyte subset frequencies in myasthenia gravis patients are related to immunosuppression.

Surface antigens on peripheral blood lymphocytes from myasthenia gravis patients were investigated. The expression of DR+ and CD8+/DR+ T lymphocytes was increased and the expression of CD4+ T cells reduced. Neither thymectomy, clinical condition nor anti-acetylcholine receptor antibody titre correlated with any of the changes in peripheral blood lymphocyte subsets observed. However, immunosuppressive therapy correlated with the significant reduction in CD4+ and CD2+/CD4+ T cells in these patients.

Adolescent↗

Differential activity of P-glycoprotein in normal blood lymphocyte subsets.

To better understand the phenomenon of P-glycoprotein (P-170) expression we investigated lymphocyte subpopulations for P-170 function in healthy volunteers. Studies were based on three-colour flow cytometry including the fluorescent probe rhodamine 123 (Rh123), which is transported by P-170. Marked Rh123 efflux was detected in CD8+ T lymphocytes with CD8+/CD45RA+ T cells (naive cells) showing significantly higher P-170 activity as compared with CD8+/CD45RA- cells (P<0.04). Vice versa, CD8+/CD45RO+ T cells (memory cells) demonstrated less P-170 activity than CD8+/CD45RO- cells (P<0.04). P-170 function was less prominent in CD4+ T cells, however, Rh123 efflux was higher in the CD4+/CD45RA+ and CD4+/CD45RO- subpopulations (P<0.025) corresponding to the CD8+ results. Dye efflux differed significantly between activated and non-activated CD8+ and CD4+ as well as CD8+/CD11b+ and CD8+/CD11b- T lymphocytes. Since CD16+ natural killer cells (NK) expressed the highest level of P-170, the NK cytotoxicity against 51Cr-labelled K562 target cells was assayed in the presence or absence of P-170 inhibitors. NK related cytotoxicity was significantly reduced in the presence of R-verapamil and dexnigaldipine-HCP in a dose-dependent manner. The differential expression of P-170 activity in naive and memory T cells together with the reduced NK related cytotoxicity in the presence of MDR-modulators suggest a physiological role of P-170 in immunological functions of these lymphocyte subsets. Consequently, the addition of MDR modulators to conventional chemotherapy as a strategy to overcome drug resistance should consider possible adverse immunosuppressive effects.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

Kinetic responses of parasite-specific antibody isotypes, blood leucocyte pattern and lymphocyte subsets in rats during primary infestation with Fasciola hepatica.

Antibody responses, blood leucocyte and splenic lymphocyte subset patterns were studied during a primary infection with Fasciola hepatica in the rat. The infection induced parasite-specific IgM by 2 weeks after infection. High levels of IgM antibodies were maintained for many weeks. The IgE response was biphasic with peaks at 5 and 9 weeks after infection which were correlated with different phases in the development of F. hepatica in the rat. Both IgG2a and IgG1 antibodies were detected but the titre of IgG2a augmented slightly and rose more slowly than did that of IgG1. There was a rise in neutrophil and eosinophil numbers. Neutrophils did not increase before the fourth week but eosinophil numbers were raised by the second week after infection and remained high during the whole migratory phase of the parasite. In the spleen, the percentage of B lymphocytes increased and there was a decrease in the percentages of CD4+ and CD(8+)-like T lymphocytes. These results suggested that, in the rat infested with F. hepatica, TH2-like lymphocytes could be preferentially stimulated, as has been reported in murine schistosomiasis.

Analysis of Variance↗

Observation of T lymphocyte subsets in the liver of patients with advanced schistosomiasis and advanced schistosomiasis accompanied with hepatitis B.

T lymphocyte subsets in the liver were detected by Avidin-Biotin Complex (ABC) assay in 22 patients with advanced schistosomiasis (AS) and 5 cases of AS accompanied with hepatitis B. T lymphocytes in the liver of AS patients were distributed in the peripheral layer of egg granuloma or the area near eggs in non-granuloma. No infiltrative T lymphocytes were observed in area with extensive fibrosis. There was infiltration of many T cells in the portal tract, piecemeal and focal necrosis area as well as in hepatic sinus in AS patients accompanied with hepatitis B. CD8+ T cells (suppressor/cytotoxic T cells, Ts/Tc) in the liver were predominant in the two groups. In AS patients, marked hepatic fibrosis, a small number of T cell infiltration and slight hepatocellular degeneration and necrosis were observed. However, obvious hepatocellular degeneration and necrosis were seen in AS patients accompanied with hepatitis B, and 3 cases of them developed active liver cirrhosis. The results indicated immune response was weak in the liver in AS patients and Ts cells might be predominant in the subset of CD8+ T lymphocytes. Cellular immune response was relatively strong in AS patients accompanied with hepatitis B and the infiltrative CD8+ T lymphocytes might be mainly Tc cells.

Adolescent↗

The natural history of lymphocyte subsets infiltrating the pancreas of NOD mice.

A longitudinal study of lymphocytic infiltration in the endocrine pancreas of non-obese diabetic mice was performed to investigate the role of different lymphocyte subsets in the pathogenesis of diabetes. The incidence of insulitis and the percentage of mononuclear cell subsets in the pancreas were evaluated in non-obese diabetic mice of various ages (5, 9, 13, 17, 22, 29 and 36 weeks). Cryostat sections of pancreas were stained with heamatoxilin-eosin or with different monoclonal antibodies against total T lymphocytes, helper T lymphocytes, cytotoxic/suppressor T lymphocytes, activated interleukin 2 receptor positive lymphocytes and B lymphocytes. A monoclonal antibody against Class-II antigens was also used. Positive cells were revealed by the immunoperoxidase technique. Insulitis was found in 5 weeks old mice but to a lesser extent than in adult animals. No significant variation between infiltrating cell subsets was found in different age groups. T lymphocytes ranged between 20.4% and 28.1%, B lymphocytes between 28.8% and 30.8% and Class-II positive cells between 22.8% and 32.2%. Interleukin 2 receptor positive cells ranged between 5.5% and 8.5% as detected with AMT-13 monoclonal antibody which recognise the interleukin 2 binding site. A higher percentage of activated cells was observed using another monoclonal antibody (7D4) directed against a different epitope of the interleukin 2 receptor, suggesting the presence of activated lymphocytes with interleukin 2 receptors saturated by interleukin 2. No insulin-containing cells were found to express Class-II molecules as demonstrated by a double immunofluorescence technique. Most infiltrating mononuclear cells were found to be positive for Class-II and L3T4 antigens or to be Class-II positive and express surface immunoglobulins.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Lymphocyte subsets in patients with lung cancer treated with thiophosphoric acid alkaloid derivatives from Chelidonium majus L. (Ukrain).

Lymphocyte subsets were evaluated in nine men (aged 42-68 years, mean 57 years) with histologically proven lung cancer, previously untreated. Lymphocyte subpopulations were quantified by immunofluorescence using monoclonal antibodies against total T-cells, T-helper and T-suppressor cells. In addition, the percentage of NK cells and the helper/suppressor (H/S) ratio were evaluated. The number of B-cells was determined by surface immunoglobulin immunofluorescence. Chelidonium majus L. (preparation Ukrain) was applied as an intravenous injection every three days. One course consisted of 10 applications of 10 mg each. All immunological tests were performed before and after drug administration. The treatment was generally well tolerated. The results showed an increase in the proportion of total T-cells, and a significant decrease in the percentage of T-suppressor cells. The normalization of the H/S ratio was also noted. However, there were no signs of activation of NK, T-helper and B-cells. The restoration of cellular immunity was accompanied by an improvement in the clinical course of the disease. This effect was particularly pronounced in patients who responded to further chemotherapy. Objective tumour regression (CR+PR) was seen in 44.4% of treated patients. Four out of nine patients (44.4%) died of progressive disease during the course of this study. It is concluded that Ukrain can be immunologically effective in lung cancer patients and can improve human cellular response.

Adjuvants, Immunologic↗

Assessment of lymphocyte subsets and neutrophil leukocyte function in chronic psychiatric patients on long-term drug therapy.

1. Lymphocyte subsets and neutrophil function were studied in chronic psychiatric patients on long-term multiple drug therapy, and the correlation with disease and drug treatments was also investigated. 2. Patients showed decreased CD4+ percentage, CD4+/CD8+ ratio, B lymphocyte percentage, increased absolute and percentage NK, and impaired neutrophil chemotaxis, phagocytosis, and oxidative metabolism. Correlations were found between: patient status and lower CD4+ percentage, CD4+/CD8+ ratio, neutrophil chemotaxis, phagocytosis, and oxidative metabolism; schizophrenia and higher absolute and percentage NK, and neutrophil random migration; benzodiazepines and higher CD8+ absolute number and lower NBT reduction frequency; carbamazepine and higher NK percentage, neutrophil chemotaxis, phagocytosis, stimulated O2-production, and lower microbicidal activity. 3. The present study provides direct evidence for the existence of immunologic abnormalities in a population of chronic psychiatric patients on long-term drug therapy. Although a possible role by other factors could not be excluded, such abnormalities are strongly associated with chronic administration of benzodiazepines and carbamazepine. Further investigations are warranted to confirm these findings and disclose the possible mechanism(s) involved.

Adult↗

[Distribution of lymphocyte subsets in regional lymph nodes in uterine cervical cancer and its immunological significance].

We performed an immunohistochemical study on the distribution of lymphocyte subsets in the regional lymph nodes in uterine cervical cancer. The lymph nodes from patients with cervical cancer were stained with monoclonal antibodies to Leu 1,2,3 and 7 as the indicators of pan T, killer/suppressor T, helper/inducer T, and NK/K cells, respectively. On the lymph nodes without tumor involvement, there were a greater number of lymphocytes of each subset in the nodes in stage Ib without nodal metastasis than in the nodes in stage Ia and stage Ib with nodal metastasis and stage II, suggesting that the regional lymph nodes have active immune reactions in stage Ib cancer without nodal metastasis. On the lymph nodes with tumor involvement, there were great numbers of lymphocytes of each subset, especially Leu 7+ cells, in the nodes with local invasion, suggesting that very strong immune reactions occur in these nodes. On the other hand, there were very few lymphocytes in the nodes with diffused tumor. The present findings suggest that regional lymph nodes in cervical cancer play important roles in antitumor immune response and, furthermore, even the tumor-involved lymph nodes are of great immunological significance if the metastatic tumor is localized.

Adult↗