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Flow cytofluorimetric analysis of lymphocyte subset alterations in cattle infected with bovine viral diarrhea virus.

Clinically normal, bovine viral diarrhea virus (BVDV)-seronegative, 7 to 9-month-old steers were inoculated intranasally with NY-1, a noncytopathic strain of BVDV, or exposed intramuscularly to killed BVDV. Indirect immunofluorescence staining with monoclonal antibodies specific for bovine leukocyte subsets followed by flow cytometric analysis was used to monitor subsequent hematologic alterations. Infection with BVDV resulted in a transient leukopenia which was characterized by decreases in the absolute numbers of circulating T lymphocytes, including BoT4+ ("helper") and BoT8+ ("cytotoxic/suppressor") subsets, B lymphocytes, and neutrophils. There was no significant variation in numbers of non-T, non-B ("null") lymphocytes or monocytes. Exposure to inactivated BVDV in a combination vaccine did not cause significant alteration in the circulating numbers of any major leukocyte subset; however, significant variation was seen in the BoT4/BoT8 ratios and in the numbers of cells expressing major histocompatibility complex (MHC) class II antigens.

Animals↗

Immunoregulatory CD4+ CD45R+ suppressor/inducer T lymphocyte subsets and impaired cell-mediated immunity in patients with Down's syndrome.

The monoclonal antibodies 2H4 and 4B4 allow CD4+ and CD8+ T lymphocytes to be subdivided into CD45R+ and CDW29+ functional subpopulations. The CD4+ CD45R+ lymphocytes are designated as suppressor/inducer and CD4+ CDW29+ as helper/inducer subsets. Peripheral blood lymphocytes from 19 patients with Down's syndrome and 19 age- and sex-matched normal controls were analysed for the CD45R+ and CDW29+ subsets from the CD4+ and CD8+ T lymphocytes. The percentage of CD4+ CD45R+ cells (suppressor inducer) was markedly increased and of CD4+ CDW29+ cells (helper/inducer) decreased in all patients with Down's syndrome. In contract, the percentage of CD8+ CD45R+ and CD8+ CDW29+ subsets showed no major differences between patients with Down's syndrome and normal controls. Moreover, an alteration in the CD4+ and CD45R+ and CD4+ CDW29+ T cell subsets was accompanied by a markedly reduced proliferative response to phytohaemagglutinin and concanavalin A stimulation of the CD4+ T lymphocytes. Thus, a deficiency exists in patients with Down's syndrome in the CD4+ CDW29+ helper/inducer T cell subset which may contribute to their impaired cell-mediated immunity.

Antigens, Differentiation↗

Age- and sex-related changes of the lymphocyte subsets in healthy individuals: an analysis by two-dimensional flow cytometry.

The relative and absolute numbers of the peripheral blood lymphocyte (PBL) subpopulations from 156 healthy men and women of different ages (20-99 years old) were studied by the use of monoclonal antibodies (MoAbs) and two-dimensional flow cytometry. The percentage of pan-T MoAb-positive cells decreased with age, which was attributable to a relative decline in the CD8+ suppressor/cytotoxic T cells, more precisely in the CD8+ CD11- cytotoxic T cells. Those of CD4+ Leu-8- helper T cells, CD3+ HLA-DR+-activated T cells, and natural killer subsets (Leu-7+, CD16+, Leu-7+ CD16+ or Leu-7+ CD16-) increased with age. The absolute numbers of most of the lymphocyte subsets examined declined with age except that those of natural killer cell subsets and helper T cells remained unchanged. It should be noted that the PBL subsets differed markedly according to age and sex, the changes being more evident among women.

Adult↗

Immunoregulation in onchocerciasis. Functional and phenotypic abnormalities of lymphocyte subsets and changes with therapy.

To help define the immunoregulatory defects in patients with onchocerciasis, flow cytometric analysis of circulating lymphocyte subpopulations was performed in parallel with functional assays. No significant differences in CD4/CD8 ratios were seen when microfilariae-positive individuals from Guatemala were compared with Guatemalan controls. However, the infected individuals had significantly increased numbers of circulating CD4+CD45RA+ lymphocytes (mean 38.3%) when compared with controls (mean 16.0%). Coexpression of the activation marker HLA-DR was significantly increased on CD4+ cells from infected individuals. In contrast, no up-regulation of HLA-DR was seen on CD8+ or CD19+ cells. At 1 year after initiation of treatment with semiannual doses of the microfilaricide ivermectin, there were significant increases (P less than 0.05) in the percentage of CD4+CD45RA- cells, the percentage of CD4+HLA-DR+ cells, and mitogen-induced lymphokine production (IL-2, IL-4). Despite these changes, parasite-specific IL-2 and IL-4 production which had been undetectable before treatment did not manifest itself even by the 2-yr follow-up. Defects in the T-cell activation pathway in Onchocerca volvulus-infected individuals may thus exist at several independent points; a state of parasite antigen-specific tolerance appears to remain even after the relative reversal of other generalized immunoregulatory defects.

Adolescent↗

[Evolution of CD4 and CD8 lymphocyte subsets in HIV seronegative and seropositive hemophiliacs].

PURPOSE: To asses the evolution over time of the CD4, CD8 and total lymphocyte counts in both HIV seronegative and HIV seropositive haemophiliacs. PATIENTS AND METHODS: A total of 124 haemophilic patients (77 HIV seropositive and 47 HIV seronegative) regularly controlled at our Hospital were studied. All patients have had several (range 2-15) CD4 and CD8 lymphocyte counts, with a minimum interval of 2 or more years between the first count and the last one (median 7.7 years). A linear regression analysis of the serial cell counts against the time was made and the corresponding slopes were estimated for each patient and expressed as a percentage of the initial values (standardized slopes). RESULTS: In HIV seronegative patients the number of CD4, CD8 and total lymphocytes decreased (p < 0.05), but the decrease of the CD4/CD8 ratio was not statistically significant. The decline of the CD4 cells showed a trend to remain above 500 cells/microL. HIV seropositive hemophiliacs had also a substantial decline of lymphocyte counts (p < 0.0001), but essentially due to changes of the CD4 cells, which declined with a nearly constant rate during the follow-up (median values decreased from 793/microL to 324/microL). CD8 cell counts diminished in lower grade than the CD4 cells counts, and consequently the percentage of the total lymphocyte number increased from 42% to 55%. The CD4/CD8 ratio decreased with a standardized slope of -5.7% per year. At the initial evaluation the seropositive patients had CD4 cell counts and CD4/CD8 ratios lower than seronegative ones, but CD8 cell counts were higher in the first group. Total lymphocyte counts were not statistically different at this first evaluation between infected and non-infected patients. At the final evaluation, differences on CD4 cell counts and CD4/CD8 ratio increased, and lymphocyte counts were significantly lower in the HIV-seropositive haemophiliacs. However, the final absolute counts of CD8 cells were not dissimilar in the two groups of patients. The median standardized slopes of the total lymphocyte counts, CD4 cell counts, and CD4/CD8 ratios were significantly more negative for the HIV-infected patients (-5.7%, -9.4 and -5.7%, respectively) than were these for the non-infected ones (-2.0%, -3.2% and -2.5%, respectively). CONCLUSIONS: Lymphocyte counts, mainly the CD4 cell counts, decline in treated haemophiliacs independently of their HIV status. In the HIV seronegative patients the CD4 cell counts generally stabilize above 500 cells/microL. However, in the seropositive haemophiliacs these counts show a continuous fall, with a median rate of 9% of the initial value per year.

Adolescent↗

Modification of lymphocyte subsets in the intestinal-associated immune system and thymus by chronic ethanol consumption.

Modification of the mucosa-associated intestinal immune system of female C57BL/6 mice was studied during consumption of the Lieber-DeCarli diet supplemented with 5% v/v ethanol or laboratory chow with ethanol (20% w/v) in the drinking water. All groups received ethanol for 11 weeks. Mice fed the Lieber-DeCarli diet had fewer CD8+ cells/villus than the chow-fed controls. Mice that received ethanol in the drinking water had fewer IgA-containing cells and CD8+ cells than controls. There were no differences in the number of cells in the mesenteric lymph nodes between ethanol-treated mice and their respective controls. Nevertheless, chow-fed control mice had more cells than those fed the Lieber-DeCarli control diet. Although no differences were detected in the percentages of CD4+, CD8+, LECAM-1+, and LECAM-1+ CD4+ cells, there was a decrease in the percentage of LECAM-1+ CD8+ cells in ethanol-fed mice when compared with their Lieber-DeCarli controls. Mice receiving ethanol in the drinking water showed alterations in the CD4 CD45RC subsets and in the CD8 CD45RC subsets. Similar results were observed in mice receiving Lieber-DeCarli diets alone or supplemented with ethanol. The low dose, chronic exposure of dietary ethanol in the Lieber-DeCarli-fed mice did not significantly affect the numbers of various thymocyte subsets. But, a decrease in the percentage of CD4- CD8+ cells was observed in the thymus of mice receiving ethanol in the drinking water. Chronic ethanol consumption caused significant decreases in the number of CD8+ and IgA+ cells in the intestinal lamina propria, important in mucosal immune defenses.

Alcoholism↗

Abnormalities of T lymphocyte subsets in Behçet's disease demonstrated with anti-CD45RA and anti-CD29 monoclonal antibodies.

We assessed T cell subpopulations using 2-color flow cytometry with phycoerythrin conjugated anti-CD45RA and anti-CD29 and fluorescein conjugated anti-CD4 and anti-CD8 monoclonal antibodies, on peripheral blood lymphocytes from 19 patients with Behçet's disease (BD) and 18 healthy control subjects. The percentage of CD4+ cells was significantly lower in patients with BD (34 +/- 2%) than in control subjects (46 +/- 3%) (p less than 0.001). Among CD4+ cells, the percentage of suppressor-inducer (CD4+CD45RA+) cells was significantly lower in patients with BD (31 +/- 4%) than in control subjects (45 +/- 2%) (p less than 0.01), while the percentages of helper-inducer (CD4+CD29+) cells were similar in patients and controls. The percentage of CD8+ cells was significantly higher in patients with BD (36 +/- 2%) than in control subjects (26 +/- 2%) (p less than 0.001) involving both CD45RA+ and CD29+ subpopulations. Within CD4+ cells, the percentage of suppressor-inducer (CD4+CD45RA+) cells was significantly decreased in patients with active BD (26 +/- 3%) compared with control subjects (45 +/- 2%) (p less than 0.001), whereas in patients with inactive BD the difference was statistically insignificant. Our results suggest that the defective suppressive function in patients with active BD may be related to the decreased suppressor-inducer subpopulation (CD4+CD45RA+).

Adult↗

Expression of the Rho-family GTPase gene RHOF in lymphocyte subsets and malignant lymphomas.

We have studied the expression of RHOF, a member of the Rho-GTPase family, in an array of lymphoid cells and tissues. Previous microarray studies demonstrated RHOF upregulation in a subset of transformed follicular lymphomas. Real-time quantitative polymerase chain reaction evaluated RHOF expression in lymphocyte subpopulations, and normal and malignant lymphoid tissue. Cells and tissues of B-cell origin expressed higher RHOF levels than their T-cell counterparts. Neoplastic cells and tissues of B-cell origin expressed higher levels of RHOF than their benign cellular counterparts. Relatively elevated levels of RHOF were seen in lymphomas derived from germinal centre origin.

Analysis of Variance↗

Distribution of lymphocyte subsets in rat milk from normal and Trichinella spiralis-infected rats.

We have shown that T. spiralis-specific T lymphocytes can mediate maternal-to-neonatal immunity during lactation. This study addresses the change of lymphocyte populations in rat milk during normal and disease conditions. Two color flow cytometric analysis was performed for milk lymphocytes. T cells (OX19+) made up 45% of rat milk lymphocyte population. T helper cells (Th) composed 35% of total T cells while T cytotoxic/suppressor (Tcs) cells constituted 34%, giving a Th/Tcs ratio of 1.03. The corresponding ratio Th/Tcs in peripheral blood was 2.8. Approximately 21% of OX8+ cells in rat milk were OX19- natural killer (NK) cells. When using the monoclonal antibody 3.2.3 (NKR-P1), 43% of lymphocytes in control rat milk and 14% of blood lymphocytes were NK cells. This indicates a selective passage of these cells into milk. In T. spiralis-immunized rats, the percentage of total T cells was slightly decreased; however, Th and Tcs cells were consistent as compared to control milk. The percentage of NK cells (OX8+OX19-) in milk from T. spiralis-immunized rats was significantly higher than that from control milk (65% vs. 21%, respectively, P less than 0.01). This result was confirmed using the monoclonal antibody 3.2.3 which showed that milk from immunized rats contained 63% NK cells compared to 43% in normal milk (P less than 0.01). This study suggests that NK cells are selectively passaged into rat milk and T. spiralis infection induces an increase of NK cells in milk.

Animals↗

Cloning, characterization, and antigen specificity of T-lymphocyte subsets extracted from gingival tissue of chronic adult periodontitis patients.

Chronic periodontitis is characterized by dense infiltrations of B and T lymphocytes within the gingival connective tissue. Distinct anaerobic gram-negative bacteria as well as autoimmunity to collagen have been reported to play a role in the etiology and the pathogenesis of this disease. Here we describe the cloning and characterization of CD4+ and CD8+ T lymphocytes isolated from inflamed gingival tissue obtained from four patients with chronic periodontitis. Clones were raised with phytohemagglutinin and interleukin-2 and tested for proliferation in response to whole-cell antigens of Porphyromonas gingivalis, Prevotella intermedia, Actinobacillus actinomycetemcomitans, human collagen type I, and two bacterial heat shock proteins. CD4+ T-cell clones reactive with collagen type I were obtained from all four patients. Eighty percent of these clones had phenotypes resembling the mouse type 2 T helper (Th) phenotype, i.e., they produced high levels of interleukin-4 and low levels of gamma interferon. No collagen-type-I-reactive CD8+ clones were obtained. Bacterial-antigen-reactive CD4+ and/or CD8+ T-cell clones were also obtained from each patient, and the majority of the clones showed a Th0-like cytokine pattern and produced equal amounts of interleukin-4 and gamma interferon. Although most clones were reactive with P. intermedia, it seems that the immune response is not strictly directed against this particular microorganism, as clones reactive with one of the other bacteria were also obtained from two patients. We propose that collagen-specific CD4+ Th2-like T cells contribute to the chronicity of periodontitis but that their modes of activation might be controlled by Th0-like T cells specific for periodontitis-associated bacteria.

Adult↗

Effect of thymopentin on peripheral blood T-lymphocytes subsets and on the clinical course of the disease in patients affected by multiple sclerosis.

The effect on peripheral T-lymphocytes of Thymopentin (TP-5), asynthetic pentapeptide reproducing the biological activity of Thymopoietin, is known in Herpes Simplex infections and in Rheumatoid Arthritis. The aim of this study was to observe the effect of TP-5 on the OKT4 and OKT8 lymphocytes in Multiple Sclerosis. The AA. have studied this effect in two patients affected by definite MS, whose lymphocytes subpopulation, observed for 33 and 13.5 months respectively, showed a constant OKT4/OKT8 ratio greater than 2.5 in peripheral blood and whose clinical course was chronically progressive. TP-5 was administered during a period of one month. A decrease of the OKT4/OKT8 ratio in both patients (significant in one, p less than 0.01) due to the increase of OKT8 was observed. Also the clinical symptomatology improved in one patient.

Amino Acid Sequence↗

Epinephrine-induced changes in the distribution of lymphocyte subsets in peripheral blood of humans.

We have previously demonstrated that mitogen responsiveness of mononuclear cells (MNC) from peripheral blood is reduced after a single injection of epinephrine to human subjects. The purpose of the present study was to characterize the relative distributions of MNC subsets after epinephrine administration using monoclonal antibodies and conventional cell markers. The absolute number of circulating MNC increased 64% within 30 min after injection of epinephrine, and returned to baseline by 2 hr. Analysis of MNC subsets revealed that there were no changes in the relative percentages of total T lymphocytes [T3+ cells, or neuraminidase-treated sheep red blood cell rosettes (EN-rosettes)], B lymphocytes (B1+, or cells with surface-bound immunoglobulin), or monocytes (by morphologic criteria) after epinephrine administration. The percentage of inducer T cells (T4+) declined at 30 and 60 min postinjection. Overall, the percentage of suppressor/cytotoxic T cells (T8+) did not change after injection of epinephrine; however, analysis of individual subjects revealed opposing responses of this subset. The T4:T8 ratio was 2.19 before injection, declined to 1.56 at 60 min, then increased to 3.10 2 hr postinjection. The percentage of natural killer/killer cells (HNK-1+) increased from a baseline of 15.5% before epinephrine injection to 29.6% at 30 min postinjection, then declined to 11.4% at 2 hr. Therefore, the administration of physiologic doses of epinephrine results in changes in the relative proportions of lymphocyte subsets in peripheral blood, in addition to reduced mitogen responsiveness as reported previously.

B-Lymphocytes↗

Bovine leukemia virus-induced lymphocytosis and increased cell survival mainly involve the CD11b+ B-lymphocyte subset in sheep.

In this study, we show that bovine leukemia virus (BLV)-induced persistent lymphocytosis (PL) results from the in vivo expansion of the CD11b+ B-lymphocyte population. This subset shares phenotypic characteristics with murine and human B-1 cells. BLV interactions with the sheep B-1-like subset were explored. We found that B-1- and B-2-like cells are initially infected to similar extents. However, in long-term-infected sheep, the viral load is higher in B-1-like cells and only B-1- and not B-2-like cells show increased ex vivo survival compared to that in uninfected sheep. Ex vivo viral expression was found in both B-1- and B-2-like cells, indicating that both cell types support viral replication. Finally, cycloheximide and a protein kinase C inhibitor (H7) that blocks the ex vivo activation of viral expression did not affect the increased survival in B-1-like cells, suggesting that resistance to apoptosis is acquired in vivo. Collectively, these results indicate a peculiar susceptibility of sheep B-1-like cells to BLV transforming effects and further support the involvement of increased survival in BLV pathogenesis.

Animals↗

Phenotypic identification of T lymphocytes subset in recurrent aphthous stomatitis.

Recurrent aphthous stomatitis (RAS) is characterized by recurring ulcers in the oral mucosa as abnormality of the immune response, autoimmune or abnormal immunological reaction to antigens of oral bacteria. This study was conducted on 25 patients 15 suffering RAS of the minor form and 10 of the major form and 15 student as control. The T lymphocytes were determined by the monoclonal antibodies (OKT) using the indirect immunofluorescent technique of Hoffman percentage of cells having surface fluorescence was calculated and the monoclonal antibody coated cells visualized and enumerated. It was concluded from this study that the two clinical entities of RAS may be of the same immunopathologic etiology but the difference lies in the T cell subpopulation and the ratio of T helper to T suppressor T4/T8.

Adolescent↗

Lymphocyte subset and cellular immune responses to a brief experimental stressor.

To evaluate effects of acute mental stress on aspects of cellular immunity, lymphocyte populations and phytohemagglutinin (PHA)-stimulated T-cell mitogenesis were measured in 33 healthy young men, both before and immediately following subjects' performance of a frustrating, 21-minute laboratory task (Stroop test). Relative to baseline evaluations, post-task measurements showed a significant reduction in mitogenesis and alterations in various circulating lymphocyte populations; the latter included a diminished T-helper/T-suppressor cell ratio and an elevation in the number of natural killer cells. Eleven subjects assigned to a control (unstressed) condition exhibited no changes in lymphocyte populations, but did show an increase in T-cell proliferation, compared with pretask measurements.

Adult↗