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Flow cytometric analysis of lymphocyte subsets in the bronchoalveolar lavage fluid and peripheral blood of healthy volunteers.

The lymphocyte subsets in the bronchoalveolar lavage fluid (BALF) and the peripheral blood of 25 healthy volunteers were examined by analysis with a fluorescence-activated cell sorter. Comparison of the lymphocyte subsets in the BALF with those of the peripheral blood revealed much higher values for the ratios of each Leu 3a+ (CD4), Leu 3+8-, and Leu 2+15- cells, while the ratios of Leu 1+ (CD5), Leu 2a+ (CD8), Leu 7+, Leu 8+, Leu 10+, Leu 11a+ (CD16), Leu 12+, and Leu 2+15+ cells were low in the BALF. The above results indicate that the lymphocyte subsets in the BALF from healthy individuals are mainly composed of cells with surface phenotypes of helper T cells and cytotoxic T cells with virtual absence of cells carrying suppressor T and NK cell phenotypes, and with low B cell ratio. Therefore, it is assumed that the local immune mechanism of the lung is different from that of the peripheral blood.

Adult↗

Multicentre evaluation of stable reference whole blood for enumeration of lymphocyte subsets by flow cytometry.

BACKGROUND: Clinical indications for lymphocyte subset enumeration by flow cytometry include monitoring of disease progression and timing of therapeutic intervention in infection with human immunodeficiency virus. Until recently international standardisation has not been possible due to a lack of suitable stable reference material. METHODS: This study consisted of two trials of a stabilised whole blood preparation. Eleven participants were sent two standard protocols for staining plus gating strategy and asked to report absolute counts for lymphocyte subsets. RESULTS: No significant difference was detected between the two methods when results from the two assays and all partners were pooled. Significant differences in results from the different partners were observed. However, representative mean counts were obtained for geometric means, geometric coefficient of variation, and 95% confidence interval for CD3 910 cells/mul, 9%, and 888 to 933, respectively), CD4 (495 cells/mul, 12%, and 483 to 507), and CD8 (408 cells/mul, 13%, and 393 to 422). CONCLUSION: We have introduced a stabilised blood preparation and a well-characterized biological standard. The availability of this reference material greatly simplifies the validation of new techniques for CD4(+) T-cell enumeration and the expansion of external quality assurance programmes for clinical laboratories, including those that operate in resource-restricted environments. (c) 2005 Wiley-Liss, Inc.

Journal Article↗

T-lymphocyte subsets in HIV-infected and high-risk HIV-uninfected adolescents: retention of naive T lymphocytes in HIV-infected adolescents. The Adolescent Medicine HIV/AIDS Research Network.

BACKGROUND: The capacity of the immune system of adolescents to generate and repopulate naive and memory cell populations under conditions of normal homeostasis and human immunodeficiency virus (HIV) infection is largely unknown. OBJECTIVE: To assess lymphocyte subsets in HIV-infected and high-risk HIV-negative adolescents. DESIGN: The Reaching for Excellence in Adolescent Care and Health Project of the Adolescent Medicine HIV/AIDS Research Network recruits a cohort of HIV-infected and high-risk HIV-uninfected adolescents, aged 13 to 18 years 364 days, into a study of biomedical and behavioral features of HIV infection as seen in the context of full availability of primary care and HIV-related consultative services. Lymphocyte phenotypes were determined using standard 3-color flow cytometry. SETTING: The Reaching for Excellence in Adolescent Care and Health Project is carried out at 16 clinical sites in 14 urban areas. PARTICIPANTS: T-lymphocyte subsets are reported in 192 HIV-positive and 78 HIV-negative youths. RESULTS: For HIV-positive subjects, the total CD4+ cell count and the percentage of CD4+ cells are decreased when compared with those of the HIV-negative controls (P<.001). The reduction in total CD4+ cells reflects a loss of naive, and memory, CD4+ cells compared with HIV-negative youths. Human immunodeficiency virus-infected adolescents, many of whom have been infected recently (ie, those with CD4+ cell counts > or =0.500 x 10(9)/L [500/microL]), have a significant increase in naive CD8+ cells compared with HIV-negative youths (P<.01). There also is a significant increase in memory CD8+ cells at all strata of total CD4+ cells compared with HIV-negative youths (P<.01). The increase in naive CD8+ cells in those subjects with CD4+ cell counts of 0.500 x 10(9)/L or greater is a unique finding in this cohort. CONCLUSIONS: This study demonstrates high levels of naive CD8+ cells in response to HIV infection in adolescents with CD4+ cell counts of 0.500 X 10(9)/L or greater. The presence of high levels of naive CD8+ cells suggests functioning thymic tissue in some adolescents infected with HIV. Furthermore, the normal level of naive CD4+ cells in adolescents with CD4+ levels of 0.500 x 10(9)/L or greater provides additional support for the concept of a more robust immune system in HIV-infected adolescents compared with HIV-infected adults. These observations suggest that the immune system of HIV-infected adolescents may be capable of better responses to neoantigens and cytotoxic T-lymphocyte responses to HIV than the immune system of infected children or adults. Human immunodeficiency virus-infected adolescents may have an immune system that is capable of reconstitution following highly active antiretroviral therapy.

Adolescent↗

[Measurement of T-lymphocyte subsets in bronchial alveolar lavage fluid on smokers and patients with lung cancer].

The measurements of T-lymphocyte subsets (as expressed by, CD3 CD4 and CD8) in the peripheral blood and bronchial alveolar lavage fluid (BALF) on 30 cases of lung cancer, 26 cases of smoker and 25 cases of nonsmoker have been done. The results showed that there existed no differences in the peripheral blood of the above said subsets of lymphocyte. In BALF the percentage of CD3 in all the lymphocytes did not show any significant difference among the three tested groups, either (P > 0.05). But the percentage of CD4 and the ratio of CD4/CD8 manifested the following peculiarity: patients with lung cancers < smokers < nonsmokers (P < 0.01). On the contrary, the percentage of CD8 showed just the opposite: patients with lung cancer > smokers nonsmokers (P < 0.01). It was suggested that the cellular immunity as shown in the peripheral blood did not correspond with the findings in the BALF. The cellular immunity of the lungs was decreased both in patients with lung cancer and smokers, but more severely in the group of lung cancer patients. Smokers who have had changes of above said subsets of lymphocytes in the lungs may develop lung cancer. As the lung cancer advances, it may suppress body immunity and in turn enhance the cancer growth. Both the cause and the effect are interrelated. Therefore, the provocative agents for cellular immunity may act as one of the adjunctive therapies for patients with lung cancer. Giving up smoking should play an important role in recovering or promoting the immunity of the lungs and decreasing the incidence as well as improving the prognosis in patients with lung cancer.

Adult↗

Lymphocyte subset enumeration in HIV seronegative and HIV-1 seropositive adults in Dar es Salaam, Tanzania: determination of reference values in males and females and comparison of two flow cytometric methods.

The level of CD4(+) T-lymphocytes represents a useful marker with which to monitor the progression of HIV infection. Sex and geographical differences in the reference values of lymphocyte subsets have been reported. We have compared two flow cytometric methods (MultiSET and SimulSET) for the quantification of lymphocyte subsets using whole blood from 92 HIV seropositive and 241 seronegative adults, and determined the reference values of lymphocyte subsets in HIV seronegative Tanzanian subjects. In seronegative Tanzanian subjects, the percentages of CD3(+) and CD4(+) T-lymphocytes and the CD4(+):CD8(+) T-lymphocyte ratios were lower while the percentage of natural killer cells was higher compared to the levels of the corresponding parameters reported for Europeans. Seronegative Tanzanian females had significantly higher levels of CD3(+) and CD4(+) T-lymphocytes and CD4(+):CD8(+) T-lymphocyte ratios compared to seronegative males. The correlation coefficients of CD3(+), CD4(+) and CD8(+) T lymphocyte counts and percentages obtained by the two flow cytometric methods were high. The median values of the number of CD4(+) T-lymphocytes obtained by the two methods were not significantly different. In conclusion, determination of the reference values of lymphocyte subsets in HIV seronegative Tanzanian adults showed significant sex differences and differences in percentage values compared to those reported in certain other geographical areas. There was acceptable agreement in the levels of CD4(+) T-lymphocyte values obtained by the two flow cytometric methods.

Adult↗

[T-lymphocyte subsets in peripheral blood of patients with herpes simplex keratitis].

By indirect immunofluorescence assay, the authors examined the lymphocyte subsets in peripheral blood of patients with HSK and normal individuals, using monoclonal antibodies. The relation between the alterations of T-lymphocyte subsets and the mechanism of HSK relapse was discussed. It is postulated that the alterations of T-lymphocyte subsets may help guide the clinical treatment of HSK and estimate its prognosis.

Adolescent↗

Quantitative and qualitative modifications of lymphocyte subsets after sublingual administration of isosorbide dinitrate in migraineurs. Preliminary report.

In 22 migraine patients and 5 healthy controls an investigation has been performed on lymphocyte subsets in basal conditions and 90 minutes after sublingual isosorbide dinitrate (IDN) administration. In all patients the drug assumption induced a typical migraine attack with increase in all examined lymphocyte subsets with except of NK cells. A statistically significant increase (p < 0.05) in T lymphocyte subsets we found during the crisis, whereas in healthy controls neither a migraine attack nor any modification in lymphocyte subsets could be observed. These results seem to confirm the immune alteration we have previously found in migraineous patients and suggest the possible role of IDN-stimulated endothelium not only in vasodilatation but also in production of substances responsible of the observed immune modifications.

Administration, Sublingual↗

Zinc supplementation has no effect on circulating levels of peripheral blood leucocytes and lymphocyte subsets in healthy adult men.

As a result of evidence documenting harmful effects of Zn supplementation on immune function and Cu status, thirty-eight men were recruited onto a Zn supplementation trial. The aim was to examine the effects of chronic Zn supplementation on circulating levels of peripheral blood leucocytes and lymphocyte subsets. Subjects (n 19) took 30 mg Zn/d for 14 weeks followed by 3 mg Cu/d for 8 weeks to counteract adverse effects, if any, of Zn supplementation on immune status resulting from lowered Cu status. A control group (n 19) took placebo supplements for the duration of the trial. Dietary intakes of Zn approximated 10 mg/d. Blood samples, taken throughout the trial, were assessed for full blood profiles and flow cytometric analyses of lymphocyte subsets. Putative indices of Cu status were also examined. Results indicate that there was no effect of Zn supplementation on circulating levels of peripheral blood leucocytes or on lymphocyte subsets. Cu status was also unaltered. Independent of supplement, there appeared to be seasonal variations in selected lymphocyte subsets in both placebo and supplemented groups. Alterations in circulating levels of B cells (cluster of differentiation (CD) 19), memory T cells (CD45RO) and expression of the intracellular adhesion molecule-1 (CD54) on T cells were observed. Findings indicated no adverse effects of Zn supplementation on immune status or Cu status and support the US upper level of Zn tolerance of 40 mg/d. The seasonal variations observed in lymphocyte subsets in the group as a whole could have implications for seasonal variability in the incidence of infectious diseases.

Adult↗

Adenoidal and tonsillar lymphocyte subsets in AIDS children.

Tonsils and adenoids are secondary lymphoid organs exposed to the environment. The most important classifications of AIDS include the lymphocyte subsets of peripheral blood. We have studied the lymphocyte subsets in peripheral blood and secondary lymphoid organs in a control group of children suffering adenotonsillar pathology and in five children with AIDS and the same adenotonsillar pathology. The antigen surface markers were determined by flow cytometry in lymphocytes isolated from peripheral blood, and from tonsils and adenoids after tonsillectomy and adenoidectomy, in the control group and in children diagnosed with AIDS. The most important findings in tonsils and adenoids were a decrease of the total T lymphocytes, helper T lymphocytes and CD4/CD8 ratio; an increase of cytotoxic T lymphocytes and B lymphocytes, as well as a 200% increase in monocytes of AIDS-affected children. These observations show the value of analyzing the lymphocyte subsets of the tonsils and adenoids of AIDS-affected children, and establishing an earlier relation to clinical symptoms.

Acquired Immunodeficiency Syndrome↗

Identification of lymphocyte subsets in pregnancy gingivitis.

The percentage of gingival tissue and peripheral T, T Subsets and B lymphocytes were assessed in ten pregnant women during the first, second and third trimesters. Clinically the gingival index (GI) was found to be higher during pregnancy than in the control group. The percentage of the T3, T4 and B cells appeared to decrease in peripheral blood and gingival tissues during pregnancy as compared to control group.

Adult↗

Effects of chromium on lymphocyte subsets and immunoglobulins from normal population and exposed workers.

Blood lymphocyte subsets and serum immunoglobulins were studied in 15 men (mean age: 35 years), not exposed to toxic agents, and in 15 healthy men, exposed to dust containing several compounds (including lead chromate), working in a factory producing plastic materials. Worker blood lead and urine chromium (Cr) levels were significantly higher than controls, while serum Cr concentration was unchanged. In the worker blood, CD4+ helper-inducer (mainly CD4+-CD45RO- "virgin"), CD5--CD19+ B, CD3--CD25+ activated B and CD3--HLA-DR+ activated B and natural killer (NK) lymphocytes were significantly reduced (about 30-50 %). The investigated workers were exposed to hexavalent Cr, as lead chromate, whereas normal population (control group) was mainly exposed to trivalent Cr. In the control group, urinary Cr showed a significant positive correlation with CD16+-56+ NK, CD5+-CD19+ B and HLA-DR+ activated T, B and NK lymphocytes and a negative correlation with serum IgA immunoglobulins; moreover, serum Cr was significantly correlated with all blood lymphocytes and HLA-DR+, CD3--HLA--DR+ and CD3--CD25+ lymphocyte subsets. These data suggest that trivalent Cr may be involved in mechanisms regulating the immune response in humans.

Adult↗

Selective recruitment of T-cell subsets to the udder during staphylococcal and streptococcal mastitis: analysis of lymphocyte subsets and adhesion molecule expression.

During bacterial infection of the bovine mammary gland, large numbers of leukocytes migrate into the udder, resulting in the establishment of a host response against the pathogen. Currently, the specific leukocyte populations mediating this immune response are not well defined. In the studies described here, we analyzed blood and milk from healthy cows and cows with naturally occurring mastitis to determine if distinct alphabeta and gammadelta T-lymphocyte subsets were involved in the response of the udder to a mastitis pathogen and if the type of mastitis pathogen influenced the subset composition of these responding leukocytes. Although blood samples from cows with confirmed staphylococcal and streptococcal mastitis were characterized by increased numbers of gammadelta T cells, the most dramatic changes in leukocyte distributions occurred in milk samples from these cows, with a 75% increase in alphabeta T-cell levels and a 100% increase in gammadelta T-cell levels relative to the levels in milk samples from healthy animals. Interestingly, the increase in alphabeta T-cell numbers observed in milk from cows with staphylococcal mastitis was primarily due to increased numbers of CD4(+) T cells, while the increase in alphabeta T-cell numbers observed in cows with streptococcal mastitis was due to a parallel increase in both CD4(+) and CD8(+) T-cell numbers. The increased numbers of gammadelta T cells in milk from cows with staphylococcal and streptococcal mastitis were due to a selective recruitment of a distinct gammadelta T-cell subset (GD3.1(+)), while no change in the numbers of GD197(+) gammadelta T cells was observed. We also analyzed adhesion protein expression on blood and milk leukocytes and found that, in comparison to the situation for healthy cows, L-selectin was down-regulated and CD18 was up-regulated on leukocytes from cows with mastitis. Thus, shedding of L-selectin and up-regulation of CD18 by neutrophils may provide a sensitive indicator of early inflammatory responses during bovine mastitis. Overall, these studies suggest that distinct alphabeta and gammadelta T-cell subsets are involved in the host defense of the udder against mastitis infection and that selective recruitment of these T-cell subsets depends on the infectious agent involved.

Animals↗

[The difference of intracellular cytokines produced by different lymphocyte subsets of cord blood and adult peripheral blood].

OBJECTIVE: To explore the difference of intracellular cytokines between cord blood and adult peripheral blood and elucidate its significance for GVHD. METHOD: Different lymphocyte subsets were categorized in cord and adult peripheral blood by monoclonal antibodies with 3 color fluorescence using flow cytometry. The IL-2, TNF-alpha and IFN-gamma produced by various lymphocyte subsets were compared between the two groups. RESULT: (1) In cord blood, dominant T cells were CD(4)(+)CD(45RA)(+), while the major T cells were both CD(4)(+)CD(45RO)(+) and CD(8)(+)CD(45RO)(+) in adult peripheral blood. (2) The quantities of IL-2, TNF-alpha and IFN-gamma produced by cord blood lymphocytes were lower than that of adult peripheral blood lymphocytes. Moreover, the majority of cells that produced cytokines in the cord blood were CD(4)(+)CD(45RA)(+), but in adult peripheral blood were mainly CD(4)(+)CD(45RO)(+) and CD(8)(+)CD(45RO)(+). CONCLUSION: As compared with adult peripheral blood, the cord blood not only possessed different proportion of lymphocyte subsets, but also significant low level of cytokines. The differences of various lymphocyte subset proportions and lower level of cytokines in cord blood probably related to the lower incidence of GVHD in cord blood stem cell transplantation.

Adult↗

Functional studies of T4+ lymphocyte subsets distinguished by the monoclonal antibody 5/9: suppressor-effector T4+ lymphocytes derive from the 5/9+ subset.

Human T lymphocytes bearing the cell surface antigen T4 are functionally heterogeneous, exerting helper/inducer, suppressor-inducer, suppressor-effector, and cytotoxic activities. Other cell surface antigens with a more restricted expression may help separate T4+ lymphocytes into functionally distinct subsets. This report describes the regulatory functions of T4+ lymphocytes fractionated by the monoclonal antibody 5/9, which detects a cell surface antigen present on 50-60% of T4+ lymphocytes. The results indicate that both 5/9+ and 5/9- T4 subsets contain helper/inducer and suppressor-inducer cells. Suppressor-effector activity, however, is found predominantly within the 5/9+ T4 subset. The 5/9 antibody thus identifies the suppressor-effector subset of T4+ lymphocytes, although it does not distinguish between T4+ cells with or without helper/inducer and suppressor-inducer functions.

Antibodies, Monoclonal↗

Lymphocyte subsets and natural killer cell cytotoxicity after laparoscopically assisted resection of rectosigmoid carcinoma.

BACKGROUND: Laparoscopically assisted resection of colorectal carcinoma is technically feasible and minimally invasive. Postoperative immunosuppression also may be reduced. This study compared the lymphocyte subsets and natural killer (NK) cell cytotoxicity in patients after laparoscopically assisted resection with those after open resection of rectosigmoid carcinoma. METHODS: In this study, 40 patients with rectosigmoid carcinoma, but no evidence of metastasis, were randomized to receive either laparoscopically assisted or conventional open resection of the tumor. Blood was collected before the operation, then 24 h, 72 h, and 8 days after the operation for studies of lymphocyte subsets and NK cell cytotoxicity. RESULTS: The lymphocyte subsets and NK cell cytotoxicity of both groups showed typical suppression after surgery. The suppression of T cell activation and NK-like T cells was significantly less after laparoscopically assisted resection than in after open resection, whereas the difference in other lymphocyte subsets and NK cell cytotoxicity was not significant. CONCLUSION: This study showed that some cellular components of the immune system are less suppressed after laparoscopically assisted than after conventional open resection of rectosigmoid carcinoma. This may have implications for tumor recurrence and long-term patient survival.

Aged↗

[NK cell activity and counts of T-lymphocyte subsets in pulmonary tuberculosis].

NK cell activity and counts of T lymphocyte subsets examined in 42 pulmonary tuberculosis cases and 32 sex- and age-matched healthy controls. The mean NK cell activity in patients (62.78 + 27.54) were significantly higher than that in healthy controls (39.39 + 12.01) While counts of CD4+T cells and CD3+T cells in patients (35.5 + 9.3, 51.5 + 9.4) were lower than that in controls (43.8 + 9.0, 65.6 + 11.1). The CD4+T cell/CD8+T cell ratio had no significant change both in patients and in controls. Nk cell activity increased with the progression of tuberculosis lesion in X-ray. Counts of T lymphocyte subsets were unrelated with the severity and the extent of the lesion radiologically.

Adult↗

Peripheral blood lymphocyte subsets after allogeneic bone marrow transplantation: reconstitution and correlation with the occurrence of acute graft-versus-host disease.

Peripheral blood lymphocyte subsets were enumerated at regular intervals during the first year after allogeneic bone marrow transplantation (BMT) in 21 Chinese patients. Eight of these patients had acute graft-versus-host disease (GVHD) while they were assessed at the time of engraftment. Our results show in patients receiving allogeneic BMT: (1) T and NK cells were the predominant lymphocyte subsets in the early reconstitution stage while B cells were severely depleted; (2) absolute numbers of the major lymphocyte subsets normalised in 4-5 months; (3) an increased percentage of T cells that expressed the activation antigen HLA-DR and a reversed CD4:CD8 ratio were observed throughout the first 12 months after BMT; (4) patients with acute GVHD had significantly higher white cell count and NK cell percentage than those not complicated by acute GVHD.

Acute Disease↗