[The effect of UFT on lymphocyte subsets and lymphocyte response to PHA].
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Antineoplastic Combined Chemotherapy Protocols↗
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Ultrastructure of lymphocyte subsets and of activated lymphocytes in type 1 diabetes as defined by monoclonal antibodies and the immunogold technique.
The ultrastructure of peripheral blood lymphocyte subsets and activated lymphocytes from 5 patients with recent onset insulin-dependent diabetes as identified by monoclonal antibodies (CD4, CD8 and 4F2) and labelled with gold coupled goat anti-mouse IgG are described and depicted. Electron microscopy revealed no differences in appearance between investigated lymphocyte subsets at the single cell level. Activated lymphocytes as defined by an early activation antigen (4F2) do not always have a characteristic appearance nor do they show morphological signs of activation in all cases. We would conclude that it is not possible to recognize different lymphocyte subsets based only on their ultrastructure.
Antibodies, Monoclonal↗
Expression and release of IL-2 receptor and production of IL-2 by activated T lymphocyte subsets.
T lymphocyte subsets differ in expression of cell surface antigen and functional properties. Both CD4+ and CD8+ subsets express interleukin-2 receptor (IL-2R) following their activation in vitro. In the present investigation T lymphocyte subsets were activated by different mitogens and IL-2R expression was enumerated on these stimulated subsets. Peripheral blood mononucleated cells (PBMC) were stimulated with phytohaemagglutinin (PHA) and pokeweed mitogen (PWM) and then stained with anti-CD4 or anti-CD8 antibodies conjugated with fluorescein isothiocyanate and anti-IL-2R monoclonal antibody conjugated with phycoerythrin using a direct immunofluorescence technique. The percentage of IL-2R positive lymphocytes was enumerated by flow cytometry. The results showed that mitogen activated lymphocytes expressed variable degrees of IL-2R which were significantly higher than the control. 53% of CD4+ lymphocytes and 28% of CD8+ expressed IL-2R following PHA stimulation in vitro. Similarly, 47% of CD4+ lymphocytes and 23% of CD8+ lymphocytes expressed IL-2R following PWM stimulation. The present study also revealed that the release of soluble IL-2R (sIL-2R) and IL-2 production in supernatant from cultured PBMC varied with different mitogen stimulation. Using the same concentration of PHA and PWM as used to study IL-2R expression, higher activity of sIL-2R was detected in PHA stimulated lymphocytes as compared to PWM treated lymphocytes. However, IL-2 production was more in culture medium from PMW treated PBMC. Thus, there was a significant correlation between the cellular and soluble IL-2R but the production of IL-2 from activated PBMC cells had no good correlation with either the cellular IL-2R expression or the release of sIL-2R.(ABSTRACT TRUNCATED AT 250 WORDS)
Adolescent↗
Serum immunoglobulins and lymphocyte subsets in chronic lymphocytic leukemia.
The concentrations of serum immunoglobulins were correlated to the stage of disease and the proportions of peripheral blood lymphocyte subsets in 25 untreated patients with B-cell chronic lymphocytic leukemia (CLL). Diminished levels of at least one serum immunoglobulin were present in 77% of all patients with CLL and 73% of patients with Stage 0 disease. The mean concentration of IgG, IgM, and IgA decreased with advancing stage of CLL. The percentages of total T, T-helper (TH), and T-suppressor (TS) cells in the peripheral blood were less in patients with CLL than in healthy persons, but the absolute concentrations of total T, TH, and TS cells were greater in patients with CLL than controls (P less than 0.02). The absolute number of B-cells (P less than 0.01) and null cells (P less than 0.001) was also increased in patients with CLL, particularly those patients in advanced stages of CLL. These findings suggest that the hypogammaglobulinemia associated with CLL first occurs during the earliest stage of disease and may be related to the alterations in the proportion of peripheral blood lymphocytes.
Adult↗
Concentration-dependent effects of prednisolone on lymphocyte subsets and mixed lymphocyte culture in humans.
In a previous study the feasibility of pharmacokinetic-pharmacodynamic modeling for a quantitative description of the lymphocytopenic effect of prednisolone was demonstrated. We now applied this technique to compare the lymphocytopenia of T-lymphocyte subsets, namely CD8 and CD4. The finding of similar rate constants for the delay of the effect on different T-lymphocyte categories supports the explanation of this delay on the basis of pharmacokinetics rather than cellkinetics. The time course of the responsiveness of remaining lymphocytes in mixed lymphocyte culture after prednisolone administration could be described with the same model as the lymphocytopenia. The concurrence of both effects suggests that total lymphocyte counts, if considered intraindividually, could be used as a measure for monitoring the indirect immunosuppressive effect of prednisolone. The inhibitory effect on mixed lymphocyte culture of plasma from subjects who received prednisolone was directly related with the prednisolone concentrations in plasma. Just as for the indirect effect, a threshold concentration could be observed in the concerning concentration-effect relation. This is attributed to the decrease of endogenous hydrocortisone levels under the influence of prednisolone. Possible consequences of our results and those from related studies for the use of prednisolone as an immunosuppressive drug are discussed.
Adult↗
Immunologic identification of lymphocyte subsets in experimental murine myocarditis with encephalomyocarditis virus. Different kinetics of lymphocyte subsets between the heart and the peripheral blood, and significance of Thy 1.2+ (pan T) and Lyt 1+, 23+ (immature T) subsets in the development of myocarditis.
To clarify the immune mechanism in myocarditis, immunofluorescence techniques with laser flow cytometry were used to examine serial changes in lymphocyte subsets in the heart, spleen, and peripheral blood of DBA/2 and BALB/c mice inoculated with encephalomyocarditis virus (Experiment I). B cells were identified by staining with fluorescein isothiocyanate-labelled rabbit anti-mouse immunoglobulin. T-cell subsets were identified with rat anti-Thy 1.2, and nonpolymorphic Lyt 1 and Lyt 2 monoclonal antibodies plus fluorescein isothiocyanate-labelled anti-mouse immunoglobulin. On days 7 and 14 postinfection, the percentage of Thy 1.2+ (pan T) cells in both strains had decreased in the peripheral blood; B cells showed no significant changes throughout the entire period. On the other hand, Thy 1.2+ (pan T) and Lyt 1+, 23+ (precursor and immature) T cells appeared to occupy the major portion of the myocardium on days 7 and 14 when congestive heart failure developed. To confirm this, serial immunohistologic studies (immunoperoxidase staining) of the hearts of DBA/2 and BALB/c mice with encephalomyocarditis virus-induced myocarditis were performed (Experiment II). In Experiment II, most of the stained cells in the hearts of both strains were Thy 1.2 positive and Lyt 1 and Lyt 2 positive on days 7 and 14. Thus, Experiments I and II demonstrated that lymphocytes at the site of inflammation in acute viral myocarditis carried antigenic markers that differed from those of peripheral lymphocytes and suggested that Thy 1.2+ (pan T) cells, especially the Lyt 1+, 23+ subset (immature T cells and T-cell subset precursors) were involved in the development of myocarditis in these animals.
Animals↗
Serial immunologic identification of lymphocyte subsets in murine coxsackievirus B3 myocarditis: different kinetics and significance of lymphocyte subsets in the heart and in peripheral blood.
To elucidate possible immune mechanisms in the pathogenesis of myocarditis, we examined, by immunofluorescence techniques, the serial changes in lymphocyte subsets in the heart, spleen, and peripheral blood of C3H/He mice inoculated coxsackievirus B3 (experiment I). B cells were identified by staining with fluorescein isothiocyanate-labeled rabbit antimouse immunoglobulin. T cell subsets were demonstrated with rat anti-Thy 1.2, anti-Lyt 1, anti-Lyt 2, and anti-L3T4 monoclonal antibodies, respectively, plus fluorescein isothiocyanate-labeled antimouse immunoglobin. The percent of Thy 1.2+, Lyt 1+, and L3T4+ cells was decreased in the peripheral blood on days 3, 7, and 14. B cells were increased on day 3. In contrast, Thy 1.2+ (pan T) or Lyt 1+ cells appeared to occupy the major portion of the myocardium on days 7 and 14, when myocarditis was most severe, while B cells were sparse. For confirmation, serial immunohistologic studies (immunoperoxidase staining) of the hearts of C3H/He mice with coxsackievirus B3 myocarditis were performed (experiment II). Most of the stained cells in the heart were Thy 1.2, Lyt 1, and Lyt 2 positive on days 7 and 14. Thus, independent methods demonstrate that specific antigenic markers on lymphocytes at the site of inflammation in the acute stage of viral myocarditis differ from those on corresponding peripheral lymphocytes, and suggest the possible involvement of Thy 1.2+ (pan T) cells, mainly the Lyt 1+ and Lyt 2+ subsets (immature T cells), in the development of myocarditis in this preparation.
Animals↗
[Laboratory tests for cellular immunology: flow cytometric analysis of lymphocyte subsets].
Flow cytometric analysis for lymphocyte subsets provides two kinds of data: proportion of fluorescence-positive cells (lymphocyte subsets) and the fluorescence intensity. For proportional analysis of lymphocyte subsets, lymphocytes are fractionated with a conventional gate window in scatter analysis, but if activated or enlarged lymphocytes are examined, lymphocytes should be fractionated with an extended gate window. On the other hand, fluorescence intensity of a lymphocyte subset, which represents density of cell surface antigen detected by fluorescence-conjugated specific antibody, is not measured in routine tests yet. However, data on fluorescence intensity are often more valuable than the proportions of lymphocyte subsets: e.g., decrease in fluorescence intensity, but not in proportion, of CD8+ cells in active Graves' disease, and marked decrease in fluorescence intensity of OKT4+ cells in a carrier of familial OKT 4 epitope deficiency. Furthermore, two-color flow cytometry has enabled to measure more valuable, classified subsets of lymphocytes than single color flow cytometry. For example, measurement of peripheral CD5 (Leu-1)+ B cells is very useful for diagnosis of Graves' disease and its differentiation from destruction-induced thyrotoxicosis. In conclusion, we showed that measurement of fluorescence intensity of lymphocyte subsets and two-color flow cytometry are useful for laboratory tests of lymphocyte subsets.
Flow Cytometry↗
[Effect of splenectomy in gastric carcinoma surgery on cell-mediated immunity; evaluation with alteration of peripheral blood lymphocyte subsets].
Peripheral blood lymphocyte subsets were studied in 70 patients who underwent gastrectomy for gastric carcinoma. Lymphocyte subsets were analysed before and after gastrectomy by use of monoclonal antibody for cell membrane markers. There were significant differences in the gastrectomized patients with splenectomy compared to the patients without splenectomy in postoperative changes of lymphocyte subsets as follows. Decrease of OKT4 and increase of OKT8. Temporary decrease of B-1. Continuous increase of Leu-7. These results suggest that gastrectomized patients with splenectomy show a decrease in helper activity, an increase of suppressor activity of T lymphocytes and a compensatory increase of population of natural killer cells in association with suppression of natural killer activity.
Female↗
Lymphocyte subsets, phytohaemagglutinin responsiveness of blood lymphocytes, and interleukin 2 production in sarcoidosis.
To test the possibility that T lymphocyte subset imbalance and interleukin 2 (IL2) play a part in the impairment of cellular immune response in sarcoidosis, the proportion of T lymphocyte subsets in peripheral blood and alveolar lavage fluid from 21 patients with sarcoidosis was studied, monoclonal antibodies OKT3, OKT4, and OKT8 being used. Lectin induced production of IL2 and phytohaemagglutinin (PHA) responsiveness of peripheral blood lymphocytes were investigated. The percentage of both OKT3+ and OKT4+ T lymphocytes was significantly lower in peripheral blood from patients with sarcoidosis than in control subjects (control 63% and 46%), more so in patients with chronic sarcoidosis (44% and 23%) than in patients with recent sarcoidosis (56% and 38%). PHA induced IL2 production from peripheral blood lymphocytes did not differ between patients with sarcoidosis and control subjects. There was a significant positive correlation between PHA responsiveness and the percentage of blood OKT3+ and OKT4+ cells. Peripheral blood lymphocyte PHA responsiveness was decreased only in patients with an OKT4/OKT8 ratio of less than 1.5. Finally, late addition of exogenous IL2 to the culture medium on day 5 increased 3(H)Tdr incorporation by PHA stimulated blasts in peripheral blood lymphocytes from normal subjects, but not from those of patients with sarcoidosis. The data suggest that the impairment of cellular immune response in patients with sarcoidosis could in part reflect a decrease in the percentage of blood T helper lymphocytes and impaired IL2 receptors at the surface of stimulated lymphocytes.
Adult↗
T lymphocyte subsets and B lymphocytes in tubercular pleural effusion.
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Adult↗
Stressor-induced changes in mitogenic activity are not associated with decreased interleukin 2 production or changes in lymphocyte subsets.
Splenic lymphocytes from Lewis rats that received presentations of physically aversive electric shock demonstrated a marked reduction in responsiveness to T-cell mitogens such as concanavalin A. This study examined cellular mechanisms which may be responsible for this functional alteration. There was no difference in distribution of T-cell subsets from shocked and nonshocked rats. There was no difference in the production of interleukin 2 (IL-2) nor was there a difference in the percentage of IL-2 receptor positive T cells or T-cell subsets after culture for 24 hr. However, there was a marked lack of mitogenic stimulation in splenocytes from shocked rats when stimulated with the calcium ionophore A23187. This indicates a defect in the biochemical pathways necessary to activate T-cell mitogenesis.
Animals↗
Histological analysis of lymphocyte subsets infiltrated into mouse tumor tissue exposed to local irradiation.
The subsets of lymphocytes infiltrated into mouse tumor tissue exposed to irradiation were analyzed by the avidin-biotin-peroxidase procedure using monoclonal antibodies. The effect of PSK administration was also analyzed. Microscopic observation revealed the intense lymphocytic reactions in the exposed tumor, especially 7 days after irradiation of 20 Gy, which mainly consisted of anti-L3T4 positive cells, whereas the degree of infiltration was lower in the nonexposed tumor. PSK was found to enhance T lymphocyte subset infiltration into the exposed tumor tissue.
Animals↗
T-lymphocyte subsets: major histocompatibility antigens rejection of H-2K or H-2I region disparate skin allografts by either Lyt-2+ or L3T4+ T-lymphocyte subset.
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Animals↗
Analysis of responding lymphocyte subsets in allogeneic mixed lymphocyte cultures in the neonates and adults.
An analysis of proliferating cell subsets in the allogeneic mixed lymphocyte culture (MLC) in adults (n = 19) and newborns (n = 12) was performed by a method combining immunoperoxidase staining and autoradiography. The main cell type responding in adult cells were OKT8-positive (38%), whereas in neonates OKT4-positive cells (31%) predominated. Furthermore, in the neonatal allogeneic MLC there were significantly more 3H-thymidine incorporating Leu 12-positive B cells than in adults (12% vs 4%).
Adult↗
Detection of lymphocyte subsets using three-color/single-laser flow cytometry and the fluorescent dye peridinin chlorophyll-alpha protein.
The fluorescent dye, Peridinin chlorophyll A protein (PerCP) derived from dinoflagellate organisms (Glenodinium sp.) can be excited by a 488 nm laser and emits light with a large Stokes shift and no major spectral overlap with commonly used chromophores such as fluorescein isothiocyanate (FITC) and R-phycoerythrin (PE). PerCP was conjugated directly to various mouse monoclonal antibodies (mAb) specific for human leukocyte markers or to avidin for use with biotinylated-mAb, and used to perform three color single-laser flow cytometry. The efficacy of this method was demonstrated by analyzing the heterogeneity of thymus T lineage subsets and B lymphocyte subsets in blood. CD4-CD8-, CD4+CD8+ and CD4+CD8- or CD4-CD8+ subsets differ in their expression of cell-cell interaction markers including CD18, CD28, CD44 and Leu 8, and activation/subset markers CD45RO, CD45RA and CD26. Some CD5+ peripheral blood B cells, unlike CD5-B cells, expressed CD45RO or high levels of CD54 (ICAM-1) suggesting the CD5+ B cell population contains activated lymphocytes. The availability of such an accessible method for three color analysis will make it possible to do routine three color monitoring of immunologic diseases such as AIDS, and autoimmune or periodontal diseases.
Antibodies, Monoclonal↗
Flow cytometric monitoring of human immunodeficiency virus-infected patients. Simultaneous enumeration of five lymphocyte subsets.
The utility of CD4 lymphocytes in monitoring disease progression and prognosis of human immunodeficiency virus (HIV)-infected patients is well established. We have modified a previously described antibody cocktail to provide complete lymphocyte subset analysis on 100-200-microL samples of whole blood. This method optimizes accuracy of CD4 lymphocyte assessments and provides simultaneous assessment of four other lymphocyte subtypes of interest in specimens with absolute lymphocyte counts as low as 300 X 10(6)/L. Lymphocytes are classified as Thelper (CD3+CD4+); Tsuppressor (CD3+CD8+); Tnull (CD3+CD4-CD8-, putative gamma delta T-cell receptor); B (CD19+CD20+); or natural killer (CD3-CD16+CD56+). The method positively discriminates against contamination of lymphocyte scatter gates by monocytes and unlysed erythrocytes and is compatible with a variety of cell preparation procedures. Increased accuracy of CD4 lymphocyte determinations and simultaneous identification of other lymphocyte subsets whose relationship to disease progression is under study make this an efficient and informative method for disease monitoring and evaluation of therapy in HIV-infected patients.
Antibodies, Monoclonal↗
Helper-inducer and suppressor-inducer lymphocyte subsets in alcoholic cirrhosis.
Peripheral blood lymphocytes from patients with alcoholic cirrhosis, alcohol-induced fatty liver, and healthy controls were analyzed for helper-inducer (CD4+CD29w+) and suppressor-inducer (CD4+CD45R+) T lymphocytes. In confirmation of earlier reports, patients with alcoholic cirrhosis were found to have a significantly reduced absolute number of peripheral lymphocytes (p = 0.03), an elevated relative percentage of CD3+ cells (median, 76% versus 68%; p = 0.0004) and CD4+ T cells (median, 56% versus 51%; p = 0.0011), and a reduced percentage of CD8+ T lymphocytes (median, 11% versus 20%; p = 0.0007) as compared with the control group. No difference in lymphocyte subsets was observed between controls and patients with alcohol-induced fatty liver. Within the CD4+ T-cell population a change in the relative proportion of two complementary lymphocyte subsets (CD4+CD29+ helper-inducer and CD4+CD45R+ suppressor-inducer T cells) was observed in patients with alcoholic cirrhosis: a higher percentage of CD4+CD29w+ helper-inducer T cells were circulating in their peripheral blood than in healthy controls (median, 33% versus 28%; p = 0.0036), whereas the CD4+CD45R+ suppressor-inducer T-cell subset did not differ (median, 21% versus 21%) between the two groups. Owing to the reduction of lymphocyte counts in cirrhotic patients the absolute number of CD4+CD29w+ cells was not different from that of control individuals; however, CD4+CD45R+ T cells in peripheral blood (p = 0.0063) were absolutely reduced. More CD4+ cells were simultaneously CD29w+ in cirrhotic patients (61%) than in controls (52%), whereas a lower percentage of CD4+ lymphocytes was also CD45R+ in these patients (33%) as compared with controls (40%).(ABSTRACT TRUNCATED AT 250 WORDS)
Adult↗