Lymphocyte subpopulations in the neonate: identification of an immature subset of OKT8-positive, OKT3-negative cells

T cell subpopulations of lymphocytes from cord blood (CBL) of 24 newborns and from peripheral blood (a-PBL) of 24 healthy adult volunteers were assessed in T cell-enriched, T cell depleted and unseparated lymphocyte fractions by using OKT3, 4, 6, and 8 monoclonal antibodies. The results show that T cell-enriched CBL include adult numbers of OKT3+, OKT4+, OKT6+ and OKT8+ lymphocytes whereas the T cell-depleted fraction consists of a high percentage of OKT8+, OKT3-, non-E rosette-forming cells bearing a PNA receptor. The presence of the PNA receptor and the lack of the OKT3+ antigen strongly support the hypothesis that the subset of OKT8+ cells in cord blood includes immature T lymphocytes that may represent an intermediate stage between thymocytes and mature peripheral T cells.     

Reversal of cell surface abnormalities of T lymphocytes in hodgkin's disease after in vitro incubation in fetal sera.

The capacity of periphal blood lymphocytes from patients with untreated Hodgkin's disease to form E rosettes with sheep erythrocytes and to respond in vitro to PHA stimulation were found to be profoundly impaired. In 49% of the patients, the percentage of E rosette-forming cells (E-RFC) was more than two standard deviations below the mean for normal donors. Overnight incubation of the peripheral blood lymhocytes from these patients in culture media containing 20% fetal calf serum was followed by restoration of the percentage of E-RFC up to normal levels. Similar results have been observed after incubation in fetal human serum, but not in adult human AB serum or adult bovine serum. Incubation of peripheral blood lymphocytes from untreated patients in20% fetal calf serum also resulted in a remarkable restoration of their capacity to respond normally to PHA. Possible mechanisms involved in these reversible cell surface and in vitro lymphocyte function abnormalities in Hodgkin's disease are discussed.     

The cellular immune response in different forms of diphtheria in adults

In 109 adult diphtheria patients with different forms of the disease (toxic, subtoxic, localized) and carriers, the latter including 6 persons having had the localized form of diphtheria, 12 different subpopulations of T and B lymphocytes and neutrophils were studied by the methods of rosette formation with sheep, mouse and bovine red blood cells. The study showed that, starting from week 2 of the disease, an increase in the content of the subpopulations under study was registered in all forms of diphtheria; the disease, as well as in patients with the localized form of the disease accompanied by complications and in long-term carriers. In all forms of diphtheria a decrease in the content of neutrophil rosette-forming cells was shown. The marker tests for prognostication of the complicated course of the disease, viz. the elevated content of M-rosette-forming cell and T gamma-rosette-forming cells during the first week of then disease, were established.     

The neutrophil receptor apparatus in typhoid fever

In 157 typhoid fever patients and 36 practically healthy persons the content of neutrophils forming complement-dependent rosettes (NEAC rosette-forming cells), as well as rosettes with sheep red blood cells (NE rosette-forming cells) and with typhoid erythrocyte diagnosticum (NS rosette-forming cells), has been studied. The data obtained in this investigation indicate that antigen-binding neutrophils play an active functional role in the pathogenesis of typhoid fever at its acute stage. The values characterizing the content of N rosette-forming cells may be used as a criterion for the evaluation of the severity of infection, as well as for the prognostication of complications and relapses.     

Surface characteristics of thymocytes in glucocorticoid treated rats using rosette-formation technique and surface marker analysis.

Glucocorticoid (GC) treatment is known to induce destruction of cortical thymocytes and then their reconstitution. By using the rats treated with GC, we examined the relationship between rosette-formation and surface markers (CD4 and CD8) for clarifying the processes of differentiation and maturation in rat thymocytes. Thymus weight and thymocyte count began to decrease immediately after GC administration and became minimal on 5-7 days, followed gradual recovery. The percentage of rosette-forming thymocytes began to decrease immediately after GC treatment and became minimal on 5 days, followed by recovery to the normal level by the 10th to 14th day after treatment. During the analysis of the changes in the percentage of 4 subsets (CD4-8-, CD4+8+, CD4+8+, CD4-8+) of rat thymocytes after GC treatment, the percentage of CD4+8+ cells was found to change in close relation to the change in the percentage of rosette-forming lymphocytes, suggesting that rosette-forming thymocytes are CD4+8+ cells. These results suggest that the treatment induces destruction of GC-sensitive thymocytes, possibly rosette-forming cells, followed by migration of precursor T cells (CD4-8- cells) in the thymus, and that the precursors change into rosette-forming cells (CD4+8+ cells) in the thymus, followed by differentiation and maturation into non-rosette-forming cells (CD4+8- or CD4-8+ cells).     

Myelin basic protein-stimulated rosette-forming T cells in multiple sclerosis

A subpopulation of T lymphocytes sensitized to human myelin basic protein in peripheral blood of patients with multiple sclerosis, central nervous system (CNS) tumors, and cerebrovascular accidents was demonstrated by the antigen-stimulated, rosette-forming T cell assay. A significant increase in the percent of active rosette-forming T cells was detected after in vitro exposure of peripheral blood lymphocytes to human myelin basic protein but not to histones. In contrast, peripheral blood lymphocytes from healthy controls and from patients with benign and malignant breast diseases were unresponsive to stimulation by either antigen. These results demonstrate a functionally active T-lymphocyte subpopulation sensitized to myelin basic protein in patients with multiple sclerosis and in patients with certain other CNS diseases.     

Evaluation of three E-rosette assays in melanoma patients

Summary The percentage of E-rosette-forming cells (E-RFC) was determined repeatedly in the peripheral blood of 43 patients with malignant melanoma. Three methods of E-rosette assaying were used: total E-rosette test, active E-rosette test, and 29° C E-rosette assay.Depressed E-RFC values were found in fewer assays than normal values. The mean values of E-RFC and the proportion of depressed E-RFC in patients in stage I did not differ from the values in healthy control persons whatever method of assay was used.The frequency of depressed E-RFC values was higher in advanced stages. Significant differences were demonstrated between stage I and stages II and III combined in the values for total and for active E-rosettes (P<0.01).The active E-rosette test and the 29° C E-rosette assay gave no more positive results (i.e., depressed E-RFC values) than the total E-rosette test in melanoma patients.     

T- and B-lymphocytes of guinea pigs sensitized to ragweed pollen]

Functional activity of T and B lymphocytes in guinea pigs sensitized with common ragweed pollen was investigated. The content of T and B rosette-forming cells (T RFC and B RFC) was studied in guinea pigs following sensitization with common ragweed pollen. It was shown that the amount of B RFC in the regional lymph nodes at the early period of sensitization was over 4 times greater than that of the B RFC in normal animals. Functional capacity of T cells during the sensitization determined in the rosette-formation test altered less than the analogous capacity of B cells.     

Natural cytotoxic reactivity of human lymphocytes against a myeloid cell line: characterization of effector cells.

Using a series of techniques to identify and deplete various peripheral blood lymphocyte subpopulations, we studied the cytotoxic reactivity of normal individuals against the myeloid cell line K-562 in a 4-hr 51chromium-release assay. Depletion of lymphocytes bearing complement receptors had a variable, usually negligible effect on cytotoxicity. In contrast, depletion of lymphocytes bearing Fc receptors abrogated target cell lysis. Separation of lymphocytes with high-affinity binding of sheep red blood cells (SRBC) evidenced by rosette formation at 29 degrees C yielded a population of rosette-forming cells containing few cytotoxic cells, whereas separation of total E-RFC under optimal rosetting conditions produced a rosette fraction containing a major proportion of the effector cells. These data indicate that the cytotoxic lymphocyte in this system is Fc receptor positive, largely complement receptor negative, and may possess low density or low affinity receptors for SRBC.     

Inhibition of rosette formation by antithymocyte globulin

Summary The peripheral blood leukocytes from 29 patients with adenocarcinoma of the colon were studied sequentially for the T-cell level, the rosette-inhibition titer of antithymocyte globulin and the blastogenic response to PHA and Con A to evaluate the T-cell immunocompetence. The level of E-rosette-forming cells and the blastogenic response did not reflect the immunocompetence of the T-cell population. Fluctuations in the rosette-inhibition titer of antithymocyte globulin (ATG) were observed; an increased ATG titer indicated an unfavorable clinical course, while a decreased ATG titer was observed with patients who had a favorable clinical course. The rosette-inhibition titer with antithymocyte globulin was observed to be an important indicator of competent T cells in patients with colorectal cancer.Supported by the Physicians' Medical Education and Research Foundation and General Research Supports FundsAbbreviations used that are not spelled out in the text are: AET 2-aminoethylisothiouronium bromide; ATG Antithymocyte globulin; ALG Antilymphocyte globulin; E-RFC Erythrocyte rosette-forming cells; PHA Phytohemagglutinin; Con A Concanavalin A; E Sheep erythrocyte     

Changes in the ratio of T-lymphocyte subpopulations in various cytological samples of Plummer's thyroid adenoma

In order to correlate possible alterations of cell-mediated immune response with the evolutive phases of Plummer Adenoma (P. A.), T lymphocytes subpopulations in FNA samples and in peripheral blood lymphocytes (PBL) have been studied in 5 patients with autonomous nodules. The lymphocyte component in FNA and peripheral blood has been isolated by Lymphoprep gradient centrifugation; the analysis of T helper and T suppressor subpopulations was made by indirect immunofluorescence with OKT8 and OKT4 monoclonal antibodies. Our results show a reduction in OKT4/OKT8 ratio in cytological samples compared with PBL in patients with P. A., while in control subjects there was not statistically significant difference. In the patients with P. A., the relative increase of OKT8 lymphocytes in FNA compared with PBL is correlated with the functional state, that is toxic adenomas have a lower OKT4/OKT8 ratio compared with nodules in pre-toxic phase. In conclusion: T lymphocyte subpopulations typing in FNA demonstrate that, even in this type of hyperthyroidism, immune response disorders are present and consist of relative increase of suppressor/cytotoxic T cells, compared to T helper cells.     

An epidemiological analysis of monitoring of the immune status of chernobyl nuclear accident liquidators for early detection of risk groups and diagnosis of cancer diseases. Communication 2. Dependence of the rate and changes in the immune status on radiation accident risk factors

It was demonstrated that malignant neoplasms (MNs) most frequently develop in the liquidators involved in the cleanup activities after the Chernobyl nuclear accident (ChNA) in 1986 (43.75%) and the liquidators involved in long-term activities, one-quarter of whom also participated in the cleanup in 1986. The specific features of the immune status (IS) depending on the period and year of the cleanup activities in the ChNA area were determined. In the cohort with a confirmed MN diagnosis, the observed deviations in the people who participated in these activities only in 1986 and on a permanent basis, including 1986, coincided in their pattern but differed in the magnitude of changes in immunological characteristics. Long-term participation during the extreme period and, correspondingly, higher exposure caused an increase in the contents of CD8⁺ T cells, CD16⁺ lymphocytes, and activated T lymphocytes, as well as decreases in the total immunoregulatory index, percentage of CD3-16/56⁺ (NK), and total IgE and a more pronounced deficiency in B lymphocytes. Differences between the groups of liquidators who participated in the cleanup activities in 1986 and 1987 were detected. The liquidators-1987 display the most pronounced deviations in the IS characteristics. Permanent presence in the ChNA area in 1986 and 1987 caused a similar effect but the magnitude of changes in the content of CD4⁺ T lymphocytes (↓), content of CD8⁺ T lymphocytes (↑), and immunoregulatory index (↓) was considerably higher in liquidators-1987. The permanent cleanup activities in Chernobyl in 1987 caused a deficiency in CD4⁺ T lymphocytes; more pronounced increase in the content of CD8⁺ T lymphocytes; decrease in CD4⁺/CD8⁺ index; change in the ratio of NK-T to NK cells; elevation in the counts of CD95⁺, HLA-DR⁺, and activated T lymphocytes; and decrease in the level of total IgE. Any increase in the expression of cell activation markers was undetectable in the liquidators-1986 involved in the cleanup activities on a long-term basis. The specific features of the IS changes depending on the dose of external γ-irradiation were determined. It was shown that the MN rate in liquidators increases with age relative to the number of examined people in each age cohort. The differences in the IS age-related dynamics in liquidators with and without MNs were detectable in stable contents of CD3⁺, CD4⁺, and CD8⁺ T lymphocytes; immunoregulatory index; and contents of CD95⁺ cells and serum IgA at ages of 40 to 70 years old with a subsequent decrease in these parameters and elevation in the content of CD8⁺ T lymphocytes with age in the absence of MNs; continuous increase in CD3-16/56⁺ (NK) cells in the presence of MNs; and decreases in these values after 70 years in the absence of MNs. In both groups of liquidators older than 70 years (with and without MNs), a deficiency in the T-cell component (CD3⁺ and CD4⁺ T lymphocytes and CD4⁺/CD8⁺ ratio) and an increase in the counts of CD8⁺ T lymphocytes are characteristic of the IS. A growing deficiency in CD4⁺ T lymphocytes during the monitoring on the background of increasing content of CD8⁺ T lymphocytes, which additionally contributes to the weakening of immune regulation due to progressing abnormalities in the ratio of regulatory T-lymphocyte subpopulations, can be regarded as one of the characteristics that suggest an adverse life expectancy prognosis for people with MNs.     

Lymphocyte subpopulation state of the human lung during prenatal development]

Distribution of surface lymphocyte markers was assessed in foetus lung by cytofluorimetry using monoclonal antibodies produced by "Ortho Diagnostic systems" and "Becton Dickinson". Seven lymphocyte subpopulations were detected in developing lungs even at the pseudoglandular stage (8-15 weeks). There was a significant increase in these lymphocyte subpopulations in canalicular stage. Quantity of T3+ cells increased 8-fold and B-cells 10-fold. There was a tendency to increased number of cortical thymocytes (T6+), and to decreased ratio T6+/T3+ lymphocytes. The ratio of immunoregulatory lymphocyte subpopulations (T4+/T8+) resembled that of these subpopulations in adult human lung rather than in cord blood. In canalicular stage the share of lung lymphoid cells in S-phase was decreased.     

Interrelation between the activity of hepatitis, liver fibrosis and immune status in children with chronic hepatitis B and C

The lesion of the liver in viral hepatitis was found to depend on the state of the immune system. Relationship between the content of lymphocyte subpopulations (CD3+, CD4+, CD8+, CD20+) in the blood and immunoglobulins (IgG, IgM, IgA) with parameters of semi-quantitative evaluation of the activity of hepatitis and the stage of liver fibrosis in children with chronic virus hepatitis B, C, B + C was studied. The characteristic feature of all hepatitis was a decrease in the number of T lymphocytes CD4+ below the normal level and an increase in the content of B lymphocytes. The correlation between the morphological activity of hepatitis and the amount of T lymphocytes CD8+ was established only in chronic hepatitis B. In chronic hepatitis B and B + C the absolute amount of blood lymphocytes decreased with the increase of the age of the patients, but in chronic hepatitis B this was accompanied by the decrease of the morphological activity of hepatitis and in hepatitis B + C by its increase. The amount of lymphocytes CD4+ rose with the increase of liver fibrosis in chronic hepatitis B. In children with chronic hepatitis C and B + C the amount of blood lymphocytes was found to be unrelated to the morphological activity of hepatitis.     

Immune status of Greek patients with beta-thalassemia major negative for anti-HIV

Patients with thalassemia who receive multiple blood transfusions are at risk for the acquired immunodeficiency syndrome. Peripheral blood lymphocyte subpopulations were studied in 22 multitransfused thalassemic patients; 10 patients were without splenectomy and 12 were studied after splenectomy. Both groups were negative for anti-HIV. Four additional patients who were found positive for anti-HIV and ten healthy controls were also included in this study. Patients without splenectomy compared to controls and to patients after splenectomy showed a significant decrease of both percentage (p less than 0.001) and absolute numbers (p less than 0.001) of Leu-7+ cells without significant abnormalities of T4/T8 ratio (1.56 +/- 0.4). Patients after splenectomy compared to controls and to patients without splenectomy showed a significant increase of the absolute numbers of lymphocytes and lymphocytes subsets T11+, T3+, T4+, T8+ and SmIg+ cells. In the seropositive patients for HIV only a significant increase of the absolute number of T8+ cells was observed while the T4/T8 ratio was 1.24 +/- 0.73. The decrease in the percentage of Leu-7+ cells in patients without splenectomy correlated inversely to the total amount of blood transfused. In conclusion patients with thalassemia had normal T4/T8 ratio and did not show the abnormal immunologic profile that has been reported in haemophiliacs.     

Adoptive immunotherapy of murine cytomegalovirus adrenalitis in the immunocompromised host: CD4-helper-independent antiviral function of CD8-positive memory T lymphocytes derived from latently infected donors.

The ability of memory T lymphocytes derived from latently infected mice to control murine cytomegalovirus disease in the immunocompromised host was studied by adoptive transfer experiments. At a stage of pathogenesis when virus had already colonized target tissues, a therapeutic antiviral function could be ascribed to the CD8+ subset. This in vivo function was not restricted to sites in which intravenously infused lymphocytes usually are trapped or home in, such as the lungs or the spleen, respectively, but was also evident in the adrenal glands, a site to which antiviral effector cells have to specifically migrate. Specific infiltration of adrenal gland cortical tissue by donor-derived CD8+ memory T lymphocytes was demonstrated. CD4+ memory T lymphocytes had no antiviral effect by themselves and also were not required for the function of the CD8+ effector cells in this short-term immunotherapy model. These findings should help settle the debate about which subset of T lymphocytes comprises the effector cells that can directly control cytomegalovirus infection in the murine model system.     

Flow cytometric evaluation of lymphocyte subpopulations in BALF of healthy smokers and nonsmokers

The purpose of this work was to evaluate the normal lymphocyte phenotype in the bronchoalveolar lavage fluid (BALF). BAL was carried out in 12 untreated healthy nonsmoking volunteers and in 9 cigarette smokers. For the analysis of lymphocyte subsets by two-color flow cytometry, the monoclonal antibodies used were directed anti: CD3, CD4, CD8, CD16, CD19, D25, CD45, CD56 and anti HLA-DR. An increase in the total number of cells in BALF of smoking persons and increased proportion of macrophages was observed. The percentage of CD8+ lymphocytes was 1.7 times higher, whereas the proportions of CD4+ cells, and a CD4+/CD8+ ratio were lower 1.5 and 2.6 times, respectively, in the BALF of cigarette smoking persons when compared with nonsmoking volunteers. The changes did not depend on the age of the person. In conclusion, we suggest that the decreased CD4/CD8 ratio and the elevated CD8 T cell subset may be regarded as a potential risk factor associated with clinically asymptomatic lung cancer. Moreover, in the interpretation of BALF from patients with pulmonary diseases cell proportions of nonsmoking and of smoking persons should be compared with the respective controls.     

Multiple myeloma: an immunologic profile. I. Peripheral blood studies.

Seventy-four patients with multiple myeloma, 17 untreated and 57 treated, were studied to characterize their peripheral blood lymphocytes. PBL were studied for E, EAC, and EA rosette-forming cells, SIg, and Fc receptor-bearing cells. The responses to HA, Con A, and PWM were assessed as well as their ability to stimulate or to respond in a MLC. Finally, the capacity of mitogen-stimulated lymphocytes to lyse Chang cells, CRBC, and PHA-stimulated lymphoblasts was examined. These results were compared with a group of normals and patients with benign monoclonal gammopathy. In untreated myeloma patients there was a normal percentage of T cells, but an abnormal distribution of B cells as judged by a decrease in SIg-bearing cells, as well as an increase in EAC rosette-forming cells. Subpopulation analysis showed a marked increase in EAC rosette-forming cells without SIg. PHA, Con A, and PWM, and response in MLC were all normal. However, the ability to stimulate in MLC was significantly depressed. Treated myeloma patients had similar findings, except that the response to PWM was significantly depressed. The capacity of PWM-stimulated cells to lyse target cells was depressed in both groups. The results indicate that, in the peripheral blood of myeloma patients, there are populations of lymphocytes characterized by the presence of the EAC receptor without SIg, which are deficient in the capacity to stimulate an MLC response and the ability to be cytotoxic when stimulated by PWM. The results form a baseline for the study of abnormal lymphoid function in human myeloma.     

Effector activity of OKT4+ and OKT8+ T-cell subsets in lectin-dependent cell-mediated cytotoxicity against adherent HEp-2 cells

The role of OKT4+ and OKT8+ T-cell subsets was studied in lectin-dependent cell-mediated cytotoxicity (LDCC) against adherent HEp-2 human epipharynx carcinoma target cells. LDCC was evaluated by detachment from the monolayer of [3H]thymidine prelabeled HEp-2 cells in a 24-hr assay with a concanavalin A (Con A) dose of 25 microgram/ml at effector:target cell ratios of 5:1, 25:1, and 50:1. Under these conditions but without Con A considerable natural cell-mediated cytotoxicity (NCMC) was not elicited; however, the cytotoxicity was significantly augmented in the presence of Con A (=LDCC) by sheep erythrocyte rosette-forming T lymphocytes and by both OKT4+ and OKT8+ T-cell fractions. LDCC activity by isolated OKT8+ T cells was superior to that by OKT4+ T cells and unfractionated T lymphocytes. By contrast, addition of either OKT4+ or OKT8+ T cells together with unfractionated T lymphocytes, or OKT4+ and OKT8+ T cells mixed at ratios of 1:1, 1:2, and 2:1, to target cells did not result in major differences in comparison of LDCC activities by these mixed effector cell populations with each other or with that by unfractionated T lymphocytes. Parallel studies were carried out to determine the effect of OKT4+ and OKT8+ T-cell subsets on the Con A-induced proliferation of peripheral blood mononuclear cells (PBMC). While OKT8+ T cells inhibited the mitogenic response to Con A, OKT4+ T lymphocytes had no major effect. A higher responsiveness of the OKT8+ to OKT4+ T-cell subset in LDCC to HEp-2 targets and in Con A-induced lymphocyte proliferation is suggested.     

Low affinity E-rosette formation by the human K cell.

Human T lymphocytes were separated into two subsets on the basis of relative affinity for sheep red blood cells (E), and these T cell fractions were examined for cytotoxic reactivity against antibody-sensitized Change liver cells (ADCC). High affinity E-rosette-forming cells (E-RFC) (55 +/- 6% of peripheral blood mononuclear cells), capable of rosette formation despite elevated temperatures of incubation (29 degrees C) and a limited concentration of E, contained few antibody-dependent cytotoxic cells (K cells). In contrast, low affinity E-RFC (23+/-7% of mononuclear cell suspensions) requiring cool temperatures of incubation (4 degrees C) and an excess of E to form rosettes, were highly enriched for ADCC activity. The majority of K cells exhibited low affinity interactions with E. T cells in thymus, tonsil, and lymph node formed high affinity E-rosettes and exhibited little reactivity in ADCC. Only peripheral blood and spleen contained easily identified low affinity E-RFC and anti-body-dependent cytotoxic cells. The proportion of low affinity E-RFC in the peripheral blood of normal subjects correlated closely with reactivity in ADCC, making it possible to predict cytotoxic potential for the E-rosette pattern. These data indicate that the human K cell may belong to a previously unappreciated but functionally important subset of thymic dependent mononuclear cells.