T and B lymphocytes in South American pemphigus foliaceus.

T and B lymphocytes were quantified in peripheral blood of thirty patients with South American pemphigus foliaceus according to their ability to form rosettes with sheep erythrocytes (E) or sheep erythrocytes sensitized with antibody an complement (EAC). When compared with the counts obtained from thirty normal subjects, a decrease was found in the total T-lymphocyte count (32-33+/-7-7 versus 46-7+/-8-7) and in the T functional lymphocyte count as detected by the active rosette test. The mean percentage of B lymphocytes within the total number of lymphocytes was not significantly different from that of normal subjects (24-0 +/- 8-2 versus 25-3+/-8-0). Lymph node sections from three pemphigus patients examined for E or EAC adherence showed depletion of T cells in the paracortical areas. The low percentage of E cells in the peripheral blood and the depletion of E cells in paracortical areas of lymph nodes from patients with South American pemphigus foliaceus may reflect an impaired cellular immunity.     

T and CR+ lymphocyte profile in leprosy and the effect of treatment.

Thymus-derived lymphocytes (T lymphocytes) and complement receptor-bearing lymphocytes (CR+ lymphocytes) were estimated by using erythrocyte rosettes and erythrocyte-antibody-complement rosettes as markers in untreated lepromatous and untreated tuberculoid patients and in healthy controls. Treated lepromatous cases were also investigated. Ten cases of untreated lepromatous patients were reassessed 6 months or more after therapy commenced. A significant decrease in both percentages and absolute numbers of CR+ cells in the untreated lepromatous leprosy subjects was observed. This decrease showed a return to normal levels after treatment. The percentage of T cells in the untreated lepromatous cases was normal; however, the absolute numbers of T cells and the total lymphocyte count showed a significant decrease. After therapy, the T cell population was unchanged but the total number of lymphocytes increased significantly with treatment. The absolute number of T and CR+ cells was significantly less in the untreated than in the treated lepromatous patients.     

T- and B-lymphocyte subpopulations following radiotherapy for invasive squamous cell carcinoma of the uterine cervix.

Lymphocyte subpopulations in patients with cervical carcinoma were studied before and after radiotherapy. T lymphocytes were recognized by their ability to form spontaneous rosettes with sheep erythrocytes (E rosettes). Two surface marker characteristics were used to detect B lymphocytes: receptors for activated complement responsible for erythrocyte--antibody--complement (EAC) rosette formation, and surface membrane immunoglobulin (SmIg), readily stainable by immunofluorescence. We have demonstrated a significant depression of total lymphocytes after radiotherapy which persists for more than 5 years. This reduction in lymphocytes is due to a loss of E rosette-forming T cells, SmIg-bearing B cells and null cells. Absolute numbers of EAC rosette-forming B cells are not altered by treatment, and there is a rise in this cell type when the results are expressed as percentages of the total lymphocyte count. The possible functional immunological significance of these changes is discussed.     

Increase of peripheral B lymphocytes in Graves' disease.

Peripheral T and B lymphocytes were examined in autoimmune thyroid diseases. The percentages of T and B lymphocytes were calculated from the proportions of E and EAC rosette-forming cells and peroxidase-positive cells determined by micromethods. In thyrotoxic Graves' disease, the percentage of T cells was significantly lower, and the percentage of B cells was higher than in normal controls. The absolute count of B lymphocytes was also markedly increased. The serum levels of thyroid hormones showed a significant correlation with the percentage of B cells and an inverse correlation with that of T cells in untreated cases of Graves' disease. Similar abnormalities of lymphocyte subpopulations were observed in patients with thyrotoxic Graves' disease under drug therapy, but the proportions and absolute counts of T and B lymphocytes were normal in euthyroid patients with Graves' disease, either under drug therapy or in remission. No abnormalities in T and B cells were found in Hashimoto's disease. The data indicate that the main feature of the abnormality of the lymphocyte subpopulations in thyrotoxic Graves' disease is an increase of B lymphocytes. The reasons for the discrepancy between our results and those of earlier reports and for the B cell abnormality in Graves' disease are discussed.     

The Effect of Levamisole on Peripheral T-Lymphocytes of Patients with Malignant Lymphomas

The effect of various concentrations (0.015-10 μg/ml) of Levamisole (LMS) on the peripheral lymphocytes of patients with malignant lymphomas (ML) and normal donors was investigated in vitro. The parameters studied include : E rosettes forming cells (total T lymphocytes), active E rosettes (early T lymphocytes) and DNA synthesis induced by pitogens PHA and Con A.

LMS improved significantly lymphocyte response both in patients with ML and normal donors when the cells were stimulated by Con A. In both groups no significant effect was observed on the response to PHA nor on the percentage of E-rosettes, whereas the mean number of active E rosettes was significantly increased on all concentrations of the drug.

While in the normal subjects a positive statistical correlation between active E-rosettes and Con A response was observed, in patients with ML an inverse correlation was found. This latter correlation was partially reversed by LMS.     

T and B lymphocytes in myasthenia gravis.

Peripheral blood lymphocytes from seventeen non-thymectomized and nine thymectomized patients with myasthenia gravis (MG) and thirteen healthy controls were examined for the presence of surface markers characteristic of T and B lymphocytes by rosette formation with sheep red blood cells (SRBC). T cells were identified by their capacity to spontaneously form rosettes with SRBCs. The percentage of B lymphocytes was determined by the erythrocyte antibody complement (EAC) rosette-forming test. The EAC complex was prepared with either whole rabbit anti-SRBC serum or with the IgM fraction of rabbit anti-SRBC serum. The two kind of erythrocyte complement rosette-forming cells (EAC-RFC) are designated erythrocyte-haemolysin-complement RFC (EA(H)C-RFC), and erythrocyte-IgM-complement RFC (EA(M)C-RFC). The percentage of total lymphocytes and T cells was not altered in MG patients. The percentage of 'active' T cells, which have been considered to be more actively involved in cellular immunity, was also similar in MG patients and controls. A significant increase in EA(H)C-RFC occurred in both thymectomized and non-thymectomized MG patients, while in B cells detected by EA(M)C-RFC no alterations were found. The increase in EA(H)C-RFC in lymphocytes from MG patients may be due to an increase in the 19S antibody-forming B lymphocytes or to an increase in T cells which have Fc receptors on their surface.     

The effect of immunosuppressive and anti-inflammatory drugs on lymphocyte population and MIF in glomerulonephritis.

The effect of immunosuppressive and anti-inflammatory drugs on the T and B lymphocyte populations, as well as cellular hypersensitivity in relation to GBM antigens (measured by MIF activity) in 60 patients with different types of glomerulopathy has been tested. The results were compared with a control group of 47 untreated patients and 32 healthy subjects. The treatment was carried out using the following drugs: azathioprine, cyclophosphamide, prednisone, ibuprofen and indomethacin. In a sample of 14 patients receiving ibuprofen and in 7 receiving arathioprine, indomethacin and prednisone, the T and B lymphocyte kinetics as well as MIF activity were repeatedly tested before and during the treatment of 150 days. In the patients under this treatment, a decreased percentage of lymphocytes with receptors for a complement as well as, in some cases. MIF suppression was observed. A comparison of healthy subjects and untreated patients with a group of 39 subjects tested for T and B lymphocytes as well as MIF activity after several months of the above treatment was made. It was found that supportive therapy with azathioprine and prednisone led to a normalization of B lymphocytes (EA and EAC rosettes) and to a small increase of T lymphocytes number. On the other hand, treatment with indomethacin and ibuprofen led to an increased B lymphocyte count (EA and EAC rosettes). Cyclophosphamide given with azathioprine caused a decrease in the proportion of T lymphocytes and an increase in the number of null cells. In addition, it was found that immunosuppressive drugs as well as anti-inflammatory drugs, even when administered in supportive doses caused in some cases, the suppression of cellular immunity as measured by MIF activity.     

The Effect of Levamisole on Peripheral T-Lymphocytes of Patients with Malignant Lymphomas

Abstract

The effect of various concentrations (0.015–10 μg/ml) of Levamisole (LMS) on the peripheral lymphocytes of patients with malignant lymphomas (ML) and normal donors was investigated in vitro. The parameters studied include : E rosettes forming cells (total T lymphocytes), active E rosettes (early T lymphocytes) and DNA synthesis induced by pitogens PHA and Con A.

LMS improved significantly lymphocyte response both in patients with ML and normal donors when the cells were stimulated by Con A. In both groups no significant effect was observed on the response to PHA nor on the percentage of E-rosettes, whereas the mean number of active E rosettes was significantly increased on all concentrations of the drug.

While in the normal subjects a positive statistical correlation between active E-rosettes and Con A response was observed, in patients with ML an inverse correlation was found. This latter correlation was partially reversed by LMS.     

Alterations in the Distribution and Proliferative Responses of Rhesus Monkey Peripheral Blood and Spleen Cells During Malaria (Plasmodium knowlesi) Infection

Rhesus monkeys are used frequently as animal models in malaria research, but few studies have evaluated lymphocyte functions in these animals after experimental infections with the primate malarial parasite Plasmodium knowlesi. In this study, the distribution and mitogen responses of mononuclear cells in the peripheral blood and spleens of 16 P. knowlesi-infected rhesus monkeys were followed. All animals included in the study developed acute infections and were bled out with parasitemias of more than 50%. With progression of the infection, alterations in the peripheral blood mononuclear cells were observed, including decreases in the percentage of T cells (measured by E rosette formation) and the total numbers of E and EAC rosette-forming cells per cubic millimeter. In addition, peripheral blood mononuclear cells displayed reduced responses to mitogen stimulation with phytohemagglutinin, concanavalin A, and pokeweed mitogen. Peripheral blood mononuclear cells from infected animals showed similar reductions in mitogen responses when cultured in media containing 15% autologous pre- or postinfection plasma. The mitogen responses of spleen cells did not appear to be affected, but a significant reduction in the proportion of splenic T cells was observed. These lymphocyte changes in P. knowlesi-infected rhesus monkeys are similar to those reported for mice with acute rodent malaria and for humans with chronic Plasmodium falciparum infections.     

Effect of radiotherapy on lymphocyte cytotoxicity in vitro.

The cytotoxic functions of highly purified blood lymphocytes from patients with breast cancer were studied before and after radiotherapy. Addition of PHA or of rabbit antibodies to target cells (chicken erythrocytes) were chosen as two means of inducing lymphocyte cytotoxicity in vitro. The proportion of T and non-T-lymphocytes was determined by means of E and EAC rosette tests. The antibody-induced cytotoxocity of lymphocytes decreased following radiotherapy while that mediated by PHA remained unchanged. There was some reduction in the percentage of EAC rosette-forming cells. These results, as well as our earlier observations, suggest that the decrease in the peripheral blood of the proportion of lymphocytes with receptors for activated complement is responsible for changes in the antibody-mediated lymphocyte cytotoxicity.     

Natural cytotoxic reactivity of human lymphocyte subpopulations.

The spontaneous cytotoxicity of human peripheral blood lymphocyte preparations from normal donors for K562 target cells was examined. Effector cells were separated into SRBC rosette forming cell (RFC) and non-rosette forming cell (non-RFC) fractions using optimal and suboptimal rosetting procedures. RFC and non-RFC fractions both had high cytotoxic activity irrespective of the rosetting procedure. Owing to the larger size of the RFC fraction, it contained a higher proportion of the total activity in the preparation. Nylon fibre column adherent and non-adherent fractions also both produced cytotoxicity. Nylon fibre non-adherent cells separated by SRBC separation gave a RFC fraction with low activity and a non-RFC fraction with high activity. Separation of nylon fibre adherent cells gave RFC and non-RFC fractions with high cytotoxic activity. Therefore cytotoxic cells did not form a discrete subpopulation and either occur in several lymphocyte subsets or show a variable capacity to form SRBC rosettes and adhere to nylon fibre.     

Demonstration of T Lymphocytes in Cerebrospinal Fluid

Cerebrospinal fluid (CSF) was allowed to drop straight into Hanks's balanced salt solution. After centrifugation the pellet was resuspended and mixed with sheep erythrocytes. The mixture was further handled as in the E-rosette test with peripheral blood lymphocytes. CSF from 20 individuals were investigated, and rosette-forming cells (RFC) were found in all. Six patients with normal fluid had between 46% and 83% RFC. Four patients with multiple sclerosis had increased numbers of RFC (94%-96%). Low numbers of RFC were found in one patient with cerebellar ataxia and in one of two patients with acute viral meningitis. With this technique RFC can be counted even in normal CSF with a 3-ml sample.     

Age-related changes in T- and B-lymphocyte subpopulations in the peripheral blood

Quantitation of T and B lymphocytes in infants and children is an important test in the diagnosis of a suspected immuno-deficiency. Previous studies indicated that the absolute and relative numbers of lymphocyte subpopulations vary with age, but these data in the pediatric age group are incomplete and often contradictory. We reviewed the literature and investigated the relationship between age and lymphocyte subpopulations in healthy infants and children using common methods and recent methodologic improvements. We found that absolute numbers of T and B cells followed the same trend as the total lymphocyte count, which was elevated at birth, increased in the first six months, and then gradually decreased to adult levels at approximately 13 years of age. Compared with adult values, the percentage of B cells also was higher at birth and continued to increase for six months, followed by a gradual decrease to adult levels by late childhood or early adolescence. The percentage of T cells gradually increased to adult levels by the same age range.     

T and B lymphocytes in the liver and peripheral blood of subjects with acute and chronic hepatitis.

E and EAC rosette-forming cells in the peripheral blood and in the liver of subjects with acute and chronic hepatitis were studied. We found a highly significant reduction (P less than 0.01) of E rosette percentage in the lymphocytes isolated from the liver patients with chronic persistent, and chronic active, hepatitis. EAC rosette-forming cells were significantly increased in the liver of patients with chronic active hepatis (P less than 0.01). In this condition lymphocytes with Fc receptor were also found.     

A sex difference in the leucocyte count

In a study of a group of 325 blood donors between the ages of 18 and 65 years, the total leucocyte and polymorph counts of the females aged 50 to 65 years were found to be significantly lower than those of the corresponding group of male donors. In the whole series of female donors, there were significantly lower total leucocyte and lymphocyte counts with increasing age and a less significant (P [unk] 0.1) fall in the polymorph count. No age variation in the male counts was found.Analysis of large numbers of leucocyte counts performed on hospital patients has confirmed the sex difference in the total leucocyte count found in the blood donors.The factors responsible for the differences are discussed.     

In vivo and in vitro action of chloroquine on surface markers of human peripheral lymphocytes.

Chloroquine was administered orally to twenty normal individuals and the effect of the drug on surface markers of peripheral bloof lymphocytes was studied. The total number of circulating lymphocytes and leucocytes in the blood did not change significantly after chloroquine administration. However, there was a significant fall in the percentage and number of lymphocytes with erythrocyte (E) and C'3 markers and an increase in cells lacking both these markers. In vitro experiments were carried out to study the mechanism of action of the drug on the expression of the lymphocyte receptors. Lymphocytes treated with chloroquine in vitro failed to show any change in their capacity to bind erythrocytes or erythrocytes coated with Ab and complement. The sera from chloroquine-treated individuals failed to show any factor inhibiting E and EAC rosette formation. The studies indicate that chloroquine may not act directly on the lymphocyte surface markers and cause inhibition of their expression but that it may act in some indirect way affecting one or more of the many factors involved in the normal expression of the markers.     

Effect of Ku80 depletion on the preintegrative steps of HIV-1 replication in human cells

To study the decreasing responsiveness of the immune system during aging, influenza virus specific cellular immunity was investigated in a cohort of healthy blood donors between 18 and 70 years of age. The percentage of influenza A virus specific T cells was determined by flow cytometry and found not to change during aging. After stimulation with phorbol 12-myristate 13-acetate and ionomycin, an increase in the percentage of IFN-gamma and IL-4 producing CD8(+) T cells was observed during aging. In addition, the cytotoxic T lymphocyte (CTL) activity was investigated in two additional groups of five donors, 18-20 and 68-70 years of age. The lytic capacity of purified CD8(+) T cells, after in vitro stimulation of peripheral blood mononuclear cells with influenza A virus, seemed lower in 68- to 70-year-old donors than in 18- to 20-year-old donors. Therefore we conclude that the reduced CTL activity in the elderly is not the result of a lower frequency of virus-specific T cells, but more likely the result of impaired antigen-specific proliferation or lower lytic capacity of these cells.     

Serial measurement of nonspecific immune parameters in chronically hemodialyzed renal failure patients

Nonspecific immune monitoring of the percentage active T (A-T) rosette-forming cells (RFC) and spontaneous blastogenesis (SB) appear to be useful indexes of host reaction toward allografts. In order to assess the significance of the observed changes, renal failure patients were serially evaluated before and after hemodialysis for the percentage of total T (T-T) and A-T RFC and for peripheral blood leukocyte metabolic activity measured by a whole blood spontaneous blastogenesis (SB) assay. Renal failure patients had a significantly lower (P<0.05) percentage of T-T RFC than normal persons, while the percentage A-T RFC and SB remained stable. A hemodialysis treatment did not change the mean values of any of the three parameters. On the other hand, serial patient evaluation over 3 months revealed significant fluctuations in the percentage T-T RFC, but not the percentage A-T RFC and SB. Since the percentage A-T RFC and SB are relatively constant measures of immune status in ungrafted patients, significant changes in these nonspecific immune probes may reflect allograft rejection.     

RRBC and EAC rosette formation after DNCB contact sensitization and cyclophosphamide treatment of guinea-pigs.

Rabbit erythrocyte (RRBC) and EAC rosette formation, for detection of T and B lymphocytes respectively, was studied in thymus, lymph nodes and peripheral blood of guinea-pigs. Using two different methods for preparation of RRBC rosette-forming cells (RFC) it was found that, without an incubation period at 37 degrees, a smaller part of the pool of T cells is detected which is particularly sensitive to cyclophosphamide (cy). RRBC rosette formation was studied for 2 weeks after treatment with one high dose of cy and appeared to be minimal in the thymus after 1 week and maximal in the lymph nodes at about day 3. EAC rosette formation was completely blocked by cy treatment. Four days after contact sensitization with 2,4-dinitrochlorobenzene (DNCB) no percentual increase was found in RRBC-RFC, suggesting a parallel increase of B and T cells in stimulated lymph nodes. Cytological screening of the rosettes at day 4 after DNCB showed that of the RRBC-RFC approximately 17 per cent were blast cells, while no difference was detected in the number of blast cells participating in EAC rosette formation as compared to the unstimulated lymph nodes. In contrast to RRBC-RFC the percentage of EAC-RFC in draining lymph nodes is increased after contact sensitization. The mechanism of this increase is discussed.     

Receptors for Helix pomatia A haemagglutinin on leukaemic lymphocytes from patients with chronic lymphocytic leukaemia (CLL).

Blood lymphocytes from thirteen patients with CLL were studied for surface-bound Ig (SIg), Fc receptors (EA rosettes), receptors for sheep erythrocytes (E rosettes) and receptors for Helix pomatia A haemagglutinin (HP), a carbohydrate-binding protein with specificity for N-acetyl-D-galactosamine and related sugars. Fluorescein-labelled HP binds to subpopulations of human peripheral blood lymphocytes (PBL) treated with neuraminidase. In normal peripheral blood, HP binds to the T-lymphocytes while the majority of the B cells bearing surface-bound immunoglobulin do not have receptors for HP. In untreated CLL, HP binds to 90-100 percent of the neuraminidase-treated PBL. Almost all of the SI-G-POSITIVE CELLS IN CLL patients also have receptors for HP. Two groups of patients were found: in one the total fraction of SIg+ cells was less than or equal to 50 percent and about 30 percent of these lost their Ig during incubation at 37 degrees C. No such loss of SIg was revealed in the remaining patients where total SIg+ fraction was approximately 70 percent. These patients usually had higher blood lymphocyte counts, probably reflecting a more advanced disease. CLL patients in remission with low numbers of leukaemic cells also had low numbers of blood lymphocytes carrying both SIg and HP-receptors. It is concluded that leukaemic cells carry both HP receptors and SIg. Testing of this combination therefore provides a valuable new tool for monitoring patients with CLL.