Immunological function of tonsils in atopic and nonatopic children, with special reference to E- and EAC-binding lymphocytes.

This investigation was carried out to determine the immunological function of the human tonsils, especially the role of lymphocytes of the atopic and nonatopic children. Authors introduced E- and EAC-rosettes forming technique with semi-quantitative evaluation of T and B lymphocytes in peripheral blood of 56 normal subjects, 28 atopic and 5 nonaptoic children of both sexes and different age. The percentage of T and B lymphocytes in removed palatine tonsils in 28 atopic and 5 nonatopic children was calculated. The percentage of rosettes forming cells in E and EAC suspensions appeared to be independent of age and sex. The percentage of T lymphocytes in peripheral blood is significantly higher than in the palatine tonsils whereas the percentage of B lymphocytes is significantly higher in the tonsils of atopic and nonatopic children than in the peripheral blood. The mean percentage of B lymphocytes in the tonsils of atopic children is significantly higher than in nonatopic children. The percentage of B lymphocytes was found to be significantly higher in hypertrophic than in small tonsils. Also, in the tonsils with adhesions there was a statistically significant higher percentage of EAC rosettes than in the tonsils free of adhesions. The percentage of T lymphocytes is statistically significant higher in the small tonsils.     

The effect of sera from chronic lymphocytic leukemia patients on normal mitogen-stimulated lymphocytes. A possible correlation with the patients' T and B lymphocytes.

Serum from chronic lymphocytic leukemia (CLL) patients was examined for its effect on normal lymphocytes stimulated by phytohemagglutinin and pokeweed. Responses of the patients' lymphocytes were similarly examined. E and EAC rosettes were also tested. A significantly increased transformation response was found when normal lymphocytes were exposed to sera from those patients who had a reduced number of E-rosette-forming lymphocytes (mean 8%) as compared to the response to sera from patients with a higher number of E-rosette-forming cells. The results suggest that a factor is present in sera from some CLL patients which stimulates the transformation of mitogen-stimulated lymphocytes and which seems to be related to the number of the patient's T and B lymphocytes and perhaps with a T suppressor effect on B lymphocytes.     

Relationship of lymphocytotoxic antibodies to lymphopenia and parameters of disease activity in systemic lupus erythematosus.

Peripheral blood lymphocyte subpopulations determined in 40 patients with systemic lupus erythematosus, revealed significant lymphopenia in comparison to normal controls. The lymphopenia was caused by decreases in the absolute numbers of both T cells and surface immunoglobulin bearing (SIg) cells. There were significant percentage decreases in T cells and normal percentages of SIg cells, resulting in an increased percentage of "null" cells. The lymphopenia was significantly greater in patients considered to have active disease on the basis of clinical parameters, and was strongly correlated with lymphocytotoxic antibodies, DNA antibodies, and hypocomplementemia. The lymphocytotoxic antibodies were reactive with both T and B lymphocytes from normal and ill donors, human organs, and various lymphoblastoid cell lines. In 30 patients who were followed for up to 22 months, the lymphopenia was clearly seen to develop during, but not before, periods of clinical disease activity, independent of changes in therapy.     

Technical Aspects of the Rosette Technique for Detecting Human Circulating B and T Lymphocytes Normal Values and Some Remarks on Null Lymphocytes

The effect of variations in several single steps in the rosette technique for detecting human circulating B, T and O cells is investigated in order to describe the optimal production of B and T rosettes. Variations in pH, duration of incubation, temperature, EAC cell production, age of EAC cells and definition of a rosette are investigated. Using the optimal rosette technique, normal values for 15 healthy men and 15 healthy women (age range 18–65 years) are found to be: B lymphocytes 27.5 % or 477/μ1, T lymphocytes 60.6 % or 1049/μl, O lymphocytes 11.9 % or 206/μl. It is concluded that variations in the analytical procedure may greatly affect the outcome. The existence of O cells is briefly discussed.     

Synovial fluid lymphocytes in rheumatoid arthritis

There is significant evidence implicating immunologic mechanisms in the pathogenesis of rheumatoid arthritis (RA). To investigate the possibility that a cellular imbalance of T and B lymphocytes plays a role in perpetuating synovitis, an examination of the peripheral blood and synovial fluid lymphocytes of patients with seropositive rheumatoid arthritis was undertaken. A significant lymphopenia was encountered in RA patients, with values approximately midway between those in normal controls and the severe lymphopenia seen in a group of patients with systemic lupus erythematosus. A significantly greater diminution of T than B cells was noted in RA, and normal percentage distributions of lymphocytes in peripheral blood were documented. Normal percentages of T and B cells, similar to those in peripheral blood, were found in synovial fluids of patients with seropositive RA, seronegative RA, osteoarthritis, and gout. Two seropositive patients exhibited synovial fluid, but not peripheral blood, lymphocytes that stained both for IgG and IgM. These cells regenerated predominantly IgM after trypsinization and short-term culture with significant diminution of cells staining for IgG. It is postulated that double staining was secondary to surface immunoglobulins with rheumatoid factor activity binding intraarticular IgG. Such cells were not detected in peripheral blood in any of the groups of patients studied.     

Diminished expression of complement regulatory proteins (CD55 and CD59) in lymphocytes from systemic lupus erythematosus patients with lymphopenia

CD55 and CD59 are glycophosphatidylinositol-anchored proteins with complement inhibitory properties. Lymphopenia in systemic lupus erythematosus (SLE) has been associated with autoantibodies targeting nuclear antigens. The aim of this study was to evaluate the surface density of CD55 and CD59 in T and B lymphocytes from patients with SLE and lymphopenia and its possible correlation with the presence of common SLE autoantibodies. Flow cytometric analyses were performed on CD55 and CD59 stained CD3+ and CD19+ cells from 40 SLE patients, 30 with lymphopenia and 10 without it, and 25 healthy controls. Autoantibodies were detected in the sera by enzyme linked immunosorbent assay. The mean fluorescence intensity of CD55 and CD59 in T and B cells was significantly diminished in SLE patients with lymphopenia when compared with healthy subjects. Interestingly, the opposite was found in T and B cells from non-lymphopenic SLE patients. Although there was no correlation between CD55 and CD59 surface density and the presence of any specificity of the autoantibodies tested, higher titres of anti-dsDNA, anti-SM and anti-ribosomal p antibodies were significantly associated with lymphopenia. The deficiency of CD55 and CD59 expression may play a role in the pathophysiology of lymphopenia, most likely by increasing the susceptibility of cells to complement mediated cytolysis.     

Lymphocyte phenotype and lymphokines following anti-thymocyte globulin therapy in patients with aplastic anaemia

Twenty-two patients with adult onset aplastic anaemia were analysed before and after therapy with anti-thymocyte globulin (ATG). Lymphocyte phenotype, lymphokine levels or production, and haematopoietic progenitor cell number were measured 3 months after therapy; clinical response was determined 1 year post-therapy. By flow cytometry there was a significant reduction in both the proportion and absolute number of peripheral blood lymphocytes expressing activation antigen Tac (IL-2 receptor) and in the proportion of HLA-DR+ lymphocytes. For T cells bearing HLA-DR, there were proportional decreases in both activated helper and suppressor cells. There was no statistically significant difference pre-ATG to post-ATG in the absolute numbers of total, helper and suppressor lymphocytes. In all 10 haematologic responders the number of Tac bearing lymphocytes after ATG therapy was in the normal range, but half of 12 non-responding patients continued to have abnormally elevated numbers of Tac+ T cells. The proportion of Tac+ cells were not related to transfusion history. Gamma-interferon levels in serum by radioimmunoassay were elevated in almost half the aplastic patients; post-ATG, gamma-interferon was detectable in only three patients. Haematologic response to ATG therapy was associated with increased numbers of haematopoietic progenitors post-treatment, but pre-treatment values were not predictive of a response. These results are consistent with a pathogenic role for activated T-cells and their lymphokine products and suggest that the target of ATG therapy may be a Tac+ lymphocyte.     

Conjugation of toll-like receptor-7 agonist to gastric cancer antigen MG7-Ag exerts antitumor effects

AIM: To investigate the effects of our tumor vaccines on reversing immune tolerance and generating therapeutic response.METHODS: Vaccines were synthesized by solid phase using an Fmoc strategy,where a small molecule toll-like receptor-7 agonist(T7) was conjugated to a monoclonal gastric cancer 7 antigen mono-epitope(T7-MG1) or tri-epitope(T7-MG3).Cytokines were measured in both mouse bone marrow dendritic cells and mouse spleen lymphocytes after exposed to the vaccines.BALB/c mice were intraperitoneally immunized with the vaccines every 2 wk for a total of three times,andthen subcutaneously challenged with Ehrlich ascites carcinoma(EAC) cells.Three weeks later,the mice were killed,and the tumors were surgically removed and weighed.Serum samples were collected from the mice,and antibody titers were determined by ELISA using an alkaline phosphate-conjugated detection antibody for total Ig G.Antibody-dependent cell-mediated cytotoxicity was detected by the lactate dehydrogenase method using natural killer cells as effectors and antibody-labeled EAC cells as targets.Cytotoxic T lymphocyte activities were also detected by the lactate dehydrogenase method using lymphocytes as effectors and EAC cells as targets.RESULTS: Vaccines were successfully synthesized and validated by analytical high performance liquid chromatography and electrospray mass spectrometry,including T7,T7-MG1,and T7-MG3.Rapid inductions of tumor necrosis factor-α and interleukin-12 in bone marrow dendritic cells and interferon γ and interleukin-12 in lymphocytes occurred in vitro after T7,T7-MG1,and T7-MG3 treatment.Immunization with T7-MG3 reduced the EAC tumor burden in BALB/c mice to 62.64% ± 5.55% compared with PBS control(P 0.01).Six or nine weeks after the first immunization,the monoclonal gastric cancer 7 antigen antibody increased significantly in the T7-MG3 group compared with the PBS control(P 0.01).As for antibody-dependent cell-mediated cytotoxicity,antisera obtained by immunization with T7-MG3 were able to markedly enhance cell lysis compared to PBS control(31.58% ± 2.94% vs 18.02% ± 2.26%; P 0.01).As for cytotoxic T lymphocytes,T7-MG3 exhibited obviously greater cytotoxicity compared with PBS control(40.92% ± 4.38% vs 16.29% ± 1.90%; P 0.01).CONCLUSION: A successful method is confirmed for the design of gastric cancer vaccines by chemical conjugation of T7 and multi-repeat-epitope of monoclonal gastric cancer 7 antigen.     

T and B lymphocytes in malignant melanoma patients.

The method of formation of spontaneous (E) and immune (EAC) rosettes and the test of lymphocyte survival in short-term cultures with phytohemagglutinin (PHA) have been used as parameters of cellular immunocompetence in 64 patients with malignant melanoma and 33 control donors. Significant changes have been found in the number of T lymphocytes whose values corresponded to the clinical stage as well as to the clinical postoperational course in the patients. A decreased number of T lymphocytes was a sign of progression of the disease, while a normal value indicated remission. Lowered ratios of T lymphocytes were followed by an increased proportion of both the B lymphocytes and the null, nonrosetting cell elements. The results in patients with malignant melanoma permit the relevant parameters to be considered as an effective in vitro correlate of cellular immunity of these patients.     

Spontaneous and induced immunoglobulin secretion by synovial fluid B lymphocytes in rheumatoid arthritis.

The functional properties of B lymphocytes in synovial fluid (SF) from patients with rheumatoid arthritis (RA) were analysed by means of a reverse haemolytic plaque forming cell (PFC) assay. SF mononuclear cells spontaneously secreted IgG, but little IgM or IgA. The SF cells failed to respond to the polyclonal B cell activators pokeweed mitogen (PWM) and Epstein-Barr virus. However, SF B cells cocultured with autologous T lymphocytes from the blood and stimulated with PWM secreted IgG but little IgM or IgA. The PFC responses of blood B cells cocultured with autologous SF T cells in the presence of PWM were low; irradiation of the T cells increased the blood B lymphocyte responses, but the differences were not statistically significant. It is concluded that suppressor SF T cells may be partly responsible for the poor response of SF B cells to PWM.     

Overexpression of Fas/CD95 and Fas-induced apoptosis in a patient with idiopathic CD4+ T lymphocytopenia.

The mechanisms of apoptosis have become better understood, in part with the discovery of Fas/CD95. We report the case of a patient characterized by a decreased CD4+ T cell count and an overexpression of Fas/CD95 resulting in apoptosis. A 54-year-old man presented with disseminated Mycobacterium xenopi infection. Analysis showed CD4+ T lymphopenia. Tests for human immunodeficiency virus (HIV) types 1 and 2 were negative. We compared the patient with eight healthy controls and five HIV-infected patients in terms of the expression of Fas/CD95 and Fas-mediated apoptosis of peripheral T lymphocytes. The percent of CD95+ cells in lymphocytes was 98% for the patient, and the mean percent of CD95+ cells in lymphocytes +/- SD for HIV-infected patients and healthy controls was 75% +/- 16% and 36% +/- 26%, respectively. The patient had a high level of spontaneous apoptosis, and apoptotic cells were all identified as being CD4+ T cells. Monoclonal antibodies to CD95 dramatically increased apoptosis of CD4+ T cells exclusively. CD4+ T lymphopenia observed in our patient correlated with an overexpression of Fas together with spontaneous and Fas-induced apoptosis.     

Lymphocyte Membrane Markers in Acute Lymphoblastic Leukaemia

S ummary . T and B lymphocytes can be detected by specific membrane markers. Spontaneous rosette formation with sheep RBC (E rosette) is a T cell marker, whereas receptors for activated complement component (EAC), for the Fc regions of certain immunoglobulin classes (FcR), and for membrane immunoglobulins (mIg) are specific for B cells. Using these lymphocyte membrane markers we evaluated peripheral blood lymphocytes of 18 patients in different stages of acute lymphoblastic leukacmia (ALL).
Our results indicate that in acute untreated ALL there is no set immunologic pattern; instead there is a variety of selective deficiencies. Patients in remission have normal populations of T and B cells, whereas those in relapse present a rather obscure picture.
The percentage of lymphocytes with membrane markers was compared to the percentage of blast cells and an inverse relation is observed. This indicates that these are abnormal blast cells.
The changes from the untreated acute stage through to remission was followed in one subject, and in two others additional study was performed on bone marrow and spinal fluid lymphocytes.     

Isolation and characterization of infiltrates in the nerves of patients with neuritic leprosy

A study was done on the characteristics of infiltrating cells in the nerves of 9 patients with pure neuritic leprosy, by preparing a single cell suspension. The patients had no skin lesion. Histopathological examination revealed that 2 of the 9 nerves showed granulomas characteristics of tuberculoid leprosy, while the remaining 7 had features of lepromatous granulomas. In the nerves showing tuberculoid granulomas, a high proportion of lymphocytes were T cells as they formed rosettes with sheep erythrocytes and only a few percent were EAC rosette forming cells. On the other hand, the nerves showing lepromatous granulomas contained only occasional lymphocytes which formed E and EAC rosettes. Macrophages from the granulomas of all the nerves were esterase positive, peroxidase negative, contained M. leprae and did not exhibit C3 surface receptors.     

CELLULAR AND HUMORAL IMMUNITY IN PATIENTS WITH HYPERTHYROID GRAVES'' DISEASE BEFORE, DURING AND AFTER ANTITHYROID DRUG TREATMENT

Many reports of thyroid stimulating immunoglobulins (TSI) in relation to treatment of Graves' disease have been published and with variable results concerning prediction of permanent remission or relapse after therapy. A range of methods has been used and little has been published measuring TSI by using their ability to stimulate cyclic AMP production in human thyroid cells in monolayer culture. We therefore conducted a prospective study of the predictive value of such an assay in patients with hyperthyroid Graves' disease before, during and after treatment of one year with methimazole and thyroid hormone substitution. Furthermore, the possible relationship between activated suppressor T lymphocytes and TSI in patients followed before, during and after medical therapy has been studied. Patients were divided into two groups; group I, 15 patients, who stayed in remission and group II, 14, who relapsed during the first year after discontinuation of therapy. Mean TSI activity did not differ between the two groups before and during the first half year of medication. In the second half year of treatment, however, mean TSI activity was significantly lower in group I. TSI activity at the end of treatment appeared to have no value in predicting final outcome. Increased TSI activity in group II during treatment was reflected in an increased pertechnetate thyroidal uptake as compared to that in group I. There was no relationship between changes in TSI activity and T cell subsets (Leu 1, 2a, 3a). We found no difference in T lymphocytes between the two groups at any time during observation. Subsets of T lymphocytes in both patient groups did not differ from normal.(ABSTRACT TRUNCATED AT 250 WORDS)     

SUBACUTE THYROIDITIS: ACTIVATED HLA-DR AND INTERFERON-γ EXPRESSING T CYTOTOXIC/SUPPRESSOR CELLS IN THYROID TISSUE AND PERIPHERAL BLOOD

Using a two-colour direct immunofluorescence staining technique, we investigated activated HLA-DR-expressing T helper and T cytotoxic/suppressor cells in peripheral blood of six patients with subacute thyroiditis at referral and at follow-up and in blood from 20 controls. In three of the patients, thyroid fine-needle aspirates were examined as well. At referral, all patients had elevated blood levels of activated T helper and T cytotoxic/suppressor cells 2 (2-4)%, median and range, vs 0 (0-2)%, P less than 0.001 and 12.5 (2-24)%, vs 0 (0-1)% P less than 0.001). At follow-up, the activated proportion of T helper cells had become normal whereas some activated T cytotoxic/suppressor cells remained, 7 (0-8)%. No significant changes in total T cell number were detected when data at referral and at follow-up were compared. In thyroid aspirates, HLA-DR expressing thyrocytes were observed; the total proportion of T cytotoxic/suppressor cells was elevated (70% compared with 35% in blood) and 70% of the T cytotoxic/suppressor cells were HLA-DR+. Furthermore, 55% of the thyroid-infiltrating lymphoid cells were positive for interferon (IFN-gamma+). The finding of activated T cytotoxic/suppressor cells in the blood and thyroid tissue in subacute thyroiditis is consistent with a viral aetiology. Furthermore, intrathyroidal IFN-gamma+ lymphocytes are likely to contribute to expression of major histocompatibility complex (MHC) class II antigens on thyrocytes. No autoantibodies, however, were detected, which suggests that aberrant expression of MHC class II molecules alone is not sufficient to provoke an autoimmune response.     

Autoantibodies to T and B cell lines detected in serum samples from patients with systemic lupus erythematosus with lymphopenia and hypocomplementaemia.

Antibodies to lymphocytes in serum samples from 88 patients with systemic lupus erythematosus (SLE) and 15 normal control subjects were examined by a cell enzyme linked immunosorbent assay (ELISA) with four human T and B cell lines as antigens. The antibodies reacted with the Wa B cell line and the T cell lines P12 (CD4-, CD8+), Jurkat (CD4-, CD8-), and Hut78 (CD4+, CD8-). Antibody titres in serum samples from patients with SLE were higher than in those from normal control subjects. Titres of antibodies to P12 were correlated with titres of antibodies to Wa, Jurkat, and Hut78 in serum samples from patients with SLE. IgG antibodies to P12 were associated with lymphopenia and reduced haemolytic complement. By thin layer chromatography immunostaining, the antibodies in serum samples from two of 10 patients with SLE with high titres of IgG antibodies to P12 and lymphopenia were shown to react with three monosialoglycosphingolipids and two neutral glycosphingolipids from P12 cells. Antibodies to lymphocytes in serum samples from patients with SLE react with T and B cell lines, recognise a series of cell membrane glycosphingolipids and are associated with the lymphopenia and hypocomplementaemia typical of active disease.     

T细胞在炭疽保护性抗原免疫中的作用——免疫T细胞继承转移试验

取炭疽保护性抗原(PA)免疫的小鼠脾脏,用尼龙棉柱过滤和EAC花环相结合的方法分离出免疫T细胞,继承转移给同系小鼠,可使受体鼠获得抗炭疽菌感染的保护力。     

The differences in purine metabolism between T and B lymphocytes

In this study enzyme activities involved in purine metabolism were measured in T and B lymphocytes separated by E and EAC rosetting methods. Adenosine deaminase, purine nucleoside phosphorylase and HGPRTase activities were significantly elevated in T cells, compared to the activities in B cells. There were no significant differences in adenosine kinase and APRTase activities between T and B lymphocytes. In contrast, PRPPsynthetase activities were higher in B cells than in T cells. The uptake of various radiolabeled precursors by mitogen stimulated lymphocytes was studied. The uptake of 14C-formate by the mitogen stimulated lymphocytes was markedly lower, compared to that of 14C-adenosine and or 14C-purine bases. The uptake of 14C-adenosine by PHA stimulated lymphocytes was considerably higher than that of Con A or PWM stimulated lymphocytes. However, the uptake of 14C-hypoxanthine into lymphocytes stimulated with PWM was increased by comparison with unstimulated lymphocytes. From these results it seems that adenosine plays a central role in the metabolism of T cells, and that purine bases are preferentially utilized in B cells.     

Thymosin-inducible lymphocytes in the peripheral blood of patients with malignant melanoma.

E rosette-forming (T) lymphocytes and surface immunoglobulin-bearing lymphocytes were estimated in 85 patients with malignant melanoma. The melanoma patient group had lower mean levels of T lymphocytes and higher mean levels of immunoglobulin-bearing (? B) lymphocytes than did normal subjects. The absolute and percentage depressions of T-cell levels in the melanoma patients were stage-related, as was the depression of total lymphocyte and B-lymphocyte levels. The T lymphopenia in the melanoma patients could, in vitro, be partially abolished by fetal calf serum (as used in many E rosetting methods), and could be totally abolished by thymosin fraction 5 (Hoffmann-La Roche) at optimum concentration. In view of the ability of thymosin to restore T cells to normal levels in all of the T-lymphopenic patients, a clinical trial of this hormone in selected melanoma patients of all stages appears to be warranted.     

Perioperative cimetidine administration promotes peripheral blood lymphocytes and tumor infiltrating lymphocytes in patients with gastrointestinal cancer： Results of a randomized controlled clinical trial

AIM: To study the effects of perioperative administration of cimetidine (CIM) on peripheral blood lymphocytes, natural killer (NK) cells and tumor infiltrating lymphocytes (TIL) in patients with gastrointestinal (GI) cancer. METHODS: Forty-nine GI cancer patients were randomized into treatment group, who took CIM in perioperative period, and control group, who did not take the drug. The treatment was initiated 7 days before operation and continued for 10 days after surgery. At baseline examination before operation, on the 2nd and 10th postoperative days, total T lymphocytes, T helper cells, T suppressor cells, and NK cells in peripheral blood were measured respectively by immunocytochemical method using mouse-anti human CD(3), CD(4), CD(8) and CD(57) monoclonal antibodies. Blood samples from 20 healthy volunteers were treated in the same way as normal controls. Surgical specimens were examined during routine histopathological evaluation for the presence of TIL in tumor margin. Immunohistochemical study was performed to measure the proportion of T and B lymphocytes in TIL population. T and B lymphocytes were detected respectively using mouse-anti-human CD(3) and CD(20) monoclonal antibodies. RESULTS: In comparison with normal controls, both the treatment and control groups had decreased T cells, T helper cells and NK cells at baseline. In control group, total T cells, T helper cells and NK cells declined continuously with the disease progression and the decrease became more obvious after operation. From baseline to the 2nd postoperative day, the proportion of total T cells, T helper cells, and NK cells went down from 60.5+/-4.6% to 56.2+/-3.8%, 33.4+/-3.7% to 28.1+/-3.4%, and 15.0+/-2.8% to 14.2+/-2.2%, respectively. On the other hand, there were significant improvements in these parameters after CIM treatment. On the 10th postoperative day, the treatment group had significantly higher percentages of total T cells, T helper cells and NK cells than control group. Moreover, CIM treatment also boosted TIL response, as was reflected by findings that 68% (17/25) of the patients in treatment group had significant TIL responses and only 25% (6/24) of the cases had discernible TIL responses (P<0.01). CONCLUSION: Perioperative application of CIM to GI cancer patients could help restore the diminished cellular immunity induced by tumor burden and surgical maneuver. The drug could also boost TIL responses to tumor. These effects suggest that the drug be used as an immunomodulator for GI cancer patients.