Regulation of human gut B lymphocytes by T lymphocytes

The aims of this study were first, to assess whether or not immunoglobulin secretion from human gut mucosal B lymphocytes can be modified by T lymphocytes, and second whether human gut mucosal T lymphocytes are capable of regulating mucosal B lymphocyte function. T and B lymphocyte enriched cell populations were isolated from gut mucosa and co-cultured in varying proportions. Addition of T lymphocytes to B enriched mucosal cell populations (ratio B:T = 2:1) showed that mucosal B lymphocytes were responsive to T cell 'help'. Addition of more T cells (ratio B:T = 2:10) suppressed immunoglobulin synthesis.     

T and B lymphocytes in Hodgkin's disease

Summary Phytohemagglutinin stimulation of peripheral lymphocytes from 54 patients with Hodgkin's disease and 40 normal blood donors was studied as a parameter for T-cell function. Lymphocytes carrying immunoglobulin determinants (B-cells) in peripheral blood from 67 patients with H. D. and normal blood donors were quantitated by membrane immunofluorescence. PHA stimulation was severely depressed in 48 patients with H. D. as compared to the controls. There was no correlation to clinical stage or histological type. Distribution of B-cells in the normal blood donors was 33% (S. D. ± 4) the mean value in all patients with H. D. was 27% (S. D. ± 12). No specific pattern of distribution of B-cells could be found when the data were analysed for clinical stage (stage I 36.50%±14.47, stage II 26.21%±9.63, stage III 25.77%±12.18, stage IV 25.59%±13.18), histological subtype (lymphocytic predominance 25.84%±13.88, nodular sclerosis 25.58%±11.53, mixed cellularity 29.80%±11.35, lymphocytic depletion 28.00%±13.06) or age. Since no change in the relative distribution of B- and T-cells could be found it is concluded that the disturbance in cellular immunity in these patients is caused by a functional defect of the T-cells rather than a decrease in their number.

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Zusammenfassung Bei 54 Patienten mit Morbus Hodgkin und 40 normalen Blutspendern wurde die Phytohämagglutininstimulierbarkeit peripherer Lymphocyten als Parameter für die T-Zellfunktion untersucht. Bei 67 Patienten mit Morbus Hodgkin und den normalen Blutspendern wurden die Immunglobulindetenninanten auf der Oberfläche peripherer Lymphocyten mit Hilfe der Membranimmunfluorescenz (B-Zellparameter) bestimmt. Bei 48 Patienten mit Morbus Hodgkin war die PHA-Stimulierbarkeit im Vergleich zu den Kontrollpersonen stark erniedrigt. Eine Korrelation zu klinischem Stadium oder histologischem Untertyp konnte nicht gefunden werden. Der Anteil von B-Zellen bei den normalen Blutspendern betrug 33% (S. D. ± 4), der Mittelwert bei allen Patienten mit Morbus Hodgkin 27% (S. D. ± 12). Es konnte kein spezifisches Verteilungsmuster für B-Zellen gefunden werden, wenn man die Ergebnisse nach klinischem Stadium (Stadium I 36,50%±14,47, Stadium II 26,21%±9,63, Stadium III 25,77%±12,18, Stadium IV 25,59%±13,18), dem histologischen Untertyp (lymphocytäre Prädominanz 25,84%±13,88, Noduläre Sklerose 25,58%±11,53, Mischzellform 29,80%±11,35, lymphocytäre Depletion 28,00%±13,06) oder nach Alter aufgliedert. Signifikante Veränderungen in der relativen Verteilung von B- und T-Zellen im peripheren Blut bei Patienten mit Morbus Hodgkin konnten nicht gefunden werden. Der Schluß liegt nahe, daß die bekannten Störungen der cellulären Immunität bei Patienten mit Morbus Hodgkin durch einen funktioneilen Defekt der T-Zellen und nicht durch eine T-Zellverarmung verursacht werden.

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Mit Unterstützung des Landesamtes für Forschung, NRW und der Deutschen Forschungsgemeinschaft.     

The role of T lymphocytes in autoimmune thyroid disease

Abnormalities in cellular mechanisms of immunoregulation are important factors in the initiation and/or propagation of autoimmune thyroid disease and Graves' ophthalmopathy. The primary immunologic lesion, however, remains elusive despite increasingly sophisticated investigations in the areas of antigen presentation and subsequent T-cell responses. Emphasis on functional evaluations of relevant cell populations and advances in lymphokine analysis, interpretation of DR antigen expression, and T-cell cloning offer promise for our future understanding of this complex process.     

Self-nonself concept for cancer and diseases previously known as “autoimmune” diseases

The illegitimate glycosphingolipid antigens of the P blood group system and of the Forssman (Fs) tissue antigen in adenocarcinoma which are foreign to the host suggest the self-nonself concept which applies also to numerous other diseases such as rheumatoid arthritis, lupus, gluomerulonephritis, and idiopathic acute hemolytic anemia. In the presence of the glycosphingolipid antigens such as ABO, P, and Fs, the normal serum of the homozygote recessive precursor contains antibodies for the missing antigen(s). The expected antibody to the Fs antigen was present in about 75% of normal men and women. In cancer sera, the incidence of anti-Fs was decreased to about 35-40%. On testing the normal population anti-Fs was present in 90% of the sera in the youngest group, and this value gradually diminished in the older groups; the incidence of the antibody in the 70-year age group was to about 60%. The rate of loss of anti-Fs with increasing years appears to parallel the gradual loss of anti-A and anti-B isoagglutinin titers. This phenomenon may be associated with the gradual diminution of protein synthesis with aging or the continuous accumulation of soluble immune complexes in the serum, or both. It is suggested that the self-nonself concept is also the basis for the pathogenesis of rhematoid arthritis, lupus erythematosus, idiopathic acute hemolytic anemia, and numerous other conditions classified as "autoimmune" diseases. Some of these diseases are induced by viruses or drugs or both. When a virus or drug attaches itself to the membrane of a tissue cell, the self is converted to nonself which, in rheumatoid arthiritis, alters its self Ig to nonself Ig.     

T and B lymphocytes in pleural effusions.

To determine the diagnostic significance of the determination of T and B lymphocytes in pleural fluid, we studied these cells in peripheral blood and in pleural fluid by means of surface markers. Our study comprised 30 patients suffering from pulmonary tuberculosis, pulmonary malignancy, connective tissue disease, nonspecific pleurisy or congestive cardiac failure. In pulmonary tuberculosis, both the percentage and absolute numbers of T lymphocytes in pleural fluid were significantly higher than in peripheral blood. In patients with pulmonary tuberculosis, pulmonary malignancy or nonspecific pleuritis, the percentages and absolute numbers of B lymphocytes were significantly lower in pleural fluid than in peripheral blood. Considered together with other clinical and laboratory indices, these determinations may aid in the differential diagnosis of pleurisy of various etiology.     

T and B lymphocytes in leukemia therapy.

In order to evaluate the effect of radio- and chemotherapy on immunity, T and B lymphocyte surface receptors were studies sequentially in the blood from 28 previously untreated leukemic children. Following the initiation of chemotherapy an increase in the percent T and B cells was noted in the peripheral blood. In association with sanctuary therapy and chemotherapy there was a decrease in the total number of circulating T and B cells and a relative increase in lymphocytes lacking markers. Based on total numbers at remission the reduction in B cells was greater than in T cells, and the most marked changes occurred during sanctuary therapy. A reduction in the mean serum immunoglobulin was associated with decreasing B cell numbers.     

T and B lymphocytes and blastogenesis in leprosy.

T and B cell percentages and their blastogenic response to PPD and lepromin have been studied in 107 patients of various types of leprosy. T cell counts and their blastogenic response were found to be considerably lower in all types of leprosy as compared to the normal. The counts and stimulation were the lowest for lepromatous leprosy. B cell counts were unaltered in all types of leprosy.     

B and T lymphocytes in man. IV. Circulating B, T and "null" lymphocytes in aging population.

T, B and "Null" lymphocytes were determined in the peripheral blood of 98 adults, 26 of them were between 20 to 40 years of age and 72 between 60 to 96 years of age. The total number of lymphocytes were about the same in both groups. The percent and absolute number of T cells decreased significantly in the aged. Although the percent of B lymphocytes increased with age, the absolute number of B lymphocytes remained about the same.     

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.     

Abnormalities in clonable B lymphocytes and myeloid progenitors in autoimmune NZB mice.

Cloning procedures were used to study B lymphocytes and progenitors of granulocytes and macrophages in NZB mice. Numbers of B cells that were detected in sheep erythrocyte-containing semisolid cultures were only slightly elevated in NZB tissues, and these were normally sensitive to inhibition by anti-mu or anti-delta antibodies or prostaglandin E. However, NZB mice rapidly developed large numbers of B cells that could be cloned in the presence of lipopolysaccharide, and these included unusual anti-mu resistant cells. Numbers of myeloid precursors in NZB bone marrow that were responsive to colony-stimulating activity in L-cell conditioned medium or endotoxin serum were at least normal, but at all ages granulocyte-macrophage precursors were poor responders in cultures stimulated by WEHI-3 cell conditioned medium. Almost no colonies were elicited in NZB cultures with a colony-stimulating activity moiety from WEHI-3 cells. Prostaglandin sensitivity of myeloid precursors from NZB and CBA mice was also different. Codominant genetic control of these abnormalities was suggested by their partial expression in F1 hybrid NZB X CBA and NZB X NZW mice. NZB mice expressed an unexpected IgD allotype allele.     

Discrimination of B, T and null lymphocytes by esterase cytochemistry.

A technique has been devised which optimally demonstrates non-specific esterase activity in human blood lymphocytes. The reaction is carried out on smears fixed with formalin vapour, using alpha-naphthol butyrate as substrate at pH 8 and at a low concentration for a short incubation period. The pattern of esterase activity revealed by this method provides a discriminating marker for mature T-lymphocytes, which show dense, localised, dot-like positivity, and a probable marker for 'null' cells in the form of scattered granular positivity. B cells appear to be negative. Monocytes show a clearly different pattern of granular positivity.     

HLA-E-restricted recognition of cytomegalovirus-derived peptides by human CD8+ cytolytic T lymphocytes

HLA-E-restricted T cell receptor alphabeta+ CD8+ cytolytic T lymphocytes (CTLs) exist as monoclonal expansions in the peripheral blood of some individuals. Here, we show that they recognize, with high avidity, peptides derived from the UL40 protein of different human cytomegalovirus (CMV) strains. Recognition results in the induction of cytotoxicity, IFN-gamma production and cell proliferation. Autologous cells pulsed with CMV-derived peptides become susceptible to lysis by HLA-E-restricted CTLs and induce their proliferation. The high avidity for CMV-derived peptides may explain how these cells are generated in vivo and suggest their possible role in the host defenses against CMV, a virus that evolved various mechanisms to down-regulate classical HLA class I molecules, thus escaping detection by conventional CTLs.     

T cell immunity using transgenic B lymphocytes

Adaptive immunity exists in all vertebrates and plays a defense role against microbial pathogens and tumors. T cell responses begin when precursor T cells recognize antigen on specialized antigen-presenting cells and differentiate into effector cells. Currently, dendritic cells are considered the only cells capable of stimulating T lymphocytes. Here, we show that mature naïve B lymphocytes can be genetically programmed by using nonviral DNA and turned into powerful antigen-presenting cells with a dual capacity of synthesis and presentation of antigen to T cells in vivo. A single i.v. injection of transgenic lymphocytes activates T cell responses reproducibly and specifically even at very low cell doses (≈10²). We also demonstrate that T cell priming can occur in the absence of dendritic cells and results in immunological memory with protective effector functions. These findings disclose aspects in the regulation of adaptive immunity and indicate possibilities for vaccination against viruses and cancer in humans.Adaptive immunity is based on the selection and expansion of lymphocytes bearing clonally distributed antigen receptors (1). It is based on genes having common structural features, the Ig and T cell receptor genes, that code for conformationally stable molecules at the surface of B and T lymphocytes where they serve as receptors for antigen. Clonal selection and expansion follow recognition of antigen by T cells, an event that requires the positive effect of coreceptor molecules that stabilize the interaction with the antigen-presenting cell (APC) and costimulatory molecules that provide a jump start to T cell activation (2, 3). The immune system also stores the ability to respond with greater intensity and faster kinetics upon re-encounter of the same antigen, immunological memory (4).T cells recognize antigen in the form of small peptides in the groove of the MHC molecule at the surface of the APC, dendritic cells (DCs), B lymphocytes, and monocytes/macrophages. Peptides that bind MHC class I molecules consist of 8-9 residues, whereas those that bind class II MHC molecules consist of 13-17 residues (5, 6). Antigenic peptides presented in the context of MHC class I molecules induce cytotoxic T lymphocyte (CTL) responses and are generally the product of degradation of cytosolic proteins via an ATP-dependent, ubiquitin-driven proteolytic system (7). Antigenic peptides presented in the context of MHC class II molecules induce CD4 T cell responses and result typically from the degradation of exogenous antigens internalized by receptor-mediated endocytosis (8). Currently, DCs are viewed as possibly the only APCs able to initiate a T cell adaptive response (9). DCs are endowed with an extraordinary capacity to take up antigen from the extracellular compartment, process it, and display its peptides at the cell surface in conjunction with costimulatory molecules. Owing to these properties, many new approaches to vaccination are based on DCs, notwithstanding the fact that while DC cluster with T cells in lymph nodes rapidly (≈24 h) they also leave lymph nodes rapidly (10). Thus, spatio-temporal factors and considerations on supply of peptides to MHC molecules, which depend on the half-life of the peptide/MHC on the APC (≈4 h for class I and ≈8-22 h class II) (11-13), are relevant to optimize T cell activation in vivo. From the foregoing, the ideal approach is that which uses an APC that is able to actively synthesize and process/present antigen at the same time.B lymphocytes possess distinct APC function. On a per-cell basis, activated B cells are as effective as DCs in activating naïve T cells (14, 15). In vivo expansion of T cells can be driven by activated B cells (16-18) and resting B lymphocytes depending on the availability of adequate costimulation via B7 and CD40 (19, 20). Resting B lymphocytes also activate naïve T lymphocytes specific for their endogenous light chain variable region peptide in vivo (21). This finding is consistent with the fact that lymphoma cell transfectants process and present to T cells peptides of endogenously synthesized Ig variable region via both MHC class II or class I pathways (22, 23). When plasmid DNA coding for an Ig heavy chain gene controlled by the Ig promoter is injected into the spleen of adult mice, production of antibodies and activation of T cell responses against heterologous epitopes expressed in the variable region are readily induced (24, 25).The present study sought to determine whether primary B lymphocytes rendered transgenic ex vivo with nonviral DNA can induce antigen-specific responses in vivo. Typically, murine spleen lymphocytes were rendered transgenic for plasmid DNA coding for transgenes under the control of an Ig-specific promoter by incubation in PBS without Ca²⁺/Mg²⁺ for 60 min (26). To induce antigen-specific CD4 and CD8 T cell responses, we used heavy chain genes engineered to express heterologous T cell determinants in the complementarity-determining regions (CDRs) (27). Transgenic cells were injected into adult histocompatible recipients i.v. within 24 h from in vitro transgenesis (Fig. 1A).Open in a separate windowFig. 1.(A) The process of spontaneous transgenesis and immunization with transgenic lymphocytes. Details of the two procedures are described in Materials and Methods. (B) The characteristics of the primary CD4 T cell response: kinetics and specificity. C57BL/6 mice were injected with 5 × 10³ γ1NV²NA³ transgenic cells. Spleen cells were harvested at different time points and tested in a T cell proliferation assay. Tests were run in triplicate. Values represent the mean stimulation index ± SD of four mice per group (24). The cpm at each time point are as follows: day 5, NVDP peptide (11,581)/medium alone (1,988); day 7, NVDP peptide (40,016)/medium alone (4,932); day 14, NVDP peptide (29,856)/medium alone (1,165); day 21, NVDP peptide (15,788)/medium alone (2,322); and day 28, NVDP peptide (1,014)/medium alone (732). (C) T cell proliferation in lymph nodes (LNs). Fourteen days after injection LNs were collected from the upper, middle, and lower body anatomical regions. The cpm per each group are as follows: upper LN, NVDP peptide (10,576)/NANP peptide (1,201); middle LN, NVDP peptide (10,236)/NANP peptide (1,130); and lower LN, NVDP peptide (9,786)/NANP peptide (887). Spleen cells in the same experiment (data not shown) were NVDP peptide (14,505)/NANP peptide (1,022). (D) Dose-response of CD4 T cell induction. Mice (two per group) were injected with lymphocytes transgenic for γ1NV²NA³ (▪) or γ1NANP (□) as a control, with varying numbers (20,000-70) of transgenic cells. Mice were killed on day 14, and spleen CD4 T cells were restimulated in vitro with the -NVDP- peptide. Proliferation is expressed as cpm. Two independent experiments yielded a similar result. (E) Cytokine production vs. immunizing dose. Culture supernatants from the experiment depicted in D were analyzed for cytokine content (pg/ml). (F) The CD8 T cell response: phase of induction and kinetics after injection of 5 × 10³ γ1NV²NP³ transgenic cells. Spleen cells were harvested at the time points indicated, restimulated in vitro with peptide NP_366-374 (5 μg/ml), and tested in a standard ⁵¹Cr-release assay using EL-4 cells pulsed with NP_366-374 as target at various effector-to-target cell ratios. Results are expressed as the percentage of specific lysis. (G) Dose-response of CTL induction. Groups of three mice each received a single injection of varying numbers (20,000-70) of transgenic cells. Mice were killed on day 14, and the CTL response was measured by using EL-4 cells pulsed with NP_366-374 (filled symbols) or EL-4 alone (open symbols) as targets.     

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.     

Engagement of B7 on effector T cells by regulatory T cells prevents autoimmune disease

Although there is considerable evidence that a subpopulation of regulatory CD4(+)CD25+ T cells can suppress the response of autoreactive T cells, the underlying molecular mechanism is not understood. We find that transmission of a suppressive signal by CD4CD25+ regulatory cells requires engagement of the B7 molecule expressed on target T cells. The response of T cells from B7-deficient mice is resistant to suppression in vitro, and these cells provoke a lethal wasting disease in lymphopenic mice despite the presence of regulatory T cells. Susceptibility of B7-deficient cells to suppression is restored by lentiviral-based expression of full-length, but not truncated, B7 lacking a transmembrane/cytoplasmic domain. Because expression of these B7 truncation mutants restores CD28-dependent costimulatory activity, these findings that indicate B7-based transmission of suppressive activity suggest new approaches to modifying autoimmune responses.