Experimental inflammatory bowel disease--role of T cells.

BACKGROUND: Our experiments were aimed to test: 1. which lymphocyte subpopulations participate in mouse colitis, produced by intrarectal (i.r.) deposition of trinitrobenzene sulphonic acid (TNBSA, TNP hapten); 2. the expression of cell adhesion molecules on lymphocytes draining the site of reaction; 3. the influence of mouse haplotype on the development of colitis. METHODS: CBA/J, BALB/c and C57BI/6 inbred and outbred Swiss Webster strains were used. Mesentheric lymph node (MLN) cells of immunized animals, unseparated or separated into CD4+, CD8+ or gammadelta+ and alphabeta+ T cell subpopulations or depleted of B lymphocytes, were transferred into recipients which were challenged i.r. with TNBSA. Inflammatory reaction in the colon was confirmed macro- and microscopically and by myeloperoxidase (MPO) level. MLN lymphocyte surface markers were tested cytofluorimetrically using appropriate antibodies. RESULTS: Sensitization with TNP results in chronic colitis (hapten dose-dependent colon weight gain and cellular infiltrate, significant increase of MPO level) only in CBA/J and BALB/c strains and can be adoptively transferred in a cell-dose dependent manner into syngeneic recipients by T alphabeta+ cells of both CD4+ and CD8+ subpopulations. T gammadelta+ cells were ineffective and B lymphocytes do not participate in the passive transfer reaction. In MLN the number of T lymphocytes positive for cell adhesion molecules particularly LPAM-1 (V-CAM1) and LPAM-2 increases significantly. CONCLUSIONS: Both CD4+ and CD8+ lymphocytes participate in the development of TNP-induced colitis. High MPO level may suggests that both Th1 and Th2 cells are involved. Colitis is accompanied by a significant accumulation in MLN of T lymphocytes positive for several cell surface adhesion molecules characteristic for memory T cells. Significant differences in susceptibility to develop colitis were found between different strains of mice.     

Early stages of human marrow lymphocyte differentiation: induction in vitro by thymopoietin and ubiquitin.

To study early stages of human lymphocyte differentiation, bone marrow cells were physically separated according to their density and size by gradient centrifugation and then velocity sedimentation. The isolated cell fractions were incubated with putative inducing agents and then assayed for their expression of an array of surface differentiation markers. The inducing agents used were two polypeptides, thymopoietin (Tp) and ubiquitin (Ub), and the cyclic nucleotide, dibutyril cyclic 3'5' adenosine monophosphate (cAMP). Tp, Ub, and cAMP each induced the ability to form sheep erythrocyte rosettes by small lymphocytes, which may thus represent T cell precursors. Ub and Tp induced rosette formation with mouse erythrocytes on lymphocytes of more heterogenous size, which may be "early" B cell precursors. Ub alone could induce surface IgM expression on small lymphocytes, which might be "late" B cell precursors. Both Tp and Ub induced Fc receptors on small lymphocytes. Complement receptors could not be induced on marrow lymphocytes by Tp, Ub, or cAMP. A number of lymphocyte precursors can thus be identified by their physical characteristics and their ability to respond to particular soluble factors with the expression of new differentiation markers.     

Human epidermal cells from ultraviolet light-exposed skin preferentially activate autoreactive CD4+2H4+ suppressor-inducer lymphocytes and CD8+ suppressor/cytotoxic lymphocytes

In vivo exposure of human epidermis to UV abrogates the function of T6+DR+ Langerhans cells and induces the appearance of Ag-presenting T6-DR+ OKM5+ cells in the epidermis. Since UV exposure of murine skin results in Ts lymphocyte activation, we investigated the capacity of human epidermal cells (EC) harvested 3 days after in vivo UV exposure to activate regulatory and effector autologous T lymphocyte subsets. T lymphocytes were separated into CD8+ suppressor/cytotoxic lymphocytes and CD4+ helper/inducer lymphocytes by C lysis and panning. The CD4+ subset was further divided by using the 2H4 mAB to obtain CD4+2H4+ lymphocytes (inducers of TS lymphocytes) and CD4+2H4- lymphocytes (inducers of B cell Ig production and inducers of cytotoxic T cells). Unirradiated suction blister-derived EC from control skin (C-EC) and from skin exposed in vivo to UV (UV-EC) were cultured with purified autologous T lymphocyte subsets in the absence of added Ag. The resultant T lymphocyte proliferation was detected by [3H]thymidine uptake. UV-EC were highly effective in the stimulation of CD4+ lymphocytes, whereas C-EC were poor stimulators. The stimulator effect of UV-EC was abrogated after depletion of DR+ UV-EC. When CD4+ lymphocytes were fractionated, UV-EC consistently demonstrated enhanced ability to stimulate suppressor-inducer CD4+2H4+ lymphocytes relative to C-EC. Although less responsive than CD4+2H4+ lymphocytes, CD4+2H4- lymphocytes also demonstrated greater proliferation to UV-EC than to C-EC. Neither UV-EC nor C-EC were able to activate CD8+ lymphocytes devoid of CD4+ lymphocytes. However, after addition of rIL-2 at concentrations that allow binding only to the high affinity IL-2R on T lymphocytes, UV-EC induced vigorous proliferation of CD8+ lymphocytes, whereas C-EC induced only background levels of proliferation. C lysis of leukocytes resident within UV-EC resulted in 66 to 70% reduction of CD8+ lymphocyte proliferation. In conclusion, UV-EC may activate CD8+ lymphocytes by at least two pathways: (1) UV-EC activation of CD4+2H4+ lymphocytes may induce differentiation/proliferation of CD8+ suppressor cells and (2) UV-EC activation of CD4+ cells may induce IL-2 production, that, in combination with UV-induced epidermal leukocytes, stimulates CD8+ cells.     

Development and regulation of chlamydia-responsive murine B lymphocytes

We have examined characteristics of chlamydia-stimulated mouse B cells as well as cells that regulate polyclonal responses in vitro. B lymphocyte proliferation stimulated by chlamydia arises at a similar time as Escherichia coli lipopolysaccharide (LPS)-induced proliferative responses during ontogeny. In contrast, development of immunoglobulin (Ig)-secreting cells after chlamydia stimulation is delayed by several weeks relative to ontogeny of LPS-inducible plaque-forming cells (PFC). The lack of Ig secretion by immature B cells is not due to a deficiency of Lyb5+ B lymphocytes, since X-linked immunodeficient (xid) NBF1 mice that lack this B lymphocyte population respond well to chlamydia stimulation. Adherent cells are important for chlamydia-stimulated B lymphocyte differentiation, but are not as necessary for their proliferation. Neither adult adherent cells nor T cells can correct the inability of immature spleen cells to develop into Ig-secreting cells; spleen cells from 2-wk-old mice (i.e., immature B cells) will not suppress adult B lymphocyte responses to chlamydia. When B lymphocytes are separated according to their buoyant densities, chlamydia stimulates low density (activated) B cells to proliferate and differentiate better than high density (resting) cells. Proliferative responses to chlamydia arise earlier during ontogeny, do not require adherent cells, and can proceed to a relatively greater extent in resting B cell population (compared with activated B cells) than induction of Ig-secreting cells.     

Malignant pleural effusions due to small cell carcinoma of the lung. An immunocytochemical cell-surface analysis of lymphocytes and tumor cells

Thirteen malignant pleural effusions due to small cell carcinoma (SCC) of the lung were immunocytochemically studied using the peroxidase-antiperoxidase adhesive slide assay for the determination of cell surface antigens. A panel of monoclonal antibodies (MAbs) was used to determine the lymphocyte subpopulations and the reactivity of the tumor cells. Of the lymphocytes, 87 +/- 1% were CD3+ T cells, with 72 +/- 10% CD4+ helper/inducer T cells and 20 +/- 5% CD8+ suppressor/cytotoxic T cells. Only a minority of T lymphocytes were activated in terms of expressing the surface markers CD38 and HLA-DR. The distribution of the lymphocyte subpopulations was not significantly different from the distribution in other malignant and nonmalignant pleural diseases previously studied, indicating that the reaction pattern of the lymphocytes in the pleural cavity is similar in different diseases. The tumor cells from all cases were positive for LeuM1, CD16 and HLA-DR; 10 of 11 cases were positive for HEA-125, Sam 2 and Sam 10. Positivity for epithelial membrane antigen was observed in 11 cases, for OKT9 in 8 cases and for carcinoembryonic antigen in 6 cases. A total or partial loss of the reactivity with HLA-1 was found in nine cases. The reactivity pattern of the tumor cells with the MAbs used in this study is not specific for SCC of the lung because other carcinoma cells also reacted with these markers. Additional morphologic criteria, such as cell size and cell configuration, are needed to recognize the immunocytochemically positive-reacting cells as tumor cells from SCC of the lung. However, the immunostaining allows a better identification of the tumor cells, especially in cases with a small quantity of tumor cells.     

Replication of human cytomegalovirus in human peripheral blood T cells.

The replication of human cytomegalovirus (HCMV) strain AD169 was studied in human peripheral blood granulocytes, monocytes-macrophages, B lymphocytes, and T lymphocytes. Progeny virus was produced in some T-cell cultures stimulated in the allogeneic mixed lymphocyte reaction and was regularly obtained when stimulated T cells were grown in the presence of interleukin 2. Replication of HCMV in these cultures was documented by increases in titer, expression of early and late antigen as assessed by indirect immunofluorescence and Western blot, and viral DNA synthesis as determined by dot-blot assays. Approximately 0.05% of cells in virus-producing cultures formed infectious centers, indicating that only a subset of cells takes part in active virus replication. In double-immunofluorescence experiments this subset was found to consist primarily of the T3+ and T8+ phenotype. By infection of preparatively separated T4+ and T8+ T lymphocytes, however, it could be shown that both T-cell subsets were susceptible to HCMV infection as indicated by increases in titer and by DNA kinetics. We conclude from these data that the T lymphocyte might be a target for HCMV in vitro, which is in accordance with in vivo findings in HCMV-infected patients.     

Separation of human lymphocyte subpopulations by density gradient electrophoresis. I. Different mobilities of T (T mu, T alpha) and B lymphocytes from human tonsils

T and B lymphocytes from human tonsils were separated by density gradient electrophoresis on the basis of their surface charge. The high-mobility cell fractions were found to be highly enriched in T lymphocytes with only very small proportions of B cells. In contrast, the low-mobility fractions were predominantly B lymphocytes, and had only 10 to 30% contamination of T cells. The intermediate-mobility fractions contained both T and B lymphocytes in approximately equal proportions. IgM-bearing lymphocytes, as well as cells with receptors for mouse erythrocytes, the Fc portion of IgG, and complement were found in the intermediate- and low-mobility fractions. T lymphocytes, prepared by E rosetting, were also electrophoresed by this method and found to be of higher mobility as compared with peripheral blood T lymphocytes. T cells with Fc receptors for IgM (Tμ) or IgA (Tα) were found to be considerably heterodisperse with regard to surface charge and were present in all fractions. The separated cell fractions were treated in vitro with various concentrations of concanavalin A and thereafter examined for Tμ, Tγ, and Tα phenotypes. Low concentrations of Con A (2.5 μg/ml) had no effect on cell surface phenotypes. However, higher concentrations of Con A (20μg/ml) significantly reduced the numbers of T cells having IgM receptors (Tμ), but failed to alter the expression of the Tγ phenotype. The latter finding contrasts to that observed with T cells from the peripheral blood where high concentrations of Con A increase the proportions of the Tγ cells. This study demonstrates that density gradient electrophoresis can be used for the separation and study of lymphocyte subpopulations from human tonsils.     

Sepsis-induced apoptosis causes progressive profound depletion of B and CD4+ T lymphocytes in humans

Patients with sepsis have impaired host defenses that contribute to the lethality of the disorder. Recent work implicates lymphocyte apoptosis as a potential factor in the immunosuppression of sepsis. If lymphocyte apoptosis is an important mechanism, specific subsets of lymphocytes may be more vulnerable. A prospective study of lymphocyte cell typing and apoptosis was conducted in spleens from 27 patients with sepsis and 25 patients with trauma. Spleens from 16 critically ill nonseptic (3 prospective and 13 retrospective) patients were also evaluated. Immunohistochemical staining showed a caspase-9-mediated profound progressive loss of B and CD4 T helper cells in sepsis. Interestingly, sepsis did not decrease CD8 T or NK cells. Although there was no overall effect on lymphocytes from critically ill nonseptic patients (considered as a group), certain individual patients did exhibit significant loss of B and CD4 T cells. The loss of B and CD4 T cells in sepsis is especially significant because it occurs during life-threatening infection, a state in which massive lymphocyte clonal expansion should exist. Mitochondria-dependent lymphocyte apoptosis may contribute to the immunosuppression in sepsis by decreasing the number of immune effector cells. Similar loss of lymphocytes may be occurring in critically ill patients with other disorders.     

Separation of a population of human T lymphocytes that bind prostaglandin E2 and exert a suppressor activity

Prostaglandin E2 (PGE2) is a potent inhibitor of immune functions. Two possible mechanisms of PGE2-mediated suppression have been proposed: one is a direct inhibitory effect exerted on interleukin 2-producing T cells; the second is mediated by the activation of nonspecific suppressor T lymphocytes. We previously showed that PGE2 can directly activate human T lymphocytes to suppress lymphocyte proliferation and B lymphocyte maturation. Herein is described the binding of 10 to 30% of human peripheral blood T lymphocytes to insolubilized PGE2 coated to albumin-Sepharose. The T lymphocytes that bound PGE2 (PGE2(+] could be eluted by the addition of serum and gentle shaking of the beads. The following data indicated the specificity of the binding: i) T lymphocytes after an overnight incubation, a condition known to abolish sensitivity to PGE2, lost their affinity for PGE2; ii) preincubation of T lymphocytes with PGE2 blocked the binding; iii) PGE2(+) T cells bound PGE after a 24-hr incubation, whereas PGE2(-) T cells did not. Few T cells bound albumin, and only a small percentage (7 to 9%) bound 6-keto-prostaglandin F1 alpha-coated beads. Among PGE2(+) T cells, there was a slight increase in the percentage of OKT8+ cells. Although T cells that had no affinity for PGE2 (PGE2(-] proliferated as well as unseparated T lymphocytes when stimulated with mitogens or antigens, the proliferative response of the PGE2(+) subset was poor. Moreover, PGE2(+) T lymphocytes did exert a strong suppressor activity on mitogen- or allogeneic cell-induced lymphocyte proliferation as well as on pokeweed mitogen-driven B cell maturation into Ig-containing cells. PGE2(-) T lymphocytes were shown not to exert a significant suppressor activity in these assays. The PGE2(+) subset-mediated suppression was not secondary to a carry-over of PGE2 released from the beads, because its suppressor activity was not altered by the addition of an anti-PGE2 serum. Moreover, PGE2(-) T lymphocytes were not sensitive to the inhibitory activity on cell proliferation of PGE2. These results indicate that a given functional subset of peripheral blood T lymphocytes binds PGE2, and that at least some of them are activated into suppressor T cells. The relationship between the PGE2-activatable T suppressor subset and other functionally defined suppressor T cells remains to be clarified; it is suggested, however, that PGE2 can act as an immunoregulator through the activation of identifiable suppressor T cells.     

The selective activation of T8+ cells by neuraminidase and galactose oxidase is mediated by activated T4+ cells

The response of highly enriched populations of human T8+ lymphocytes to the oxidative mitogenic enzymes neuraminidase (NA) and galactose oxidase (GO) was enhanced by NAGO-primed T4+ lymphocytes. No similar enhancement occurred when the cells were primed with phytohemagglutinin (PHA). In the absence of subclass contamination (1%), the T8+ and T4+ cells responded equally to NAGO by the criterion of DNA replication. The addition of a small number, 2-10%, of NAGO-T4+ cells to the NAGO-T8+ cells enhanced DNA synthesis by as much as 8.5-fold. Augmentation of the cellular response did not occur unless the T4+ cells were activated by NAGO. The converse situation, 2-10% of NAGO-T8+ cells in a primarily NAGO-T4+ cell population, did not increase the DNA synthetic response of the NAGO-T4+ cells. The NAGO-T4+ cells did not augment the early event of increased phosphatidylinositol metabolism or the midcycle event of induction of receptors for interleukin 2 (IL2) and transferrin. The NAGO-T4+ cells therefore increased the probability that fully activated T8+ lymphocytes crossed the G1/S boundary. The basis for this effect was not an enhanced responsiveness of the NAGO-T8+ cells to IL2 or to other soluble growth mediators in medium conditioned by NAGO-activated lymphocytes. The results of this investigation thus implicate a control point in the NAGO-T8+ lymphocyte cell cycle that is positively modulated by the NAGO-T4+ cells themselves or by a product of their activation.     

Characterization of rat bone marrow cells. II. Analysis of surface antigens in small lymphocytes with particular reference to thymus antigen-carrying cells.

Rat bone marrow (BM) small cells were enriched by velocity sedimentation, further separated by means of free-flow electrophoresis, and characterized using T- and B-cell-specific surface markers. More than 80% of these cells were small lymphocytes by morphological criteria and reacted with lymphocyte-specific antisera. A minority of cells had high electrophoretic mobility (EPM), carried surface antigens characteristic of mature T cells, and lacked B-cell markers. These cells may represent recirculating T cells. A small number of cells were found with rat B lymphocyte-specific antigen (RBLA) and surface immunoglobulin (sIg) and had medium EPM. These cell fractions also contained “null” cells which were devoid of T- and B-cell-specific antigens. More than 80% of the BM small cells had low EPM and carried the three subspecificities of the Thy-1 antigen complex and the Thy-A antigens. These antigens were found at several-fold higher concentration on the surface of all thymocytes, but are lacking in most other lymphocytes. The thymus antigen-carrying BM cells of low EPM do not carry other T- and B-cell-specific markers found in thymocytes and peripheral T and B lymphocytes. These markers comprise the T-cell antigens RTLA (rat T-lymphocyte-specific antigen) and RHLA (antigens specific for rat T cells of high EPM) and the B-cell markers RBLA and sIg. Thus the majority of rat BM lymphocytes differ from all other lymphocytes of the T- and B-cell series which makes any classification on this basis purely speculative.     

T lymphocyte subpopulations and lymphocytotoxic antibodies in patients with autoimmune haemolytic anaemia

Evaluation of T lymphocyte subpopulations was performed on peripheral blood of patients affected by idiopathic or associated autoimmune haemolytic anaemia. A marked reduction of absolute number of T gamma and T mu cells was observed in 11 of 16 patients; a decrease of both OKT4+ and OKT8+ cells was found in 8 of 10 patients. Circulating cytotoxic antibodies against autologous and allogenic T lymphocytes and/or thymocytes were found in almost all the cases. T lymphocyte subsets depletion, probably connected to antibodies against T lymphocytes and their thymic precursors, could play a role in autoimmunity because of T3+/T4+ cell depletion.     

Phenotypic and functional analysis of the cellular response in regional lymphoid tissue during an acute virus infection

A phenotypic and functional analysis has been made of the cellular response in regional lymphoid tissue of C57BL/6J mice infected with lymphocytic choriomeningitis virus. Massive recruitment of nondividing cells occurred from 3 days after infection, with total numbers of CD8+ T lymphocytes, B220+ B cells, and Thy-1- B220- null cells being high from day 4 to day 6. In contrast, the peak counts for CD4+ T cells were recorded on day 4 and declined dramatically thereafter. Enhanced expression of IL-2R and Ly-24, both of which can be regarded as T cell activation markers, was found for both the CD4+ and the CD8+ subsets, being most prominent for the CD8+ T cells on day 6. Evidence of T cell proliferation was not recognized until days 5 and 6, coincident with enhanced responsiveness of the lymphocytes to rIL-2 and the development of virus-specific cytotoxic activity. Elimination of the CD4+ T cells by treatment of mice with mAb did not modify either the pathogenesis of lymphocytic choriomeningitis, or the expression of activation markers on the CD8+ T cells which are known to be the key effectors in this disease. Thus, the pattern of responsiveness for the CD8+ population is of recruitment to the lymph node, progressive increase in the expression of activation markers and enhanced sensitivity to rIL-2, with late proliferation and generation of cytotoxic activity. This model provides a system for the rigorous in vivo analysis of parameters influencing lymphocyte differentiation and activation in a virus infection.     

Isolation of two early B lymphocyte progenitors from mouse marrow: a committed pre-pre-B cell and a clonogenic Thy-1-lo hematopoietic stem cell

Two novel early B lymphocyte precursor populations have been identified by their capacity to differentiate in Whitlock-Witte bone marrow cultures. Cells expressing neither the B lineage antigen B220 nor Thy-1 contain committed B cell precursors which differentiate in short-term culture into pre-B and B cells. The other population expresses low levels of Thy-1, and lacks B220 as well as the T cell markers L3T4 and Lyt-2. The Thy-1+ cells which initiate long-term B cell cultures contain clonogenic B cell precursors at a frequency of 1 in 11, a 100-fold enrichment over unseparated bone marrow. Thy-1+ cells are also highly enriched for myeloid-erythroid precursors (CFU-S). Thy-1+ cells allow long-term survival of lethally irradiated mice and fully reconstitute the hematopoietic system, including T and B lymphocyte compartments. These results indicate that this population (approximately 0.1% of bone marrow) may contain the pluripotent hematopoietic stem cell.     

Functional characterization of human T lymphocyte subsets distinguished by monoclonal anti-leu-8

Previous studies have shown that monoclonal anti-Leu-8 antibody identifies functionally distinct subpopulations within both the Leu-2 (T8+) and Leu-3 (T4+) lineages of human T lymphocytes. We now report in detail on the tissue distribution of the Leu-8 antigen and on extensive functional studies of T cells subsets distinguished by their expression or lack of expression of this marker. Leu-8 is present on a wide variety of hematologic cells, including granulocytes, T and B lymphocytes, monocytes, and null or NK cells. Within lymph nodes and tonsils, Leu-8 is absent from both B and T cells within germinal centers but is present on nearly all paracortical lymphocytes. Leu-8 is present on most but not all EBV-transformed B cell lines, reflecting its presence on a subset of normal peripheral blood B cells. None of six malignant T cell lines tested were Leu-8+, whereas most circulating T cells are Leu-8+. Although standard immunoprecipitation techniques failed to demonstrate any specific bands on SDS polyacrylamide gels, the antigenic determinant recognized by anti-Leu-8 is protein or protein-associated, because brief treatment of target cells with pronase abrogated binding of anti-Leu-8. Both Leu-3+8+ and Leu-3+8- cells proliferated in response to several soluble antigens and to autologous and allogeneic non-T cells. Nonetheless, nearly all of the helper T cells for PWM- and AMLR-induced PFC were contained within the Leu3+8- subset. Optimal suppression of the PWM-induced PFC response required both Leu-2+8+ and Leu-2+8- cells, and irradiation of either subset with 3000 R abrogated the capacity of the recombined subsets to effect suppression. In contrast to help for B cell differentiation, both Leu-3+8+ and Leu-3+8- cells were capable of amplifying the development of allospecific T killer cells; precursor and effector T killer cells could be found within both Leu-2+8+ and Leu-2+8- subpopulations. The correlation between Leu-8 phenotype and selected immune functions of T cells (and B cells; see companion paper) indicates that anti-Leu-8 distinguishes important immunoregulatory T and B lymphocyte subsets in man.     

Activated and memory T lymphocytes in human milk.

Since activated macrophages and cytokines are found in human milk (HM), a flow cytometry study was conducted to determine whether T cells in HM display phenotypic markers of recent or previous activation. HM was collected during the first 3 d of lactation. The Paint-a-Gate program was used to optimize gating on the lymphocyte population. A mean +/- 1 SD of 4 +/- 3% of total HM leukocytes were lymphocytes and 96 +/- 3% were macrophages and granulocytes (N = 33 subjects). HM lymphocyte populations were further analyzed in five subjects. T cells (CD3+) represented 83 +/- 11% and B cells (CD19+) were 6 +/- 4% of HM lymphocytes. The mean CD4/CD8 ratio of T cells in HM was 0.88 (range 0.40-1.25). This ratio was significantly decreased compared to the peripheral blood (PB) of control adults (P less than 0.02) and postpartum women (P less than 0.02), due mostly to a significant increase in CD8+ CD3+ cells in HM compared to the PB of control adults (P less than 0.002) and postpartum women (P less than 0.05). T cells bearing markers of recent activation were significantly increased in HM compared to the PB of control adults: 85 +/- 7% of CD3+ cells in HM were HLA-DR+ (controls, 10 +/- 4%; P less than 0.001), and 15 +/- 6% of CD3+ cells in HM were IL-2R+ (controls, 6 +/- 2%; P less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)     

Changes in blood leucocyte populations induced by acute maximal and chronic submaximal exercise

Absolute (x 10(3).mm-3) or relative (%) numbers of blood leucocyte types (monocytes, lymphocytes, neutrophils) and lymphocyte subsets (T11+, T4+, T8+, B1+, and NKH1+) reacting with specific monoclonal antibodies were determined at rest, immediately after maximal exercise on a treadmill, in six controls (C), and in six young cyclists before training (BT) and after 5 months of training (AT). Maximal exercise significantly increased the absolute number (mobilization) of virtually all the types of leucocytes and subsets of lymphocytes in C, BT and AT subjects. In these subjects mobilization of natural killer cells (NKH1+) and cytotoxic/suppressor T lymphocytes (T8+) was greater than mobilization of the other leucocyte types and lymphocyte subsets; however, maximal exercise induced no significant changes in the relative numbers of any leucocyte types and lymphocyte subsets, except in the case of T4+ lymphocytes in At cyclists. Chronic submaximal exercise induced increased mobilization of neutrophils and decreased mobilization of lymphocytes during maximal exercise, except in the case of B lymphocytes (B1+) and NKH1+ cells, and decreases in the absolute and relative number of neutrophils at rest. It remains to be seen how these results can explain the modifications of leucocyte activities noted in vitro after isolated or chronic exercise.     

Suppressor cell function of human granular lymphocytes identified by the HNK-1 (Leu 7) monoclonal antibody

The HNK-1 (Leu 7) differentiation antigen defines a subpopulation of human granular lymphocytes with natural killer (NK) and K cell function. In this study, we investigated whether HNK-1+ cells, identified with the monoclonal antibody and purified with a fluorescence-activated cell sorter (FACS), could function as suppressor cells. The results demonstrated that purified HNK-1+ cells efficiently suppressed both PWM-induced IgG production by B cells and T cell proliferation in mixed lymphocyte reactions (MLR). Manifestation of this suppressor cell activity required immune complex activation and was partially sensitive to 2000 rad irradiation. This suppressor cell activity was predominantly mediated by a subset of HNK-1+ cells that have previously been shown to have maximum NK function and lack expression of the E rosette (ER) receptor and T cell antigens (e.g., T3 and T8). Thus, HNK-1+ER- cells suppressed a MLR by an average 52%; HNK-1+ER+ were one-half as efficient, causing an average 23% suppression. For comparison, we also examined the characteristics of Leu 2a+ suppressor T lymphocytes. In contrast to HNK-1+ cells, unactivated Leu 2a+ cells suppressed both B and T cell responses. This suppressor activity was not augmented by immune complex activation and was absolutely radio-sensitive in PWM assays. HNK-1+ cells, especially the HNK+ER- subset, can therefore mediate suppressor cell function in addition to their spontaneous cytotoxic function. Furthermore, some of their suppressor cell properties are distinct from those attributed to other types of suppressor lymphocytes.     

The majority of lymphocytes in the bone marrow, thymus and extrathymic T cells in the liver are generated in situ from their own preexisting precursors.

Parabiotic pairs of B6.Ly5.1 and B6.Ly5.2 mice were used to investigate how lymphocytes in various organs and various lymphocyte subsets mixed with partner cells. The origin of partner cells was determined by using anti-Ly5.1 mAb in conjunction with immunofluorescence tests. Parabiosis was also produced after the irradiation of B6.Ly5.2 mice at various doses to prepare an immunosuppressive partner. Irrespective of irradiation, lymphocytes and other hematopoietic cells in the bone marrow and lymphocytes in the thymus showed a low mixture of partner cells in comparison with those of all other organs tested. On the other hand, lymphocytes in the blood, spleen, and lymph nodes became a half-and-half mixture of their own cells and partner cells by 14 days after parabiosis. Among lymphocyte subsets, intermediate CD3 cells (i.e., CD3int cells) and NKT cells (i.e., NK1.1+ subset of CD3int cells) in the liver also showed a low mixture of partner cells. The present results raise the possibility that lymphocytes in the bone marrow and thymus, and extrathymic T cells in the liver might be in situ generated from their own preexisting precursor cells. Another observation was that, after irradiation, partner cells showed accelerated mixture even if they showed a low mixture under non-irradiated conditions. However, only lymphocyte subsets with the same phenotype as those of preexisting cells entered the corresponding sites.     

Distinctive functional properties of human blood L lymphocytes: a comparison with T lymphocytes, B lymphocytes, and monocytes.

Human blood lymphocytes with high affinity Fc receptors have been operationally named L lymphocytes because of membrane-labile IgG markers. L lymphocytes lack membrane-incorporated immunoglobulin and do not form rosettes with sheep red blood cells coated with IgM antibody and mouse complement. These lymphocytes are capable of binding IgG in normal human serum at 4 degrees C and will form rosettes with human lymphocytes coated with Ripley IgG. In this study, functional in vitro properties of isolated L lymphocytes were compared with T lymphocytes, B lymphocytes, and monocytes. To obtain these mononuclear populations, first, plastic adherent monocytes were harvested. T lymphocytes were then isolated by centrifugation of E rosette-forming cells, and other rosetting techniques were employed to isolate L and B lymphocytes by negative selection. The functional properties of L lumphocytes were completely unlike those of T cells, B cells, or monocytes. L lymphocytes did not proliferate in response to mitogens, soluble antigens, or cell surface antigens. Moreover, this population could not replace monocytes in helping T lymphocytes respond to concanavalin A and pokeweed mitogen. Once T cells were supplemented with monocytes, however, the addition of L lymphocytes to the culture greatly enhanced the T lymphocytes proliferative response to phytohemagglutinin, concanavalinA, purified protein derivative (PPD), and streptokinase/streptodornase. L lymphocytes were not a subset of B cells. They did not spontaneously develop surface Ig in culture, and pokeweek mitogen could not induce them to transform and generate cytoplasmic Ig detectable by immunofluorescence. Mixtures of B cells and T cells responded to pokeweed mitogen better than do T cells alone. In contrast, enhanced reactivity with L and T cell combinations was not observed. Another sharp difference between these two populations was the stimulator capacity of each in mixed lymphocyte culture. When B and L lymphocytes were carefully monocyte-depleted, only B cells were effective stimulators of autologous and allogeneic lymphocytes. In comparison with T cells, B cells, and monocytes, L lymphocytes were the only effective killers of human blood lymphocytes sensitized with IgG. L lymphocytes, then, have cytotoxic potential, but cannot proliferate in response to various stimulants or become antibody-producing cells. These findings suggest that L lymphocytes comprise a third lymphocyte population.