Induction of suppressor activity in the autologous mixed lymphocyte reaction and in cultures with concanavalin A.

T lymphocytes that are activated in the autologous mixed lymphocyte reaction (MLR) have suppressor activity. Concanavalin A (Con A) augments the suppressor activity generated in cultures containing both T and non-T lymphocytes and can induce suppressor activity in T-lymphocyte preparations that contain too few (10%) non-T cells to generate a significant autologous MLR. However, when such T-lymphocyte preparations are further depleted of adherent cells and contain less than 2% non-T cells, Con A fails to induce suppressor activity. These findings support the concept that an autologous MLR may play an important role in generation of suppressor cells by Con A.     

Effect of oral colchicine on T cell subsets, monocytes and concanavalin A-induced suppressor cell function in asthmatic patients

Asthmatic patients have a deficiency of concanavalin A-(Con A) induced suppressor cell function. We tested whether oral colchicine 0.5 mg twice daily for 7 days could correct this immunoregulatory abnormality. Peripheral blood mononuclear cells were incubated with Con A and then suppression of proliferation was measured by coculture of these cells with healthy volunteers' mononuclear cells and phytohaemagglutinin. Sixteen asthmatic patients had significantly (P less than 0.002) decreased Con A-induced suppressor cell function (17.0 +/- 17.2%, mean +/- s.d.) as compared to 13 healthy volunteers (37.9 +/- 14.9%). Oral colchicine significantly (P less than 0.05) increased, though only partially corrected, these 16 asthmatic patients' Con A-induced suppressor cell function (28.1 +/- 14.3%). Asthmatic patients had an increased number of monocytes (691 +/- 289 vs 388 +/- 271/mm3 for normals, P less than 0.01) and a normal number of lymphocytes, Leu 4+ total T cells, Leu 3+ helper/inducer T cells, and Leu 2+ suppressor/cytotoxic T cells as well as a normal Leu 3/Leu 2 ratio. Oral colchicine significantly (P less than 0.005) decreased the number of monocytes (451 +/- 255/mm3) without significantly affecting the number of lymphocytes, Leu 4+, Leu 3+, or Leu 2+ T cells, or the Leu 3/Leu 2 ratio. These results are consistent with the hypothesis that the deficiency of Con A-induced suppressor cell function in asthmatic patients may be due, in part, to an increased number and/or abnormal activity of monocytes. If so, then oral colchicine may have partially corrected the deficiency of Con A-induced suppressor cell function by decreasing the number and/or modulating the activity of monocytes.     

Defective production of monocyte-activating cytokines in lepromatous leprosy

We have examined the capacity of monocytes from patients with leprosy to undergo activation and the capacity of mononuclear cells from these patients to incorporate [3H]thymidine and produce monocyte-activating cytokines. Monocytes from patients with either lepromatous or tuberculoid leprosy were activated by concanavalin A (Con A)-induced mononuclear cell supernatants generated from the leukocytes of a normal person. Monocytes activated by these supernatants strongly inhibited L. pneumophila multiplication, and the degree of inhibition was comparable in both groups of patients. Mononuclear cells from patients with either form of leprosy responded comparably to Con A with vigorous [3H]thymidine incorporation. Mononuclear cells from patients with tuberculoid leprosy also vigorously incorporated [3H]thymidine in response to M. leprae antigens. In contrast, mononuclear cells from patients with lepromatous leprosy did not exhibit significant [3H]thymidine incorporation in response to M. leprae antigens. The capacity of mononuclear cells to generate monocyte-activating cytokines generally paralleled their capacity to incorporate [3H]thymidine in response to Con A and M. leprae. Mononuclear cells from patients with either form of leprosy responded to Con A with the production of cytokines (supernatants) able to activate normal monocytes, expressed by inhibition of L. pneumophila multiplication. However Con A-induced supernatants from patients with lepromatous leprosy were less potent than Con A-induced supernatants from patients with tuberculoid leprosy. Mononuclear cells from patients with tuberculoid leprosy responded to M. leprae antigens with the production of potent monocyte-activating supernatants. In contrast, mononuclear cells from patients with lepromatous leprosy did not produce monocyte-activating cytokines in response to M. leprae antigens. These studies support the hypothesis that the immunological defect in lepromatous leprosy results from a failure to activate mononuclear phagocytes rather than from an intrinsic inability of these cells to be activated. We suggest that the failure to activate mononuclear phagocytes stems from defective production of monocyte-activating cytokines in response to M. leprae antigens.     

Separation of concanavalin A-induced human suppressor and helper T cells by the autologous erythrocyte rosette technique.

Very few normal human peripheral blood T cells are capable of binding autologous erythrocytes to form rosettes, whereas in the T cell population activated by concanavalin A (Con A) the autorosette levels are markedly enhanced. Fractionation of the Con A-activated T cells with autologous erythrocytes into autorosetting and nonrosetting cells demonstrates that suppressor, but not helper, activity resides in the autorosetting population, whereas the reverse is true of the nonrosetting population. Both these activities are found to be Con A dependent. The Con A-induced human suppressor cells can be identified and separated from the Con A-induced human helper cells by the autorosette technique. Studies on the surface properties of autorosetting and nonrosetting T cells indicate that there is little correlation between the activated suppressor and helper T cell subsets defined by autorosette technique and either those defined by monoclonal antibodies (which are able to distinguish these subsets in the resting but not activated T cells) or those defined by Fc receptors. Since the autorosetting T cell population (which acts as suppressor cells) bears receptors for peanut agglutinin, the nature of Con A-induced human suppressor cells appears to be analogous to that of Con A-induced murine suppressor cells.     

Identification and characterization of a monocyte-derived neutrophil-activating factor in corticosteroid-resistant bronchial asthma

Peripheral blood mononuclear cells (PBMC) were isolated from seven normal subjects, eight asthmatic subjects clinically sensitive to corticosteroids (CS), and eight asthmatic subjects clinically resistant to corticosteroids (CR). PBMC were cultured at 37 degrees C for 24 h in the absence or presence of 10(-16) to 10(-4) M hydrocortisone. Calcium ionophore (A23187)-activated neutrophils (PMN) primed by supernatants of PBMC from asthmatic subjects cultured in the absence of hydrocortisone generated approximately threefold more leukotriene B4 than PMN primed by supernatants of PBMC from normal subjects (P less than 0.05). Incubation of PBMC derived from CS subjects with 10(-8) M hydrocortisone completely inhibited the production of the enhancing activity (P less than 0.01), whereas in CR subjects hydrocortisone at concentrations up to 10(-4) M did not suppress the release of enhancing activity. The enhancing activity was produced by monocytes. Enhancing activity eluted with an Mr of 3,000 D and a pI of 7.1. It eluted at 10% acetonitrile after reverse-phase HPLC. The activity was destroyed by heating to 60 degrees C for 60 min and was sensitive to pronase treatment. The purified factor also enhanced superoxide generation by PMN which had been stimulated submaximally by phorbol myristate acetate.     

Immunomodulatory effect of procainamide in man. Inhibition of human suppressor T-cell activity in vitro.

Procainamide (PA) induces the production of a number of autoantibodies in a high proportion of treated individuals and in some a syndrome closely resembling systemic lupus erythematosus. The mechanism underlying this action of PA is unclear. To examine the possibility that PA might induce autoantibody formation by altering normal immunoregulatory mechanisms, the action of this drug on an in vitro model of antibody formation in man was examined. PA was found to augment the generation of immunoglobulin-secreting cells (ISC) from human peripheral blood mononuclear cells (PBM) in response to pokeweed mitogen but had no effect on pokeweed mitogen-induced tritiated thymidine incorporation. When purified populations of B and T cells were used, PA enhanced the generation of ISC in B-cell cultures supported by untreated T cells but not by T cells treated with mitomycin C. These results indicate that PA augmented B-cell responses by inhibiting suppressor T-cell activity and not by augmenting helper T-cell or B-cell function. N-Acetyl-procainamide had no effect on the generation of ISC in this system. The effect of PA on concanavalin A (Con A)-induced suppressor cell activity was also examined to determine whether PA altered the generation or expression of suppressor T-cell function. PBM were cultured with 30 microgram/ml of Con A for 48 h to generate suppressor cells. When these were co-cultured with fresh PBM, the number of ISC generated was decreased by 58.1 +/- 3.4% (mean +/- SEM, n = 6). Cells that had been similarly incubated without Con A were not inhibitory. The addition of PA to the Con A-stimulated cultures inhibited the generation of suppressor cells as indicated by the fact that the response of fresh cells co-cultured with the Con A-stimulated cells was diminished by only 27.2 +/- 4.3%. In this system too, N-acetyl-procaimamide had no effect. By contrast, adding PA only to the co-culture of Con A-stimulated cells with fresh PBM had a less marked effect on suppressor cell function. These results indicate that the major action of PA is to inhibit the generation of suppressor T-cell activity. Such an effect may explain the capacity of this agent to induce autoantibody formation in treated individuals.     

Suppressor T cell numbers and function in atopic patients with high serum IgE levels

Suppressor T cell function and the sensitivity of lymphocyte transformation to histamine has been studied in 9 atopic patients with high serum IgE levels and in 14 controls. The effect of concanavalin A (Con A) induced suppressor T cells on the proliferative response of fresh lymphocytes to mitogens and antigens was measured; in atopic subjects the median suppression to 5.0 micrograms/ml Con A was 80% to 1.0 microgram/ml Phytohaemagglutinin (PHA) was 73%, to Dermatophagoides pteronyssinus extract was 45% and to streptokinase--streptodornase was 24%. Indomethacin increased lymphocyte proliferation to mitogens, but to a variable degree, in both groups. Histamine suppressed lymphocyte transformation to PHA and Con A (median suppression in normal subjects 38 and 46%, and in atopics 51 and 60%) and to D. Pteronyssinus extract. There was no significant difference between normal subjects and atopics in any of these functional assays. The relative and absolute numbers of total T cells, helper T cells and suppressor T cells measured by monoclonal antibodies and the helper to suppressor T cell ratio were normal in the atopic group. These results show that the activity of Con A induced suppressor T cells and the effect of histamine and indomethacin on lymphocyte proliferation is normal in highly atopic subjects. No suppressor T cell defect has been identified using these assays.     

Effect of BM 12 531 on in vitro induction of suppressor cells by concanavalin A in cancer patients and normal healthy donors

The effects of BM 12 531, a 2-cyanaziridinyl derivative, on in vitro generation of Con A induced suppressor cells as well as generation of spontaneous suppressor cells in peripheral blood lymphocytes of 9 chronic myeloid leukemia (CML) patients in remission, 9 patients with active Hodgkin's disease (HD) and 12 normal healthy donors were studied. The suppressor cells generated spontaneously and with Con A, in the presence and absence of the drug, were tested for their modulating effect on mitogenic (PHA) response of autologous lymphocytes. The results indicate that the addition of the drug at the time of generation of suppressor cells decreased the spontaneous suppressor cell activity in 2/4 healthy donors, 3/7 CML patients and 4/6 HD patients. Con A induced suppressor cell activity generated in presence of the drug was significantly reduced in 7/12 healthy donors, 6/9 CML patients and 3/9 HD patients.     

Histamine regulation of interleukin-18-initiating cytokine cascade is associated with down-regulation of intercellular adhesion molecule-1 expression in human peripheral blood mononuclear cells.

In the previous study, we demonstrated that interleukin (IL)-18 up-regulated intercellular adhesion molecule-1 (ICAM-1) expression on monocytes in human peripheral blood mononuclear cells (PBMC) and that heterotypic interaction between monocytes/T or NK cells through ICAM-1/LFA-1 intensified the production of IL-12, interferon-gamma (IFN-gamma), and tumor necrosis factor-alpha (TNF-alpha) in PBMC. In the present study, we demonstrate that histamine inhibited the ICAM-1 expression in monocytes induced by IL-18 using flow cytometry and that the responses of IL-12, IFN-gamma, and TNF-alpha induced by IL-18 were concentration dependently inhibited by coexisting histamine, whereas IL-18-inhibited IL-10 production was reversed by the same concentrations of histamine. The modulatory effects of histamine on ICAM-1 expression and cytokine production were all concentration dependently antagonized by famotidine but not by d-chlorpheniramine and thioperamide, and were mimicked by selective H(2)-receptor agonists but not by H(1)- and H(3)-receptor agonists, indicating the involvement of H(2)-receptors in histamine action. The inhibition of IL-18-induced IFN-gamma by histamine was ascribed to the strong inhibition of IL-12 production by histamine. Histamine thus operates the negative feedback mechanism against IL-18-activated cytokine cascade through the strong inhibitory effect on ICAM-1 expression and IL-12 production in monocytes, contributing to the formation of diverse pattern of cytokine activation from Th1 to Th2, depending on the monocyte/macrophage activation and cytokine environment.     

Normal T cell regulation of IgG synthesis in systemic lupus erythematosus

The regulation of pokeweed mitogen (PWM) stimulated IgG synthesis by peripheral blood mononuclear cells (PBL) was studied in patients with systemic lupus erythematosus (SLE) and controls. The helper-T, suppressor-T and B cell functions of SLE-PBL were separately evaluated and compared to the functions of control lymphocyte populations. SLE-T cells function normally both in helper and suppressor modulation of PWM-stimulated IgG secretion by control or autologous B cells. The SLE-B cell, however, fails to synthesize normal quantities of IgG when cultured under conditions that induce maximal stimulation. While other studies document abnormal Con A induced suppressor cell function in SLE, this study shows that normal functioning populations of SLE-suppressor T cells exist in the peripheral blood.     

Inhibition of humoral and cell-mediated immune responses in man by distinct suppressor cell systems.

Studies were designed to investigate whether the suppressor cell systems that regulate the humoral and cell-mediated immune responses belong to the same subsets of T cells or different subsets. Mitogen-activated suppressor cells were simultaneously assayed for their ability to inhibit (a) pokeweed mitogen-induced generation of plasma cells, (b) blastogenic response of lymphocytes to allogeneic cells, and (c) generation of killer cells in the cell-mediated lymphocytotoxicity assay. We found that suppressor cells that inhibited the generation of plasma cells were activated by concanavalin A (Con A) and were both radiation and prednisone sensitive. Suppressors that inhibited the blastogenic response in lymphocytes to allogenic cells were also activated by Con A but differed in that they were both radiation and prednisone resistant. In contrast, suppressors that inhibited the generation of the killer cells were activated with phytohemagglutinin and not Con A. These suppressors were prednisone and radiation resistant. These observations cannot be explained by differences at the pro-suppressor or suppressor activator levels as both T cell subsets are radiosensitive. Alternatively, heterogeneity of suppressor cell systems may explain these differences.     

Bovine aortic endothelial cells elaborate an inhibitor of the generation of lipopolysaccharide-stimulated human blood monocyte procoagulant activity.

We examined the effect of bovine aortic endothelial cell culture supernatants upon the generation of procoagulant activity by human blood monocytes. Confluent endothelial monolayers were cultured for up to 96 h. At timed intervals, culture supernatants were collected and incubated for 5 h with lipopolysaccharide-stimulated human peripheral blood mononuclear cells. The procoagulant activity of mononuclear cell lysates was determined in a one-stage clotting assay. In five experiments, procoagulant activity with culture supernatant (time 0) was 2,294 +/- 761 U/ml (mean +/- SEM). Culture supernatants from endothelial cells incubated for 24-96 h strongly inhibited mononuclear cell generation of procoagulant activity. Indomethacin (10 microM) added to endothelial cells delayed the appearance of procoagulant inhibitor for 72 h. Bovine aortic smooth muscle cell culture supernatants did not inhibit procoagulant activity. The inhibitor was heat stable, effective at 1:50 dilution, soluble, and acid sensitive, with a molecular weight of less than 1,500. Further studies on subpopulations of mononuclear cells demonstrated that endothelial inhibitor selectively decreased the generation of monocyte procoagulant activity and interfered with T lymphocyte amplification of monocyte production of procoagulant activity. Thus, we have demonstrated that endothelial cells elaborate a potent inhibitor of monocyte procoagulant activity.     

Differential processing of proenkephalin-A by human peripheral blood monocytes and T lymphocytes.

Human peripheral blood mononuclear cells are analyzed for preproenkephalin gene expression and peptide processing. Met-enkephalin immunoreactivity as detected with a specific antiserum is found in the cytoplasm of monocytes but not in T lymphocytes. Secretion of met-enkephalin was analyzed with an RIA that is specific for the met-enkephalin pentapeptide. Unfractionated PBMC spontaneously released 40 pg/ml met-enkephalin and this increased two- to fourfold after stimulation with PHA. Lower levels (less than 100 pg/ml) of met-enkephalin were detected in supernatants from purified T cells that were activated with PHA and IL-2. In contrast, stimulation of purified monocytes with LPS or PMA resulted in the release of up to 600 pg/ml of the processed peptide. To examine whether T cells can produce met-enkephalin precursor peptides, T cell conditioned media were treated with trypsin and carboxypeptidase-B, which is known to release met-enkephalin from the propeptide. This increased levels of met-enkephalin to 400 pg/ml, indicating that lymphocytes secrete the propeptide but do not process it to met-enkephalin. The 1.4-kb preproenkephalin mRNA is detected in activated blood mononuclear cells and in purified monocytes and T cells. To determine whether monocytes or lymphocytes express met-enkephalin in vivo, lymphoid tissues were analyzed by immunohistochemistry. In human spleen tissue, positive cells were found in the red pulp but not in the follicles, which is also consistent with met-enkephalin expression in monocytes. In summary, these results show that human peripheral blood mononuclear cells express preproenkephalin mRNA and that monocytes, but not T cells, process the propeptide to metenkephalin.     

Activation of infectious virus from latent human immunodeficiency virus infection of monocytes in vivo.

Individuals infected with HIV may be asymptomatic for years before progressing to overt AIDS. Since HIV can latently infect monocytoid cell lines, we examined whether HIV latency occurs in monocytes in vivo. Freshly isolated monocytes from asymptomatic seropositive individuals examined before and after culture were positive for HIV DNA, but not RNA, as measured by polymerase chain reaction, showing that HIV latency occurs in monocytes in vivo. Coculture of these latently infected monocytes with Con A-activated T cells from HIV-negative normal donors stimulated 90% of the patients' samples and latently infected THP-1 to produce infectious virus. Neither Con A, resting T cells, nor T cell supernatants induced virus. Plasma membranes from activated T cells stimulated HIV production, suggesting cell contact induces factor(s) in monocytes to overcome latency. Thus, monocytes in AIDS patients harbor latent HIV inducible during an immune response, leading to T cell infection and viral-induced pathology.     

Cellular defenses against Toxoplasma gondii in newborns.

Mononuclear phagocytes, particularly macrophages (M phi) that have been activated by lymphokines, are the principal defense against intracellular pathogens such as Toxoplasma gondii. To determine reasons for the newborns' susceptibility to Toxoplasma infection, we compared: the interaction of Toxoplasma with newborns' mononuclear phagocytes (blood monocytes and two types of newborn M phi, those derived from blood monocytes or from placental tissue) with adults' blood monocytes and monocyte-derived M phi and the production of M phi-activating lymphokines (MAF) by Concanavalin A (ConA)-stimulated newborn and adult blood mononuclear cells (MC). Newborn and adult monocytes killed Toxoplasma with equal efficiency. Similarly, survival and replication of Toxoplasma were comparable in control newborn and adult M phi. Exposure to adult ConA supernatants significantly decreased the survival and replication of Toxoplasma both in adult and newborn M phi. In contrast, exposure to cord blood ConA supernatants failed to affect the survival or the replication of Toxoplasma in newborn M phi and decreased the replication but not the survival of Toxoplasma in adult M phi. Exposure to ConA supernatants of peripheral blood MC from 2-5-d old newborns failed to affect survival or replication of Toxoplasma in newborn or adult M phi. Thus, both generation of MAF by newborn blood MC and response to newborn MAF by newborn M phi were impaired. Generation of MAF by adult blood mononuclear cells was not inhibited by cord blood MC nor was generation of MAF by cord blood MC increased by depletion of OKT8 antibody-binding cells, by depletion of adherent cells with or without addition of adult adherent cells, or by addition of indomethacin. Depletion of OKT4 antibody-binding cells abrogated the generation of MAF both by adult and cord blood MC. The activity of adult ConA supernatants was abrogated by dialysis at pH 2 or by addition of anti-gamma-interferon but not anti-alpha-interferon antibody. However, the correlation between antiviral interferon activity and anti-Toxoplasma activity was weak (r = 0.40). Enhanced M phi anti-Toxoplasma activity was not associated with detectably enhanced superoxide anion generation, nitroblue tetrazolium reduction, or phagolysosome fusion, and was not inhibited by catalase, superoxide dismutase, or mannitol. These results indicate that generation of and response to MAF is decreased in cells from human newborns and that gamma-interferon may be the major MAF under these conditions.     

CELL-MEDIATED IMMUNE RESPONSES IN VITRO : I. SUPPRESSION OF THE GENERATION OF CYTOTOXIC LYMPHOCYTES BY CONCANAVALIN A AND CONCANAVALIN A-ACTIVATED SPLEEN CELLS

The effects of soluble concanavalin A (Con A) or Con A-activated spleen cells on the generation of cytotoxic lymphocytes (CL) in mixed leukocyte cultures (MLC) were examined. Mitogenic concentrations of soluble Con A or small numbers of Con A-activated spleen cells substantially inhibited CL responses. The suppression was partial rather than absolute and was critically dependent upon the concentration and time of addition of soluble Con A or Con A-activated spleen cells to the MLC. Suppressive effects of Con-A activated spleen cells were mediated by T cells since suppressor cell activity was abrogated by treatment of spleen cells with anti-θ serum and complement before or after Con A activation. X irradiation of spleen cells before Con A treatment also abrogated generation of suppressor cell activity. After activation by Con A, however, the function of suppressor cells was radioresistant. Although the precise mechanism(s) of suppression is, as yet, unknown, the precursors of CL must be exposed to Con A-activated cells during the early phases of the immune response for suppression to occur. Kinetic studies revealed that suppression of CL responses was not due to a failure to initiate an immune response, but represented a response which developed initially, but subsequently aborted. The relevance of these observations to the concepts of T-cell-T-cell interaction and regulatory control of immune responses by T cells is discussed.     

Decreased in vitro humoral immune responses in aged humans.

Induction of antigen-specific and non-specific (polyclonal) humoral immune responses in vitro was investigated in peripheral blood mononuclear cells of aged (65-85 yr) and young (20-30 yr) volunteers. In vitro immunization of lymphocytes with antigen (sheep erythrocytes) was performed in a recently described microculture system, and anti-sheep erythrocyte plaque forming cells were quantitated in a direct hemolytic plaque assay. Immunoglobulin secreting cells, induced polyclonally with pokeweed mitogen, were quantitated in a reverse hemolytic plaque assay. Significant depressions of antigen-specific as well as polyclonal responses were noted in relation to advancing age. Antigen-specific responses were more frequently depressed than polyclonal responses. T cell mitogen concanavalin A (Con A) was used to amplify functions of autologous immunoregulatory T cells. Addition of 10 microgram/ml Con A to lymphocytes of young donors at culture initiation resulted in activation of suppressor cells and abrogated antigen-specific responses. Delayed addition of Con A, on the other hand, enhanced responses, presumably because of activation of helper T cells. Similar manipulations of lymphocyte cultures from aged donors showed failure of Con A to suppress antigen-specific responses in approximately half of the responders. In many nonresponders, responses within normal range were elicited by the delayed addition of Con A to their lymphocyte cultures. Deviations beyond the range of expected responses were noted in 32.5% of the co-cultures between pokeweed mitogen stimulated young and aged cells. Our findings suggest that age-related deficiencies of B cell function are frequently associated with dysfunction of immunoregulatory T cells and are only occasionally due to intrinsic defects of B cells.     

Comparison of interferon-γ and interleukin-4 production by peripheral blood mononuclear cells and isolated T cells after activation with polyclonal t cell activators

Controversial data have been reported regarding the ability of peripheral blood T cells to secrete interferon-γ (IFN-γ) and interleukin-4 (IL-4) from atopic patients as compared to nonatopic healthy controls. In most of these studies, T cells in peripheral blood mononuclear cell preparations (PBMC) were stimulated with polyclonal T cell activators. Some of these activators are able to activate cells other than T cells in the PBMC preparations which may influence the lymphokine levels in supernatants of PBMC. To evaluate this, we compared the IFN-γ and IL-4 levels in PBMC and isolated T cell preparations after activation with phytohemagglutinin (PHA), Concanavalin A (ConA), anti-CD3 plus phorbol myristate acetate (PMA), or ionomycin plus PMA. The IFN-γ and IL-4 levels in the supernatants were calculated based on the percent T cells in the preparations. Whereas all activators induced significant IFN-γ secretion, only ionomycin plus PMA stimulation induced large IL-4 secretion. In virtually all cases, the IFN-γ levels calculated on a per T cell basis differed for PBMC versus isolated T cells. Whereas in some donors the IFN-γ levels were higher in PBMC preparations than in T cells, in others it was the opposite. Similarly, in about one half of both normal and atopic donors tested, the IL-4 levels of activated PBMC were 2- to 7-fold lower than levels in isolated T cells. The data suggest that non-T cells have a significant effect on the IFN-γ and IL-4 levels in supernatants of polyclonally activated PBMC. This indicates that isolated T cells rather than PBMC should be analyzed for determining possible differences in the ability of T cells from patients or normals to secrete lymphokines. © 1994 Wiley-Liss, Inc.     

Keratinocyte growth regulation by the products of immune cells

We describe a bioassay that allows the in vitro investigation of the stimulatory and suppressive factors derived from immune cells in short-term cultures of human keratinocytes. In agreement with other assays, epidermal growth factor is not mitogenic for human keratinocytes. Supernatant fluid from human PBMC stimulated with Con A, from allo-MLRs, as well as supernatants from nonstimulated PBMC, possess growth-promoting molecules. Our results show that both activated and nonactivated T cells release growth factors. Suppressive molecules are produced preferentially by monocyte cultures. Two T cell products, IFN-gamma and transforming growth factor beta are both inhibitory for keratinocyte proliferation. Two other T cell products, IL-3 and GM-CSF, stimulate keratinocyte proliferation at nanogram concentrations. These results suggest the existence of regulatory circuits between the T cells of a dermal inflammatory infiltrate and the overlying epidermal keratinocytes. This may determine the fine control of epidermal proliferation and turnover leading either to enhanced wound repair or skin pathology.