Differential effects of monoclonal antibodies anti-L3T4 and anti-LFA1 on the antigen-induced proliferation of T-helper-cell clones: correlation between their susceptibility to inhibition and their affinity for antigen

Recognition by specific T helper (TH) cells of antigen presented by antigen-presenting cells (APC) involves, in addition to the antigen-specific receptor, non-antigen-specific molecules such as L3T4 and LFA1. In the present study, we analyzed the relationship between the avidity for antigen presented by APC of three TH cell lines and the participation of L3T4 and LFA1 cell surface antigens. We found a correlation between the avidity of TH cells for the complex GAT/Ia on APC measured by two independent assays and the participation of the cell-adhesion molecules L3T4 as measured by the ability of corresponding monoclonal antibody (MAb) to block the antigen-induced proliferation of TH cells. In contrast to the situation found with cytolytic T-lymphocyte (CTL) clones, we also found a differential inhibiting effect of anti-LFA1 MAb on the GAT-specific proliferation of the three TH clones. The results indicate a direct correlation between the inhibitory effects of anti-LFA1 and anti-L3T4 MAb and the affinity of TH cells for the complex formed by antigen and Ia.     

Antigen presentation by EBV-B cells to resting and activated T cells: role of interleukin 1

We have previously demonstrated that Epstein Barr virus-transformed human B lymphocytes (EBV-B cells) present antigen to activated T cells (lines and clones) in a MHC-restricted manner. In the present study, using EBV-nonimmune donors, we demonstrate that EBV-B cells are unable to trigger tetanus toxoid (TT) antigen-specific proliferation in autologous highly purified resting T cells. EBV-B cells from these same individuals were able to present TT to autologous TT-specific activated T cell blasts (Tbl). The inability of EBV-B cells to present TT to resting T cells was not caused by defective antigen processing by EBV-B cells. Thus, paraformaldehyde treatment of antigen-pulsed EBV-B cells did not impair their ability to trigger proliferation of antigen-specific Tbl, and EBV-B cells pulsed with antigen in the presence of autologous TT-specific T cell blasts did not present antigen to resting T cells. Furthermore, antigen-specific proliferation of resting T cells triggered by monocytes was enhanced rather than suppressed by EBV-B cells. The addition of partially purified human IL 1 allowed EBV-B cells to present TT antigen to resting T cells, suggesting that failure to secrete IL 1 contributed to the failure of EBV-B cells to present antigen. IL 1 could not be detected in supernatants of EBV-B cells stimulated with Staphylococcus epidermidis, concanavalin A, and TT antigen in the presence or absence of up to 5% autologous T cells. The differential capacity of EBV-B cells to present antigen to resting T cells vs activated T cells correlated with the T cell requirement for IL 1, because a rabbit antibody to human IL 1 inhibited the monocyte-supported proliferation of resting T cells but not that of activated T cells. These results suggest that the inability of EBV-B cells to present antigen to resting T cells is related to their inability to secrete detectable IL 1.     

Antigen-specific and -nonspecific mitogenic signals in the activation of human T cell clones

We have directly compared the signals required for: induction of the [Ca+2]i response, expression of Tac antigen, and proliferation in antigen-specific human T cell clones. We have previously shown that antigen-specific activation of cloned T cells under conditions leading to proliferation is accompanied by a rapid increase in [Ca+2]i. Cloned T cells showed increased [Ca+2]i, enhanced Tac expression, and proliferated in response to specific antigen in the presence of viable, genetically appropriate antigen-presenting cells. Paraformaldehyde fixation of antigen-presenting cells after "pulsing" with antigen prevented proliferation, but did not affect MHC-restricted [Ca+2]i or Tac responses. Treatment of cloned T cells with monoclonal anti-T3 antibody also increased [Ca+2]i and Tac expression but did not induce proliferation. Proliferation was restored by viable autologous or allogenic APC or exogenous IL 2, but not by IL 1. In contrast to resting T cells, T cell clones were insensitive to the mitogenic effects of lectins or of ionophores and phorbol esters. These results suggest that activation of antigen-specific T cells requires the sequential action of at least two signals. The first is MHC restricted and is mediated by interaction of antigen + MHC class II products with the T cell receptor (T3-Ti) complex. This leads to Tac expression and increased [Ca+2]i, but is not sufficient for proliferation. This signal can be bypassed by anti-T3 monoclonal antibodies. Proliferation requires a second, nonantigen-specific, non-MHC-restricted antigen-presenting cell signal, which cannot be replaced by IL 1 in our system. This signal can be bypassed, however, by the addition of exogenous IL 2 to cells that have received the first signal and express Tac, suggesting that it is required for IL 2 synthesis and secretion. T cell clones therefore provide a useful model for studying antigen-dependent and -independent events in cell activation.     

Analysis of the role of leukocyte function-associated antigen-1 in activation of human influenza virus-specific T cell clones

The role of leukocyte function-associated Ag-1 (LFA-1) in intercellular adhesion is well documented. Previously, we demonstrated that the LFA-1 molecule (CD11a/CD18) can also regulate the induction of proliferation of peripheral blood T cells. In these studies, we observed opposite effects of antibodies against CD11a (LFA-1-alpha-chain) or CD18 (LFA-1-beta-chain). Here, we determined the effects of anti-CD11a and anti-CD18 mAb on proliferation of cloned influenza virus-specific T cells. Anti-CD18 mAb had similar inhibiting effects on the proliferative response of T cell clones induced by immobilized anti-CD3 mAb as it had on the response of peripheral blood T cells. In contrast to its costimulatory effect on resting peripheral blood T cells, anti-CD11a mAb did not increase the proliferation of cloned T cells. Similar differences in effects of anti-CD11a and anti-CD18 mAb were observed when proliferation of the T cell clones was induced by immobilized anti-TCR mAb. When proliferation was induced by influenza virus presented by monocytes as APC, both anti-CD11a and anti-CD18 mAb inhibited T cell proliferation. However, when EBV-transformed B cells were used as APC, neither anti-CD11a nor anti-CD18 mAb inhibited proliferation. These results demonstrate that the effects of antibodies against CD11a (LFA-1-alpha) or CD18 (LFA-1-beta) on T cell proliferation depend on 1) the stage of activation of the T cells, 2) the activation stimulus and its requirement for intercellular adhesion involving LFA-1, and 3) the type of cell used to present Ag.     

Requirements for T cell activation by OKT3 monoclonal antibody: role of modulation of T3 molecules and interleukin 1

The requirements for activation of human peripheral blood T cells by the mitogenic monoclonal antibody OKT3 were examined. OKT3 binds to a T cell molecule, T3, associated with the T cell antigen receptor and involved in T cell activation. Activation of T cells by OKT3 requires signals provided by accessory cells and is IL 2 dependent. In the presence of accessory cells, OKT3 induces loss of T3 molecules from the cell surface, production of IL 2, expression of IL 2 receptors, and proliferation. Modulation of T3 molecules by OKT3 can be induced in the absence of accessory cells with anti-mouse IgG. These T cells, however, are not induced to express IL 2 receptors or secrete IL 2. The addition of IL 1 induces expression of IL 2 receptors, but does not induce IL 2 secretion or proliferation. Thus, peripheral blood T cells appear to have different requirements for activation compared with antigen-specific T cell clones that can be induced to produce IL 2 when stimulated with OKT3 and IL 1. Expression of IL 2 receptors does not require modulation of T3 molecules, because the binding of OKT3 to T cells in the presence of IL 1 alone is sufficient to induce IL 2 receptor expression. The results suggest that IL 2 secretion depends on cross-linking and modulation of T3 molecules, and additional, as yet undefined, accessory cell signals. The expression of IL 2 receptors and proliferation of T cells can be induced in the absence of these signals when exogenous IL 2 is provided.     

Bronchoalveolar cells from sarcoid patients demonstrate enhanced antigen presentation

The recognition of foreign antigens by T lymphocytes in association with lung antigen-presenting cells may be critical in the initiation of the mononuclear alveolitis and granuloma formation of pulmonary sarcoidosis. However, it has been shown that bronchoalveolar cells (BAC) from normal volunteers function poorly as antigen-presenting cells. Therefore, the ability of sarcoid BAC to serve as accessory cells for antigen-dependent autologous T cell proliferation, as measured by tritiated thymidine uptake, was compared with that of normal BAC. Although irradiated sarcoid BAC supported antigen-induced T cell proliferation, normal BAC did so poorly (p less than 0.005). Because it has been shown that sarcoid BAC produce more interleukin 1 (IL 1) than normal BAC, it was considered that the enhancement of antigen-induced proliferative responses could result from an increased amount of IL 1, and that contaminating monocytes in the peripheral blood T cell preparations displayed the antigen for T cell recognition. Therefore, it was necessary to establish that antigen-induced T cell responses required HLA-D region compatibility between the sarcoid BAC and T lymphocytes. BAC from sarcoid patients stimulated antigen-specific proliferation in T cells lines matched for at least one HLA-D-region antigen, but failed to stimulate T cell lines that were unmatched for both antigens. This finding indicates that cells in bronchoalveolar lavage fluids from sarcoid patients were fully capable of acting as antigen-presenting cells. The identification of antigen-presenting cells in the lungs of patients with sarcoidosis together with the previous findings of activated T cells, enhanced IL 1 production, and spontaneous interleukin 2 release in sarcoid patients is compatible with the hypothesis that local cell-mediated immunity is involved in the pathogenesis of pulmonary sarcoidosis.     

Human alveolar macrophages: HLA-DR-positive macrophages that are poor stimulators of a primary mixed leukocyte reaction

Previous studies demonstrated that alveolar macrophages (AM) from most normal human volunteers failed to stimulate the antigen-induced proliferation of peripheral blood T lymphocytes although greater than 90% of AM expressed HLA-DR antigens. The current studies establish that AM also fail to induce allogeneic peripheral blood mononuclear cells to proliferate in a mixed leukocyte reaction (MLR). Suppressive activity by AM was not an explanation for their failure to induce an MLR. Indirect immunofluorescence established the presence of both HLA-DR and DQ antigens on the majority of AM and the persistence of these antigens on cells in culture for up to 6 days, the period of time required to observe a maximal MLR. Metabolic labeling experiments also demonstrated that HLA-DR antigens were synthesized by AM. It was recently reported that AM secrete relatively small amounts of IL 1, an important ancillary signal provided by accessory cells to enhance the stimulation of lymphocyte proliferation. However, addition of optimal concentrations of IL 1 to cultures containing AM failed to enhance the MLR. Thus, there is at least one additional, but as yet undefined, requirement for an accessory cell to induce an optimal MLR besides the display of HLA-D region antigens and the secretion of IL 1. In contrast, AM were effective in specifically stimulating proliferation of alloreactive T cell lines, suggesting that at least some cell lines do not require this nonspecific undefined second signal. We speculate that although AM may not initiate primary immune responses in the lung, they may be important in maintaining immune-mediated inflammatory responses by specifically restimulating already activated T cells.     

Antigen presentation by human antigen-presenting cells to antigen-specific xenogeneic murine T cells

Successful antigen presentation by xenogeneic human antigen-presenting cells (APC) to stimulate the proliferation of antigen-specific, keyhole limpet hemocyanin (KLH)-specific, ovalbumin (OVA)-specific, and purified protein derivative of Mycobacterium tuberculosis (PPD)-specific murine T cells was observed. Evidence indicating a direct cell interaction between antigen-specific murine T cells and xenogeneic human APC was given by experiments using antigen-specific murine T cell clones. The OVA-specific B10.S(9R) T cell line (9-0-A1) and PPD-specific B10.A(4R) T cell line (4-P-1) were stimulated by both xenogeneic human APC and murine APC from syngeneic or I-A compatible strains, while the PPD-specific human T cell line (Y-P-5) was stimulated by autologous human APC but not by murine APC. Anti-HLA-DR monoclonal antibodies (MoAb) blocked the xenogeneic human APC-antigen-specific murine T cell clone interaction. Thus, human xenogeneic APC can stimulate antigen-specific murine T cells through HLA-DR molecules in the same manner as syngeneic murine APC do through Ia molecules coded for by the I region of the H-2 complex, while murine APC failed to present antigen to stimulate human antigen-specific T cells.     

Development and characterization of interleukin-2-independent antigen-specific human T cell clones that produce multiple lymphokines

The development of antigen-specific T lymphocyte lines and clones has greatly facilitated the investigation of T-cell recognition of and response to foreign antigens. In the present study, human antigen-specific helper T cell lines and clones which are completely independent of exogenous interleukin-2 (IL-2) have been developed by cyclic restimulation with the soluble antigen keyhole limpet hemocyanin (KLH) to which the T cell donor had previously been immunized. These T cells uniformly bear the OKT4 phenotype and were shown to require both histocompatible antigen-presenting cells (APC) and antigen for optimal proliferation. The T cell line was composed of a highly antigen-specific and clonable T cell population. Following four cycles of antigen stimulation, limiting dilution cloning analysis showed a Poisson distribution of clonable T cells with a precursor frequency of 0.62, and from 88 to 92% of viable clones were specific for the stimulating antigen. Individual clones were obtained which recognized KLH with either DR 1 (one parental Ia haplotype of the donor) or DR 2 (the other parental Ia haplotype) allogeneic APC, but not both. Following stimulation with KLH, the T cell clones produced IL-2. Peak amounts of IL-2 were assayable in the first 6 to 24 hr after stimulation. In contrast, virtually no IL-2 was detectable in supernatants at 72 to 96 hr, suggesting autoutilization by the proliferating T cells. In addition, some clones were also capable of producing both B cell growth factor and IL-2 following KLH stimulation. These IL-2-independent T cells appeared to be derived from a discrete Leu 8-negative subclass of T4⁺ cells and expressed the full complement of Ia antigen of the donor. Thus, soluble antigen-specific human helper T cell clones have been produced which can be maintained in the absence of exogenous IL-2, elaborate their own growth factors and other immunoregulatory lymphokines, and show fine DR-related restriction to either one or the other parental DR haplotypes in antigen-stimulated proliferative responses.     

LPS and specific T cell responses: interleukin 1 (IL 1)-independent amplification of antigen-specific T helper (TH) cell proliferation

Lipopolysaccharide (LPS) fraction of endotoxin induces a significant potentiation of the antigen-specific proliferative response of T helper (TH) cell lines. This effect was obtained with LPS from different bacterial sources and reproduced with the lipid A moiety of endotoxin. Purified adherent spleen cells used as antigen-presenting cells (APC) support this LPS-enhanced TH cell proliferation. In addition, the effect of endotoxin on specific TH cell responses was found to be absolutely dependent on the interaction between TH lymphocytes and APC through antigen-specific recognition. Thus, it was not observed in the absence of specific antigen or when monoclonal antibodies against class II MHC products or against L3T4 antigens were used to inhibit the T cell-APC interaction. Similarly, it was found that APC from the B6.CH-2bm12 mutant do not support the LPS-mediated enhancing effect. Furthermore, interleukin 1 (IL 1) appears not to be involved in LPS-mediated enhancement, and this effect is not reproduced by muramyl dipeptide (MDP)-mediated activation of APC.     

Antigen processing requirements for T cell activation: differential requirements for presentation of soluble conventional antigen vs cell surface MHC determinants

The present studies were undertaken to characterize the antigen-processing requirements involved in the responses to T cells to soluble antigen (antigen specific), to allogeneic cell surface MHC determinants (alloreactive), and to syngeneic MHC determinants (autoreactive). T cell clones were used that have dual cross-reactive specificities either 1) for self MHC plus soluble antigen and for allogeneic MHC products or 2) for syngeneic MHC and for allogeneic MHC, in order to permit comparison of the processing requirements for responses of the same T cell to distinct antigenic stimuli. The proliferative responses of antigen-specific, Ia-restricted T cell clones to soluble antigens were sensitive to treatment of antigen-presenting cells (APC) with 125 to 250 microM chloroquine, a lysosomotropic agent previously shown to inhibit the processing of soluble antigens. In contrast, the same T cell clones were only minimally affected in their ability to respond to similarly chloroquine-treated APC expressing allogeneic MHC products. The responses of autoreactive T cell clones to syngeneic stimulating cells and their cross-reactive responses to allogeneic cells were both resistant to chloroquine treatment of stimulating cells. The failure of chloroquine to inhibit antigen presentation to autoreactive T cell clones suggests that these clones are specific for self Ia not associated with in vitro processed foreign antigen. Thus, chloroquine sensitivity distinguishes the in vitro antigen-processing requirements for presentation of the soluble antigens tested from the requirements for presentation of syngeneic or allogeneic cell surface MHC determinants to the same T cells.     

The relationship between immune interferon production and proliferation in antigen-specific, MHC-restricted T cell lines and clones

The release of immune or gamma interferon (IFN-gamma) by major histocompatibility complex (MHC)-restricted pigeon cytochrome c-specific Lyt 1+2-, interleukin 2 (IL 2)-producing proliferative T cell clones when cultured with antigen and antigen-presenting cells (APC) is a sensitive measure of the state of activation of the cell. In general, the fine specificity of T cell activation was similar when activation was measured either by IFN-gamma production or by proliferation. In response to antigen and the correct Ia molecule, the T cell clones produced both high titered IFN-gamma and a strong proliferative response. However, IFN-gamma production and the degree of proliferation of the T cell clones differed at high antigen concentrations. As antigen concentration increased, the magnitude of proliferation became submaximal whereas the IFN-gamma response became maximal suggesting that IFN-gamma produced by the cells might act as an autoregulatory molecule inhibiting the proliferative response. Stimulating the T cell to divide via its IL 2 receptor by adding exogenous IL 2 produced high levels of proliferation but only low titers of IFN-gamma activity. In addition, irradiation of the clone eliminated the IFN-gamma release induced by IL 2 but did not affect the IFN-gamma release induced by antigen and Ia. Thus proliferation is not essential for IFN-gamma production and unlike antigen and Ia, IL 2 functions predominantly as a proliferative signal and not as a signal for factor release. Two T cell clones showed a dissociation of IFN-gamma production and proliferation. In one case, a clone that proliferated in response to both allogeneic and antigenic stimuli released IFN-gamma in response to antigen but failed to produce IFN-gamma in response to the allogeneic stimulus. A second clone that showed a strong proliferative response to pigeon cytochrome c but no proliferative response to a species variant of cytochrome c, tobacco hornworm moth (THWM) cytochrome c, produced IFN-gamma when stimulated with either of these antigens. Thus, the sensitivity of detecting activation of T cell clones as measured by the release of an individual lymphokine varies from one clone to another.     

Monoclonal antibodies to CD2 and lymphocyte function-associated antigen 3 inhibit human thymic epithelial cell-dependent mature thymocyte activation

Recent study of human thymocyte-thymic epithelial (TE) cell interactions has demonstrated that thymocytes bind to TE cells, and a consequence of this binding is the provision of accessory cell signals by TE cells for phytohemagglutinin (PHA)-induced mature thymocyte activation. In this paper we report on studies of the molecules involved in TE cell-dependent mature thymocyte activation. TE-thymocyte interactions necessary for PHA-induced thymocyte activation were inhibited by monoclonal antibodies against the cluster of differentiation (CD)2 antigen on thymocytes and lymphocyte function-associated (LFA)-3 antigen on TE cells. Inhibition of TE accessory cell signals by antibodies against CD2 (alpha CD2) and LFA-3 (alpha LFA-3) antigens occurred early on during thymocyte activation and prevented thymocyte interleukin 2 receptor expression. Further, alpha CD2 and alpha LFA-3 inhibited PHA-induced thymocyte activation in whole thymic explant cultures suggesting a significant role of the CD2 and LFA-3 antigens in thymocyte activation when accessory cell signals for PHA-induced thymocyte triggering were delivered by cells within an intact thymic microenvironment.     

LFA-3 plasmid DNA enhances Ag-specific humoral- and cellular-mediated protective immunity against herpes simplex virus-2 in vivo: involvement of CD4+ T cells in protection

Adhesion molecules are important for cell trafficking and delivery of secondary signals for stimulation of T cells and antigen-presenting cells (APCs) in a variety of immune and inflammatory responses. Adhesion molecules lymphocyte function-associated antigen (LFA)-1 and CD2 on T cells recognize intercellular adhesion molecule (ICAM)-1 and LFA-3 on APCs, respectively. Recent studies have suggested that these molecules might play a regulatory role in antigen-specific immune responses. To investigate specific roles of adhesion molecules in immune induction we coimmunized LFA-3 and ICAM-1 cDNAs with a gD plasmid vaccine and then analyzed immune modulatory effects and protection against lethal herpes simplex virus (HSV)-2 challenge. We observed that gD-specific IgG production was enhanced by LFA-3 coinjection. However, little change in IgG production was observed by ICAM-1 coinjection. Furthermore, both Th1 and Th2 IgG isotype production was driven by LFA-3. LFA-3 also enhanced Th cell proliferative responses and production of interleukin (IL)-2, interferon-gamma, IL-4, and IL-10 from splenocytes. In contrast, ICAM-1 showed slightly increasing effects on T-cell proliferation responses and cytokine production. beta-Chemokine production (RANTES, MIP-1alpha, and MCP-1) was also influenced by LFA-3 or ICAM-1. When animals were challenged with a lethal dose of HSV-2, LFA-3-coimmunized animals exhibited an enhanced survival rate, as compared to animals given ICAM-1 or gD DNA vaccine alone. This enhanced protection appears to be mediated by CD4+ T cells, as determined by in vitro and in vivo T-cell subset deletion. These studies demonstrate that adhesion molecule LFA-3 can play an important role in generating protective antigen-specific immunity in the HSV model system through increased induction of CD4+ Th1 T-cell subset.     

The lymphocyte function-associated antigen (LFA)-1 and CD2/LFA-3 pathways of antigen-independent human T cell adhesion

Human cytotoxic T lymphocyte clones form conjugates with both antigen-positive and antigen-negative lymphoblastoid cells. Conjugates with antigen-negative targets form as rapidly, and are almost as frequent, as those with antigen-positive targets; both types are strong. Monoclonal antibodies against lymphocyte function-associated antigen (LFA)-1, CD2, and LFA-3 (or their Fab fragments) each consistently inhibit conjugate formation, but only partially; mixes of alpha LFA-1 with either CD2 monoclonal antibodies or alpha LFA-3 cause complete inhibition. Our previous studies have demonstrated two distinct pathways of antigen-independent conjugate (AIC) formation, one involving LFA-1 and the other involving CD2/LFA-3. The present studies showing supra-additive inhibition with mixes of Fab indicate that at least a major fraction of the conjugates involve T cells which utilize both pathways. Preincubation studies (and restricted expression for CD2) demonstrate that in the CD2/LFA-3 pathway, CD2 is critical on the effector and LFA-3 on the target and that in the LFA-1 pathway, LFA-1 is critical on the effector. Analysis of conjugate formation by primary allosensitized T cells confirms the critical findings made with T cell clones. Among a panel of antigen-negative "target" cell lines tested, there is wide variation in the number of AIC formed with cytotoxic T lymphocyte clones; this variation correlates partially with differences in level of expression of LFA-3. Both pathways of adhesion are utilized in AIC formation with all five targets tested, but there was variation between targets in the relative contribution by each pathway. Studies of inhibition of lysis (rather than conjugate formation) support the relevance of the two-pathway model to the lytic process as a whole. These studies demonstrate the general involvement of two pathways of adhesion in human T cell interactions: one involving T cell LFA-1 and the other involving T cell CD2 binding to target cell LFA-3.     

Antigen-specific inhibition of the IL-2-driven proliferation of myelin basic protein-reactive, human T cells

This report examines the antigen-specific inhibition of the IL-2-driven proliferation of autoantigen-reactive, human T cells. Human, myelin basic protein (MBP)-reactive CD4+ cell lines and clones were isolated and maintained in culture by use of IL-2 and periodic antigen stimulation. When freshly isolated antigen-presenting cells (APC) were present, MBP induced proliferation of MBP-reactive T cell populations. However, under different culture conditions, MBP reduced the IL-2-driven proliferation of some MBP-reactive T cell populations. The inhibition of IL-2-driven proliferation did not appear to require CD8+ or OKM 1+ cells since these were not detected when inhibition studies were performed at least 9 days after the last restimulation by irradiated APC and MBP. Supraoptimal concentrations of MBP were not required for inhibition of proliferation. Some heterogeneity of response was apparent since MBP inhibited the IL-2-driven proliferation of some T cell clones while for others MBP had either no effect or produced slight enhancement of proliferation. These results demonstrate an antigen-specific, in vitro immune mechanism that reduces the IL-2-dependent proliferation of autoantigen-reactive, human T cells.     

Role of LFA-1 and VLA-4 in the adhesion of cloned normal and LFA-1 (CD11/CD18)-deficient T cells to cultured endothelial cells. Indication for a new adhesion pathway.

Patients with the leukocyte adhesion deficiency (LAD) syndrome have a genetic defect in the common beta 2-chain (CD18) of the leukocyte integrins. This defect can result in the absence of cell surface expression of all three members of the leukocyte integrins. We investigated the capacity of T cell clones obtained from the blood of an LAD patient and of normal T cell clones to adhere to human umbilical vein endothelial cells (EC). Adhesion of the number of LAD T cells to unstimulated EC was approximately half of that of leukocyte function-associated antigen (LFA)-1+ T cells. Stimulation of EC with human rTNF-alpha resulted in an average 2- and 2.5-fold increase in adhesion of LFA-1+ and LFA-1- cells, respectively. This effect was maximal after 24 h and lasted for 48 to 72 h. The involvement of surface structures known to participate in cell adhesion (integrins, CD44) was tested by blocking studies with mAb directed against these structures. Adhesion of LFA-1+ T cells to unstimulated EC was inhibited (average inhibition of 58%) with mAb to CD11a or CD18. Considerably less inhibition of adhesion occurred with mAb to CD11a or CD18 (average inhibition, 20%) when LFA-1+ T cells were incubated with rTNF-alpha-stimulated EC. The adhesion of LFA-1- T cells to EC stimulated with rTNF-alpha, but not to unstimulated EC, was inhibited (average inhibition, 56%) by incubation with a mAb directed to very late antigen (VLA)-4 (CDw49d). In contrast to LAD T cell clones and the LFA-1+ T cell line Jurkat, mAb to VLA-4 did not inhibit adhesion of normal LFA-1+ T cell clones to EC, whether or not the EC had been stimulated with rTNF-alpha. We conclude that the adhesion molecule pair LFA-1/intercellular adhesion molecule (ICAM)-1 plays a major role in the adhesion of LFA-1+ T cell clones derived from normal individuals to unstimulated EC. Adhesion of LFA-1-T cells to TNF-alpha-stimulated EC is mediated by VLA-4/vascular cell adhesion molecule (VCAM)-1 interactions. Since we were unable to reduce significantly the adhesion of cultured normal LFA-1+ T cells to 24 h with TNF-alpha-stimulated endothelium with antibodies that block LFA-1/ICAM-1 or VLA-4/VCAM-1 interactions, and lectin adhesion molecule-1 and endothelial leukocyte adhesion molecule-1 appeared not to be implicated, other as yet undefined cell surface structures are likely to participate in T cell/EC interactions.     

Antigen presentation by human dermal fibroblasts: activation of resting T lymphocytes

We have shown that human dermal fibroblasts, exposed to interferon-gamma (IFN-gamma) to induce surface class II major histocompatibility complex (MHC) antigens, were capable of presenting tetanus toxoid (TT) antigen to human TT-specific T cell clones. Antigen presentation by fibroblasts was antigen dependent, required HLA-DR expression by fibroblasts, and was MHC restricted. In contrast, we now report that IFN-gamma-treated fibroblasts are unable to present TT antigen to purified resting T cells obtained from the peripheral blood of TT-immune donors. In addition, although IFN-gamma-treated fibroblasts were able to stimulate alloreactive T cell clones, they were unable by themselves to stimulate primary allogeneic responses in resting T cells. The failure of fibroblasts to stimulate resting T cells was not due to suppressor effects by fibroblasts, because induction of TT and alloantigen responses in resting T cells by monocytes was not inhibited by the presence of fibroblasts. On the contrary, IFN-treated fibroblasts were synergistic with small numbers of monocytes in activating resting T cells. In addition, the failure of antigen presentation by fibroblasts to resting T cells was reversed by the addition of recombinant human interleukin 2 (rIL 2) to cultures, but not of purified human interleukin 1 (IL 1). These results emphasize that the requirements for activation of resting T cells differ from those of T cell clones. Although fibroblasts can efficiently present antigen to T cell clones, antigen presentation by fibroblasts to resting T cells requires the addition of exogenous IL 2. It is postulated that fibroblasts differ from classical antigen-presenting cells in that fibroblasts are incapable of stimulating the production of IL 2 in resting T cells.     

T cell adhesion molecules

Cell adhesion or conjugate formation between T lymphocytes and other cells is an important early step in the generation of the immune response. Although the antigen-specific T cell receptor confers antigen recognition and specificity, a number of other molecules expressed on the T cell surface are involved in the regulation of lymphocyte adhesion. T cell molecules that function to strengthen adhesion include lymphocyte function-associated antigen (LFA)-1, CD2, CD4, and CD8. Their ligands have recently been identified. LFA-1 is a member of the integrin family of adhesion receptors and one of its ligands is intercellular adhesion molecule-1 (ICAM-1); a ligand for CD2 is LFA-3; and ligands for CD4 and CD8 appear to be major histocompatibility complex class II and class I molecules, respectively. In addition, T cells express a number of receptors thought to be involved in cell matrix adhesion. The function and significance of these T cell adhesion receptors and their ligands are reviewed.     

Role of the LFA-1 molecule in cellular interactions required for antibody production in humans

The lymphocyte function-associated antigen 1 (LFA-1) has been shown to play a role in various T cell functions in mice and humans including cytotoxicity, and proliferation to allogeneic cells and foreign antigens. These functions have been defined with specific monoclonal antibodies and were additionally confirmed by the investigation of patients with inherited deficiency in membrane LFA-1 expression. In this paper, we report our studies on the potential role of the LFA-1 molecule in T lymphocyte-dependent antibody responses. In a patient with a complete lack of membrane expression of LFA-1, there was no in vivo antibody response to vaccinal antigens such as tetanus, diphtheria toxoids, and polio virus, and no in vivo or in vitro antibody production to influenza virus, whereas serum immunoglobulin levels and antibodies to polysaccharides (isohemagglutinins, antibody to mannan, and a polysaccharide from Candida albicans) were detected in correlation with in vitro production of anti-mannan antibody. The defective antibody response to polypeptides was not secondary to poor antigen-specific T proliferation, because the latter was found to be present. Similarly, in vitro antibody production to influenza virus of normal cells was blocked by several anti LFA-1 monoclonal antibodies specific for the alpha subunit of the molecule, if they were added from the beginning of the culture. The antibody production blockade could be achieved with monoclonal antibody concentrations that partially preserved T cell proliferation. The helper effect of an influenza virus-specific helper T cell clone was also blocked. The targets of the blockade were shown by incubation experiments to be T cells and monocytes. In contrast, anti-LFA-1 monoclonal antibodies had no effect on pokeweed mitogen-induced B cell maturation into immunoglobulin-containing cells and on the anti-mannan antibody production. These combined data demonstrate that the LFA-1 molecule plays a role in T cell dependent antibody production to polypeptidic antigens but not in the antibody response to polysaccharides, although the antibody response to mannan is T cell dependent. It is proposed that the LFA-1 molecule is required to some extent for a antigen-presenting cells-T lymphocyte interaction and for the maintenance of a close association between antigen-specific helper T cells and small resting B lymphocytes. Polysaccharidic antigens that exhibit repetitive antigenic determinants might cross-link membrane immunoglobulins on B lymphocytes, thus allowing B cells to pass through a first step of activation requiring cognate T-B cell interaction.