Intercellular adhesion molecule 1 deficiency leads to impaired recruitment of T lymphocytes and enhanced host susceptibility to infection with Trypanosoma cruzi

In this study, we investigated the involvement of Th1 cytokines in the expression of cell adhesion molecules (CAM) and recruitment of inflammatory cells to the heart of mice infected with Trypanosoma cruzi. Our results show that endogenously produced IFN-gamma is essential to induce optimal expression of VCAM-1 and ICAM-1 on the cardiac vascular endothelium of infected mice. Furthermore, the influx of inflammatory cells into the cardiac tissue was impaired in Th1 cytokine-deficient infected mice, paralleling the intensity of VCAM-1 and ICAM-1 expression on the vascular endothelium. Consistent with the importance of ICAM-1 in host resistance, ICAM-1 knockout (KO) mice were highly susceptible to T. cruzi infection, as assessed by mortality rate, parasitemia, and heart tissue parasitism. The enhanced parasitism was associated with a decrease in the numbers of CD4(+) and CD8(+) T lymphocytes in the heart tissue of ICAM-1 KO mice. Additionally, ICAM-1 KO mice mounted an unimpaired IFN-gamma response and IFN-gamma-dependent production of reactive nitrogen intermediates and parasite- specific IgG2a. Supporting the participation of ICAM-1 in cell migration during T. cruzi infection, the entrance of adoptively transferred PBL from T. cruzi-infected wild-type C57BL/6 mice into the cardiac tissue of ICAM-1 KO mice was significantly abrogated. Therefore, we favor the hypothesis that ICAM-1 plays a crucial role in T lymphocyte recruitment to the cardiac tissue and host susceptibility during T. cruzi infection.     

Differential utilization of ICAM-1 and VCAM-1 during the adhesion and transendothelial migration of human T lymphocytes.

The comparative roles of the endothelial cell (EC) adhesion receptors VCAM-1 and ICAM-1 during the adhesion and transendothelial migration of T cells were examined. The adhesion of T cells to IL-1-activated EC was markedly, but not completely, inhibited by mAb to VCAM-1 as well as to its counter-receptor, VLA-4, whereas, T cell binding to IL-1-activated EC was not blocked by mAb to ICAM-1 or to its counter-receptor, LFA-1. In contrast, LFA-1/ICAM-1, but not VLA-4/VCAM-1, mediated much, but not all, of the binding of T cells to unstimulated EC. Activation of T cells with phorbol dibutyrate and ionomycin alter the receptor-counter-receptor pairs used for binding to EC. Regardless of the activation status of the EC, the binding of activated T cells was not blocked by mAb to VLA-4 or VCAM-1. Moreover, the binding of activated T cells to EC was blocked to a lesser degree by mAb to LFA-1 than that of resting T cells, and mAb to ICAM-1 blocked binding only modestly. The role of VCAM-1 and ICAM-1 during the transendothelial migration of T cells was also examined. Regardless of the activation status of the T cells or the EC, VCAM-1 was never found to function during transendothelial migration, even when it mediated the binding of resting T cells to IL-1-activated EC. In contrast, ICAM-1 played an important role in transendothelial migration under all of the conditions examined, including situations when T cell-EC binding was not mediated by ICAM-1. Immunoelectron microscopic analysis of transendothelial migration supported the conclusion that ICAM-1 but not VCAM-1 played a central role in this process. Thus, ICAM-1 was prominently and uniformly expressed at all EC membrane sites that were in contact with bound and migrating T cells, whereas VCAM-1 was localized to the luminal surface of IL-1-activated EC, but was often absent from the surface of the EC in contact with T cells undergoing transendothelial migration. These studies confirm that ICAM-1 and VCAM-1 play reciprocal roles in the binding of resting T cells to resting and IL-1-activated EC, respectively, but a less prominent role in the binding of activated T cells. Moreover, ICAM-1 but not VCAM-1 plays a role in transendothelial migration, regardless of the receptor-counter-receptor pairs used for initial binding.     

In vivo effects of monoclonal antibody to ICAM-1 (CD54) in nonhuman primates with renal allografts

These studies test whether allograft rejection can be blocked by interference with leukocyte adhesion, using a murine IgG2a mAb (R6.5) reactive with monkey ICAM-1 (CD54). In 16 Cynomolgus renal allograft recipients, R6.5 was administered prophylactically as the sole immunosuppressive agent for 12 days (0.01 to 2 mg/kg/day). Survival in 14 recipients with technically successful grafts was significantly prolonged (24.2 +/- 2.4 vs 9.2 +/- 0.6 days for controls; p less than 0.001). Intercellular adhesion molecule-1 (CD54) (ICAM-1) was expressed on vascular endothelium in the kidney and other organs in the monkey in a pattern similar to that in humans. During cellular rejection in controls, ICAM-1 expression increased on endothelial cells, infiltrating mononuclear leukocytes and tubular cells. Biopsies during R6.5 administration showed decreased T cell infiltration (CD2, CD8, CD4) compared with controls and decreased arterial endothelial inflammation. No changes occurred in circulating T cells, aside from variable coating with mIgG. In six of eight other recipients R6.5 administration (0.5 to 2 mg/kg/day for 10 days) reversed preexisting rejection that resulted from taper of Cyclosporine to subtherapeutic levels. Responding grafts showed decreased edema and hemorrhage but no consistent change in the infiltrate. At 1 h after the first dose, mouse IgG deposited primarily on the graft vascular endothelium without any change in the inflammatory infiltrate. Mouse IgG also deposited on the endothelium of normal organs without eliciting an inflammatory response and was cleared from the endothelium within 4 days. Inasmuch as the principal site of binding was the vascular endothelium, we hypothesize that the antibody blocks adhesion to graft ICAM-1 molecules on the vessels. Anti-ICAM-1 also binds to recipient cells and may interfere with Ag presentation and/or T cell interactions. Whatever the mechanism(s), these studies indicate that an anti-ICAM-1 antibody inhibits T cell mediated injury in vivo, and that ICAM-1 is a critical molecule in the pathogenesis of allograft rejection.     

B cell-associated LFA-1 and T cell-associated ICAM-1 transiently cluster in the area of contact between interacting cells.

TNP-specific B cells interact with carrier-specific T hybridoma cells in an antigen-specific, MHC-restricted manner. The formation of T cell/B cell conjugates is time and temperature dependent and results in the formation of a broad area of close contact between the interacting cells. In order to determine which surface molecules on the two cells cluster at the interaction site. T cell/B cell conjugates were formalin-fixed at different times following conjugation and were stained with antibodies directed against cell surface molecules. Results of these studies indicate that the alpha- and beta-subunits of LFA-1 on B cells transiently cluster in the area of cell contact. Maximum clustering of LFA-1 occurs at 45 min, after which time LFA-1 redistributes on the surface of the B cells. Several other B cell-associated molecules (MHC Class II, ICAM-1, Ig, B220, J11D, or CD23) do not cluster at the interaction site at any time point. T cell-associated LFA-1 does not cluster with any specific pattern, but ICAM-1 does. Maximum clustering of ICAM-1 occurs 60 to 90 min after intercellular contact. After this time, ICAM-1 redistributes on the surface of the T cells. Although both the alpha- and beta-subunits of LFA-1 cluster at the interaction site on B cells, antibodies recognizing these subunits differ in their ability to affect conjugation. One antibody recognizing the alpha chain of LFA-1 (M17/4.2) inhibits T-cell/B cell conjugation, whereas another antibody that also recognizes the alpha chain-(G-48) enhances conjugation. In contrast, an antibody that recognizes LFA-1 beta (M18/2.a.8) has no effect. An antibody that recognizes ICAM-1 (YN/1.7), the ligand for LFA-1, inhibits conjugation. These data show that, during T cell/B cell interaction. LFA-1 on B cells and ICAM-1 on T cells transiently cluster with similar, albeit not identical, kinetics to the site of cell-cell contact.     

Human endothelial-cell specific molecule-1 binds directly to the integrin CD11a/CD18 (LFA-1) and blocks binding to intercellular adhesion molecule-1.

ICAMs are ligands for LFA-1, a major integrin of mononuclear cells involved in the immune and inflammatory processes. We previously showed that endothelial cell specific molecule-1 (ESM-1) is a proteoglycan secreted by endothelial cells under the control of inflammatory cytokines. Here, we demonstrate that ESM-1 binds directly to LFA-1 onto the cell surface of human blood lymphocytes, monocytes, and Jurkat cells. The binding of ESM-1 was equally dependent on Ca(2+), Mg(2+), or Mn(2+) divalent ions, which are specific, saturable, and sensitive to temperature. An anti-CD11a mAb or PMA induced a transient increase in binding, peaking 5 min after activation. Direct binding of ESM-1 to LFA-1 integrin was demonstrated by specific coimmunoprecipitation by CD11a and CD18 mAbs. A cell-free system using a Biacore biosensor confirmed that ESM-1 and LFA-1 dynamically interacted in real time with high affinity (K(d) = 18.7 nM). ESM-1 consistently inhibited the specific binding of soluble ICAM-1 to Jurkat cells in a dose-dependent manner. These results suggest that ESM-1 and ICAM-1 interact with LFA-1 on binding sites very close to but distinct from the I domain of CD11a. Through this mechanism, ESM-1 could be implicated in the regulation of the LFA-1/ICAM-1 pathway and may therefore influence both the recruitment of circulating lymphocytes to inflammatory sites and LFA-1-dependent leukocyte adhesion and activation.     

PLGA nanoparticle--peptide conjugate effectively targets intercellular cell-adhesion molecule-1

Targeted delivery of therapeutics possesses the potential to localize therapeutic agents to a specific tissue as a mechanism to enhance treatment efficacy and abrogate side effects. Antibodies have been used clinically as therapeutic agents and are currently being explored for targeting drug-loaded nanoparticles. Peptides such as RGD peptides are also being developed as an inexpensive and stable alternative to antibodies. In this study, cyclo(1,12)PenITDGEATDSGC (cLABL) peptide was used to target nanoparticles to human umbilical cord vascular endothelial cell (HUVEC) monolayers that have upregulated intercellular cell-adhesion molecule-1 (ICAM-1) expression. The cLABL peptide has been previously demonstrated to possess high avidity for ICAM-1 receptors on the cell surface. Poly( dl-lactic-coglycolic acid) nanoparticles conjugated with polyethylene glycol and cLABL demonstrated rapid binding to HUVEC with upregulated ICAM-1, which was induced by treating cells with the proinflammatory cytokine, interferon-gamma. Binding of the nanoparticles could be efficiently blocked by preincubating cells with free peptide suggesting that the binding of the nanoparticles is specifically mediated by surface peptide binding to ICAM-1 on HUVEC. The targeted nanoparticles were rapidly endocytosed and trafficked to lysosomes to a greater extent than the untargeted PLGA-PEG nanoparticles. Verification of peptide-mediated nanoparticle targeting to ICAM-1 may ultimately lead to targeting therapeutic agents to inflammatory sites expressing upregulated ICAM-1.     

Accessible hyaluronan receptors identical to ICAM-1 in mouse mast-cell tumours

Immunohistochemical studies of the hyaluronan (HA)-receptor (R), originally found on liver endothelial cells (LEC) and related to the intercellular adhesion molecule 1 (ICAM-1), showed that polyclonal antibodies against HARLEC (HA receptor on LEC) also stain structures in mouse mastocytomas, mainly vessels. To test if intravenously administered HA might target the tumour receptorsin vivo, mice carrying an inoculated mastocytoma in one hind leg muscle were injected in the tail vein with¹²⁵I-tyrosine (T)-labelled HA and killed 75 min after injection when organs and tissues were checked for radioactivity. When doses exceeding the binding capacity of the liver were injected, a significant increase in radioactivity (up to five-fold) within the tumour tissue was found. The weight adjusted difference between control and tumour tissue was greater for smaller tumours, probably due to necrosis in the larger. HA-staining of tumours from animals receiving¹²⁵I-T-HA, showed HA in areas that also stained weakly for ICAM-1 using monoclonal antibodies. ICAM-1 staining was dramatically increased after hyaluronidase treatment of the sections, indicating that the HA is bound to these receptors and thereby blocks antibody recognition.Abbreviations ICAM-1 intercellular adhesion molecule 1 - HA hyaluronan - HARLEC hyaluronan receptor on liver endothelial cells - MW molecular weight     

Binding of human rhinovirus and T cells to intercellular adhesion molecule-1 on human fibroblasts. Discordance between effects of IL-1 and IFN-gamma.

Intercellular adhesion molecule-1 (ICAM-1) is found on the surface of many hemopoietic and non-hemopoietic cells and can function as an adhesive ligand for the integrin, leukocyte function associated molecule-1 (LFA-1, CD11a/CD18). ICAM-1/LFA-1 interaction is thought to be of importance in many immune mediated cell-cell adhesion reactions. Recently, the major human rhinovirus (HRV) receptor has been identified as ICAM-1. HRV has been shown to bind specifically to ICAM-1 on transfected COS cells and to purified ICAM-1, which has been adsorbed to plastic microtiter wells. We have compared the ability of ICAM-1 expressed on the surface of human fibroblasts (FB) to function as a receptor for HRV as well as a receptor for LFA-1-bearing human T lymphocytes. We show that FB stimulation by the cytokines IFN-gamma or IL-1, both known inducers of ICAM-1 synthesis and expression in FB, induced an increase in HRV binding to treated cells, which could be inhibited by antibody to ICAM-1. In contrast, only IFN-gamma and not IL-1 treatment of FB resulted in an increased adhesion of T lymphocytes. Binding of HRV to IFN-gamma-treated FB inhibited the subsequent adhesion of T cells. We also show that prior stimulation of FB with IL-1 enhanced the adhesion of HRV to IFN-gamma-stimulated cells, although IL-1 pretreatment was inhibitory for T cell adhesion. As these two cytokines both up-regulate ICAM-1 on the surface of human FB, the contrasting effects of IFN-gamma and IL-1 on human FB ICAM-1 adhesion to HRV and to LFA-1 suggest that qualitative as well as quantitative alterations of the ICAM-1 molecule may contribute to its specificity of ligand recognition.     

Mast cells are potent regulators of endothelial cell adhesion molecule ICAM-1 and VCAM-1 expression

To investigate the possible role of mast cells (MC) in regulating leukocyte adhesion to vascular endothelial cells (EC), microvascular and macrovascular EC were exposed to activated MC or MC conditioned medium (MCCM). Expression of intercellular and vascular adhesion molecules (ICAM-1 and VCAM-1) on EC was monitored. Incubation of human dermal microvascular endothelial cells (HDMEC) and human umbilical vein endothelial cells (HUVEC) with activated MC or MCCM markedly increased ICAM-1 and VCAM-1 surface expression, noted as éarly as 4 hr. Maximal levels were observed at 16 hr followed by a general decline over 48 hr. A dose-dependent response was noted using incremental dilutions of MCCM or by varying the number of MC in coculture with EC. At a ratio as low as 1:1,000 of MC:EC, increased ICAM-1 was observed. The ICAM-1 upregulation by MCCM was >90% neutralized by antibody to tumor necrosis factor alpha (TNF-α), suggesting that MC release of this cytokine contributes significantly to inducing EC adhesiveness. VCAM-1 expression enhanced by MCCM was partly neutralized (70%) by antibody to TNF-α; thus other substances released by MC may contribute to VCAM-1 expression. Northern blot analysis demonstrated MCCM upregulated ICAM-1 and VCAM-1 mRNA in both HDMEC and HUVEC. To evaluate the function of MCCM-enhanced EC adhesion molecules, T cells isolated from normal human donors were used in a cell adhesion assay. T-cell binding to EC was increased significantly after exposure of EC to MCCM, and inhibited by antibodies to ICAM-1 or VCAM-1. Intradermal injection of allergen in human atopic volunteers known to develop late-phase allergic reactions led to marked expression of both ICAM-1 and VCAM-1 at 6 hr, as demonstrated by immunohistochemistry. These studies indicate that MC play a critical role in regulating the expression of EC adhesion molecules, ICAM-1 and VCAM-1, and thus augment inflammatory responses by upregulating leukocyte binding. © 1995 Wiley-Liss Inc.     

VCAM-1-targeted magnetic resonance imaging reveals subclinical disease in a mouse model of multiple sclerosis

Diagnosis of multiple sclerosis (MS) currently requires lesion identification by gadolinium (Gd)-enhanced or T(2)-weighted magnetic resonance imaging (MRI). However, these methods only identify late-stage pathology associated with blood-brain barrier breakdown. There is a growing belief that more widespread, but currently undetectable, pathology is present in the MS brain. We have previously demonstrated that an anti-VCAM-1 antibody conjugated to microparticles of iron oxide (VCAM-MPIO) enables in vivo detection of VCAM-1 by MRI. Here, in an experimental autoimmune encephalomyelitis (EAE) mouse model of MS, we have shown that presymptomatic lesions can be quantified using VCAM-MPIO when they are undetectable by Gd-enhancing MRI. Moreover, in symptomatic animals VCAM-MPIO binding was present in all regions showing Gd-DTPA enhancement and also in areas of no Gd-DTPA enhancement, which were confirmed histologically to be regions of leukocyte infiltration. VCAM-MPIO binding correlated significantly with increasing disability. Negligible MPIO-induced contrast was found in either EAE animals injected with an equivalent nontargeted contrast agent (IgG-MPIO) or in control animals injected with the VCAM-MPIO. These findings describe a highly sensitive molecular imaging tool that may enable detection of currently invisible pathology in MS, thus accelerating diagnosis, guiding treatment, and enabling quantitative disease assessment.     

On the species specificity of the interaction of LFA-1 with intercellular adhesion molecules

Species restrictions in immune cell interactions have been demonstrated both in Ag-specific responses of T lymphocytes and the phenomenon of natural attachment. To determine the possible contribution of adhesion receptors to these restrictions, we have studied binding between the murine and human homologues of LFA-1 (CD11a/CD18) and ICAM employing purified human LFA-1 and ICAM-1 (CD54) bound to solid substrates. Murine cell lines bind to purified human LFA-1 through ICAM-1 and at least one other counter-receptor. This provides evidence for multiple counter-receptors for LFA-1 in the mouse as well as in the human. In contrast to binding of murine ICAM-1 to human LFA-1, murine LFA-1 does not bind to human ICAM-1. The species specificity maps to the LFA-1 alpha subunit, because mouse x human hybrid cells expressing the human alpha subunit associated with a mouse beta subunit bind to human ICAM-1, whereas those with a human beta subunit associated with a murine alpha subunit do not. Increased adhesiveness for ICAM-1 stimulated by phorbol esters could be demonstrated for hybrid LFA-1 molecules with human alpha and murine beta subunits.     

Identification of different binding sites in the dendritic cell-specific receptor DC-SIGN for intercellular adhesion molecule 3 and HIV-1.

The novel dendritic cell (DC)-specific human immunodeficiency virus type 1 (HIV-1) receptor DC-SIGN plays a key role in the dissemination of HIV-1 by DC. DC-SIGN is thought to capture HIV-1 at mucosal sites of entry, facilitating transport to lymphoid tissues, where DC-SIGN efficiently transmits HIV-1 to T cells. DC-SIGN is also important in the initiation of immune responses by regulating DC-T cell interactions through intercellular adhesion molecule 3 (ICAM-3). We have characterized the mechanism of ligand binding by DC-SIGN and identified the crucial amino acids involved in this process. Strikingly, the HIV-1 gp120 binding site in DC-SIGN is different from that of ICAM-3, consistent with the observation that glycosylation of gp120, in contrast to ICAM-3, is not crucial to the interaction with DC-SIGN. A specific mutation in DC-SIGN abrogated ICAM-3 binding, whereas the HIV-1 gp120 interaction was unaffected. This DC-SIGN mutant captured HIV-1 and infected T cells in trans as efficiently as wild-type DC-SIGN, demonstrating that ICAM-3 binding is not necessary for HIV-1 transmission. This study provides a basis for the design of drugs that inhibit or alter interactions of DC-SIGN with gp120 but not with ICAM-3 or vice versa and that have a therapeutic value in immunological diseases and/or HIV-1 infections.     

Involvement of the "I" domain of LFA-1 in selective binding to ligands ICAM-1 and ICAM-3

To analyze the binding requirements of LFA-1 for its two most homologous ligands, ICAM-1 and ICAM-3, we compared the effects of various LFA-1 activation regimes and a panel of anti-LFA-1 mAbs in T cell binding assays to ICAM-1 or ICAM-3 coated on plastic. These studies demonstrated that T cell binding to ICAM-3 was inducible both from the exterior of the cell by Mn2+ and from the interior by an agonist of the "inside-out" signaling pathway. T cells bound both ICAM ligands with comparable avidity. A screen of 29 anti-LFA-1 mAbs led to the identification of two mAbs specific for the alpha subunit of LFA-1 which selectively blocked adhesion of T cells to ICAM-3 but not ICAM-1. These two mAbs, YTH81.5 and 122.2A5, exhibited identical blocking properties in a more defined adhesion assay using LFA-1 transfected COS cells binding to immobilized ligand. Blocking was not due to a steric interference between anti-LFA-1 mAbs and N-linked carbohydrate residues present on ICAM-3 but not ICAM-1. The epitopes of mAbs YTH81.5 and 122.2A5 were shown to map to the I domain of the LFA-1 alpha subunit. A third I domain mAb, MEM-83, has been previously reported to uniquely activate LFA-1 to bind ICAM-1 (Landis, R. C., R. I. Bennett, and N. Hogg. 1993. J. Cell Biol. 120:1519-1527). We now show that mAb MEM-83 is not able to stimulate binding of T cells to ICAM-3 over a wide concentration range. Failure to induce ICAM-3 binding by mAb MEM-83 was not due to a blockade of the ICAM-3 binding site on LFA-1. This study has demonstrated that two sets of functionally distinct mAbs recognizing epitopes in the I domain of LFA-1 are able to exert differential effects on the binding of LFA-1 to its ligands ICAM-1, and ICAM-3. These results suggest for the first time that LFA-1 is capable of binding these two highly homologous ligands in a selective manner and that the I domain plays a role in this process.     

ICAM-1-Based Rabies Virus Vaccine Shows Increased Infection and Activation of Primary Murine B Cells In Vitro and Enhanced Antibody Titers In-Vivo

We have previously shown that live-attenuated rabies virus (RABV)-based vaccines infect and directly activate murine and human primary B cells in-vitro, which we propose can be exploited to help develop a single-dose RABV-based vaccine. Here we report on a novel approach to utilize the binding of Intracellular Adhesion Molecule-1 (ICAM-1) to its binding partner, Lymphocyte Function-associated Antigen-1 (LFA-1), on B cells to enhance B cell activation and RABV-specific antibody responses. We used a reverse genetics approach to clone, recover, and characterize a live-attenuated recombinant RABV-based vaccine expressing the murine Icam1 gene (rRABV-mICAM-1). We show that the murine ICAM-1 gene product is incorporated into virus particles, potentially exposing ICAM-1 to extracellular binding partners. While rRABV-mICAM-1 showed 10-100-fold decrease in viral titers on baby hamster kidney cells compared to the parental virus (rRABV), rRABV-mICAM-1 infected and activated primary murine B cells in-vitro more efficiently than rRABV, as indicated by significant upregulation of CD69, CD40, and MHCII on the surface of infected B cells. ICAM-1 expression on the virus surface was responsible for enhanced B cell infection since pre-treating rRABV-mICAM-1 with a neutralizing anti-ICAM-1 antibody reduced B cell infection to levels observed with rRABV alone. Furthermore, 100-fold less rRABV-mICAM-1 was needed to induce antibody titers in immunized mice equivalent to antibody titers observed in rRABV-immunized mice. Of note, only 10³ focus forming units (ffu)/mouse of rRABV-mICAM-1 was needed to induce significant anti-RABV antibody titers as early as five days post-immunization. As both speed and potency of antibody responses are important in controlling human RABV infection in a post-exposure setting, these data show that expression of Icam1 from the RABV genome, which is then incorporated into the virus particle, is a promising strategy for the development of a single-dose RABV vaccine that requires only a minimum of virus.     

Intrinsic ICAM-1/LFA-1 activation mediates altered responsiveness of atopic asthmatic airway smooth muscle

Cell adhesion molecules (CAMs) have been importantly implicated in the pathobiology of the airway responses in allergic asthma, including inflammatory cell recruitment into the lungs and altered bronchial responsiveness. To elucidate the mechanism of CAM-related mediation of altered airway responsiveness in the atopic asthmatic state, the expressions and actions of intercellular adhesion molecule-1 (ICAM-1) and its counterreceptor ligand lymphocyte function-associated antigen-1 (LFA-1; i.e., CD11a/CD18) were examined in isolated rabbit airway smooth muscle (ASM) tissues and cultured human ASM cells passively sensitized with sera from atopic asthmatic patients or nonatopic nonasthmatic (control) subjects. Relative to control tissues, the atopic asthmatic sensitized ASM exhibited significantly enhanced maximal contractility to acetylcholine and attenuated relaxation responses to isoproterenol. These proasthmatic changes in agonist responsiveness were ablated by pretreating the atopic sensitized tissues with a monoclonal blocking antibody (MAb) to either ICAM-1 or CD11a, whereas a MAb directed against the related beta(2)-integrin Mac-1 had no effect. Moreover, relative to control tissues, atopic asthmatic sensitized ASM cells displayed an autologously upregulated mRNA and cell surface expression of ICAM-1, whereas constitutive expression of CD11a was unaltered. Extended studies further demonstrated that 1) the enhanced expression and release of soluble ICAM-1 by atopic sensitized ASM cells was prevented when cells were pretreated with an interleukin (IL)-5-receptor-alpha blocking antibody and 2) administration of exogenous IL-5 to naive (nonsensitized) ASM cells induced a pronounced soluble ICAM-1 release from the cells. Collectively, these observations provide new evidence demonstrating that activation of the CAM counterreceptor ligands ICAM-1 and LFA-1, both of which are endogenously expressed in ASM cells, elicits autologously upregulated IL-5 release and associated changes in ICAM-1 expression and agonist responsiveness in atopic asthmatic sensitized ASM.     

Intercellular adhesion molecule-1 (ICAM-1) is involved in the cytolytic T lymphocyte interaction with a human synovial cell line

The cell surface molecules involved in the human cytolytic T lymphocyte (CTL)-synovial cell interaction may play an important role in T cell interactions with connective tissue mesenchymal cells. To examine the molecular basis for the CTL-synovial cell interaction, we immortalized synovial cell explants to establish the cell line SYN.SPP. The SYN.SPP cell line was compared to the established B lymphoblastoid cell line JY. Cell surface immunofluorescence demonstrated significantly different levels of the immunologically relevant cell surface molecules ICAM-1 and LFA-3. Both cell lines were used to stimulate CTL precursors. After several months in culture, CTL lines stimulated by the SYN.SPP and JY cell lines demonstrated HLA class I-directed cytolytic activity. The cell surface molecules utilized by the anti-SYN.SPP and anti-JY CTL lines were identified by monoclonal antibody (MAb) inhibition. MAb recognizing the CTL cell surface molecules CD3, CD8 and LFA-1 (CD11a) significantly inhibited CTL-mediated lysis of both target cells. An interesting observation was that the anti-SYN.SPP CTL line appeared to utilize the ICAM-1 and not the LFA-3 target cell molecule. In contrast, the anti-JY CTL line utilized the LFA-3 and not the ICAM-1 membrane molecule. These results indicate that CTL interactions with connective tissue mesenchymal cells may be regulated by a unique pattern of antigen nonspecific cell-cell interaction molecules.     

t-PA对EAE 病理性淋巴细胞与血脑屏障粘附的影响

目的：研究组织型纤溶酶原激活剂（t-PA）对实验性自身免疫性脑脊髓炎（EAE）小鼠病理性淋巴细胞与血脑屏障粘附的影 响。方法：用MOG35-55 肽段免疫C57BL/6小鼠建立EAE 动物模型,于发病高峰期取淋巴细胞用MOG35-55 肽段进行刺激得到 抗原特异性T淋巴细胞。通过尾静脉给予t-PA 的方法对EAE 小鼠进行干预,临床评分评价小鼠的发病情况。体外培养小鼠血脑 屏障内皮细胞系bEnd.3,应用不同浓度的t-PA 进行处理。用荧光标记MOG35-55 特异性T 细胞进行细胞粘附实验,用Transwell 小室建立体外血脑屏障模型进行细胞迁移实验。用免疫荧光化学方法检测ICAM-1 的表达情况。结果：t-PA处理可以使血脑屏障 内皮细胞与T 淋巴细胞粘附和迁移作用增强。在体外细胞培养模型中检测到t-PA诱导ICAM-1 表达升高。经过t-PA 处理的小 鼠,其血管内皮表面ICAM-1 的表达也有所上升。经t-PA 处理的EAE 小鼠发病高峰提前,症状加重。结论：t-PA 处理可以使EAE 病理性淋巴细胞与血脑屏障内皮的粘附性增加,浸润能力增强;t-PA 所引起的粘附性增加可能与bEnd.3 表面ICAM-1表达升高 有关。     

Molecular cloning of murine intercellular adhesion molecule (ICAM-1).

We have previously reported a murine lymphocyte surface antigen MALA-2 of approximately 95,000 Mr which is expressed mainly on activated lymphocytes. The rat monoclonal antibody YN1/1 that detects this antigen profoundly inhibits mixed lymphocyte response. We have now purified MALA-2 and determined its partial amino acid sequence. By using non-redundant synthetic oligonucleotides as probes, based on the amino acid sequence, we have isolated two full length cDNA clones encoding MALA-2. The two clones are identical except for the 5' end sequence. Expression of MALA-2 on transfected COS cells is only achieved with one of the two cDNA clones. The nucleotide sequence as well as the deduced amino acid sequence of MALA-2 display striking homology with those of the recently reported human intercellular adhesion molecule ICAM-1. All the unique features of the human ICAM-1, including its homology with the neural adhesion molecule NCAM, its internal repeat structure and the immunoglobulin-like structure, are found in MALA-2. Furthermore, purified MALA-2 crosslinked to a solid support binds Con A blasts that express LFA-1, the putative receptor for ICAM-1, and the binding can be blocked by YN1/1 antibody or antimurine LFA-1 antibody indicating a direct interaction of these molecules in cell adhesion. Therefore, we consider MALA-2 to be the murine homolog of human ICAM-1. Since ICAM-1 is known to be of primary importance in immune responses and inflammatory reactions, having a monoclonal antibody and a mouse model will provide the opportunity to study the functional role of ICAM-1 in vivo.     

Five tumor necrosis factor-inducible cell adhesion mechanisms on the surface of mouse endothelioma cells mediate the binding of leukocytes

We have distinguished five TNF-alpha-inducible cell adhesion mechanisms on microvasculature-derived endothelioma cells of the mouse which mediate the binding of different types of leukocytes. Three of these mechanisms could be identified as the mouse homologs of ICAM-1, VCAM-1, and E-selectin, of which the latter was defined by the novel mAb 21KC10. The fourth TNF-alpha-inducible cell adhesion mechanism was blocked by antibodies specific for mouse P-selectin. We have recently shown that TNF-alpha stimulates the synthesis of P-selectin in mouse endothelioma cells (A. Weller, S. Isenmann, D. Vestweber. 1992. J. Biol. Chem. 267:15176-15183). Here we show that this stimulation leads to maximal cell surface expression levels within 4 h after stimulation while the same endothelioma cells are also able to upregulate P- selectin at the cell surface within minutes after stimulation with PMA. Both effects are additive. The fifth TNF-induced cell adhesion mechanism is defined by mediating the binding to the mouse monocyte/macrophage cell line J774. This adhesion mechanism is not inhibited by antibodies against any of the other four CAMs; it functions well at 7 degrees C (in contrast to ICAM-1 and VCAM-1) and it is as active after 16 h of TNF induction as after 4 h (in contrast to E- and P-selectin). Furthermore, this new adhesion mechanism only functions on two of three endothelioma cell lines and is undetectable on the third, although ICAM-1, VCAM-1, E-selectin, and P-selectin could be demonstrated to function well on this cell line. Thus, in addition to the three known TNF-inducible CAMs, ICAM-1, VCAM-1, and E-selectin, also P-selectin and a fifth, as yet molecularly undefined cell adhesion mechanism, are TNF inducible at the cell surface of mouse endothelioma cells.