Adhesion-mediating molecules of human monocytes

Adhesion of monocytes to each other and to T cells and substrates is increased by phorbol esters. In the presence of these compounds monocyte aggregation was almost completely inhibited (greater than 90%) by monoclonal antibody 60.3. This antibody recognizes GP90 (CD18), a leukocyte surface glycoprotein which is separately and noncovalently associated to either GP160 (CD11a), GP155 (CD11b), or GP130 (CD11c). Anti-LFA-1 antibody (CD11a) was only partially inhibitory (35%) while antibodies 60.1 (CD11b) and anti-Leu-M5 (CD11c) had a minimal inhibitory effect (10%). Antibody LB-2 recognizing a single glycoprotein distinct from the GP90-GP160 complex and expressed on activated B and T cells, monocytes, and vascular endothelial cells was partially inhibitory (22%). Monoclonal antibodies anti-C3bR (CD35), T29/33 (CD45, leukocyte common antigen 200). TA-1 (CD11a), OKM1 (CD11b), F10-44-2 (brain-leukocyte antigen), OKM5 (monocyte-endothelial cell antigen) and to class I or class II molecules exerted no inhibition on the monocyte aggregation. Fab fragments of antibody 60.3 efficiently inhibited not only monocyte aggregation in the absence or presence of phorbol esters but also adhesion of these cells to autologous or allogeneic T lymphocytes and, to a lesser extent, to plastic surfaces. It is thus concluded that GP90, either alone or associated to the larger glycoproteins, and LB-2 antigen mediate monocyte adhesion.     

Arsenic Exposure Increases Monocyte Adhesion to the Vascular Endothelium,a Pro-Atherogenic Mechanism

Epidemiological studies have shown that arsenic exposure increases atherosclerosis, but the mechanisms underlying this relationship are unknown. Monocytes, macrophages and platelets play an important role in the initiation of atherosclerosis. Circulating monocytes and macrophages bind to the activated vascular endothelium and migrate into the sub-endothelium, where they become lipid-laden foam cells. This process can be facilitated by platelets, which favour monocyte recruitment to the lesion. Thus, we assessed the effects of low-to-moderate arsenic exposure on monocyte adhesion to endothelial cells, platelet activation and platelet-monocyte interactions. We observed that arsenic induces human monocyte adhesion to endothelial cells in vitro. These findings were confirmed ex vivo using a murine organ culture system at concentrations as low as 10 ppb. We found that both cell types need to be exposed to arsenic to maximize monocyte adhesion to the endothelium. This adhesion process is specific to monocyte/endothelium interactions. Hence, no effect of arsenic on platelet activation or platelet/leukocyte interaction was observed. We found that arsenic increases adhesion of mononuclear cells via increased CD29 binding to VCAM-1, an adhesion molecule found on activated endothelial cells. Similar results were observed in vivo, where arsenic-exposed mice exhibit increased VCAM-1 expression on endothelial cells and increased CD29 on circulating monocytes. Interestingly, expression of adhesion molecules and increased binding can be inhibited by antioxidants in vitro and in vivo. Together, these data suggest that arsenic might enhance atherosclerosis by increasing monocyte adhesion to endothelial cells, a process that is inhibited by antioxidants.     

Occupancy of CD11b/CD18 (Mac-1) divalent ion binding site(s) induces leukocyte adhesion

The group of leukocyte integrins CD11a-c/CD18 coordinate disparate adhesion reactions in the immune system through a regulated process of ligand recognition. The participation of the receptor divalent ion binding site(s) in this mechanism of ligand binding has been investigated. As compared with other divalent cations, Mn2+ ions have the unique property to dramatically stimulate the adhesive functions of the leukocyte integrin CD11b/CD18 (Mac-1), expressed on myelo-monocytic cells. This is reflected in a three- to fivefold increased early monocyte adhesion (less than 20 min) to resting, unperturbed endothelial cells, and increased association of CD11b/CD18 with its soluble ligands fibrinogen and factor X. CD11b/CD18 ligand recognition in the presence of Mn2+ ions is specific, time and concentration dependent, and inhibited by anti-CD11b mAb. At variance with Ca(2+)-containing reactions where CD11b/CD18 functions as an inducible receptor activated by adenine nucleotides or chemoattractants, Mn2+ ions induce per se a constitutive maximal ligand binding capacity of CD11b/CD18, that is not further modulated by cell stimulation. Rather than quantitative changes in surface density, Mn2+ ions increase the affinity of CD11b/CD18 for its complementary ligands up to 10-fold, as judged by Scatchard plot analysis of receptor:ligand interaction under these conditions. Furthermore, monocyte exposure to Mn2+ ions induces the expression of activation-dependent neo-antigenic epitopes on CD11b/CD18, selectively recognized by mAb 7E3. These data suggest that in addition to cell-activating stimuli, favorable engagement of divalent ion binding site(s) can provide an alternative pathway to rapidly regulate the receptor affinity of leukocyte integrins.     

Constitutive and stimulus-induced phosphorylation of CD11/CD18 leukocyte adhesion molecules

The leukocyte CD11/CD18 adhesion molecules (beta 2 integrins) are a family of three heterodimeric glycoproteins each with a distinct alpha subunit (CD11a, b, or c) and a common beta subunit (CD18). CD11/CD18 mediate crucial leukocyte adhesion functions such as chemotaxis, phagocytosis, adhesion to endothelium, aggregation, and cell-mediated cytotoxicity. The enhanced cell adhesion observed upon activation of leukocytes is associated with increased surface membrane expression of CD11/CD18, as well as a qualitative upregulation of CD11/CD18 functions. To elucidate the nature of the qualitative modifications that occur, we examined the phosphorylation status of these molecules in resting human leukocytes and upon activation with PMA or with the chemotactic peptide F-met-leu-phe (FMLP). In unstimulated cells, all three CD11 subunits were found to be constitutively phosphorylated. In contrast, phosphorylation of the common CD18 subunit was minimal. PMA induced rapid and sustained phosphorylation of CD18 that occurred at high stoichiometry, but had only minimal effects on phosphorylation of the associated CD11 subunits. FMLP also induced rapid phosphorylation of CD18, but the effect was of short duration. FMLP-induced phosphorylation of CD18 was not related to its Ca++-mobilizing effect, as CD18 phosphorylation was not observed upon treatment of leukocytes with the Ca++ ionophore, ionomycin. Phosphoamino acid analysis of CD11/CD18 in PMA- or FMLP-treated monocytes revealed a predominance of phosphoserine residues in all CD11/CD18 subunits. A small component of phosphothreonine was present in CD11c and CD18 and a minor component of phosphotyrosine was also detected in CD18 upon leukocyte activation may regulate the adhesion functions mediated by the CD11/CD18 family of molecules.     

Human monocyte adherence to cultured vascular endothelium: monoclonal antibody-defined mechanisms

We have evaluated the binding of human peripheral blood monocytes to cultured vascular endothelium as an in vitro model of monocyte interaction with the vessel wall. Monocytes were purified (91% +/- 4 SE esterase positive) by elutriation to avoid contact with surfaces before assay. Adherence of 51Cr-labeled monocytes after 45 min (36% +/- 11 SE) was significantly higher than that observed with autologous radiolabeled neutrophils (9% +/- 5 SE) and was greater on monolayers of human umbilical vein endothelium than on bovine aortic endothelium. Peripheral blood mononuclear cells treated with monoclonal antibody (MoAb) 60.3, a reagent that binds leukocyte membrane complex CDw18, implicated in multiple adherence-dependent functions, failed to adhere and flatten on artificial surfaces. Mononuclear cells treated with MoAb 60.3 simulated cells from a patient with recurrent infections whose phagocytes failed to react with MoAb 60.3 and failed to emigrate to extravascular sites in vivo. Incubation of monocytes with MoAb 60.3 inhibited (by 32 to 61%) monocyte adherence to endothelium in a dose-dependent manner for periods up to 24 hr, but had negligible effects on basal (unstimulated) neutrophil adherence. Basal monocyte adherence in the presence of MoAb 60.3 remained significantly greater than basal neutrophil adherence. Augmentation of phagocyte adherence to endothelial monolayers by autologous plasma or phorbol ester (PMA) was abrogated by incubation with MoAb 60.3. Studies with immunofluorescence flow cytometry indicated that PMA stimulation of monocytes resulted in a specific 40% increase in monocyte surface expression of the epitope recognized by MoAb 60.3. These in vitro findings, in conjunction with observations from two patients, support the hypothesis that monocyte adherence to endothelium and emigration to tissues is mediated by mechanisms both dependent upon and independent of the CDw18 complex and the epitope recognized by MoAb 60.3.     

TLR4-Mediated Expression of Mac-1 in Monocytes Plays a Pivotal Role in Monocyte Adhesion to Vascular Endothelium

Toll-like receptor 4 (TLR4) is known to mediate monocyte adhesion to endothelial cells, however, its role on the expression of monocyte adhesion molecules is unclear. In the present study, we investigated the role of TLR4 on the expression of monocyte adhesion molecules, and determined the functional role of TLR4-induced adhesion molecules on monocyte adhesion to endothelial cells. When THP-1 monocytes were stimulated with Kdo2-Lipid A (KLA), a specific TLR4 agonist, Mac-1 expression was markedly increased in association with an increased adhesion of monocytes to endothelial cells. These were attenuated by anti-Mac-1 antibody, suggesting a functional role of TLR4-induced Mac-1 on monocyte adhesion to endothelial cells. In monocytes treated with MK886, a 5-lipoxygenase (LO) inhibitor, both Mac-1 expression and monocyte adhesion to endothelial cells induced by KLA were markedly attenuated. Moreover, KLA increased the expression of mRNA and protein of 5-LO, suggesting a pivotal role of 5-LO on these processes. In in vivo studies, KLA increased monocyte adhesion to aortic endothelium of wild-type (WT) mice, which was attenuated in WT mice treated with anti-Mac-1 antibody as well as in TLR4-deficient mice. Taken together, TLR4-mediated expression of Mac-1 in monocytes plays a pivotal role on monocyte adhesion to vascular endothelium, leading to increased foam cell formation in the development of atherosclerosis.     

A unique recognition site mediates the interaction of fibrinogen with the leukocyte integrin Mac-1 (CD11b/CD18)

Mac-1 (CD11b/CD18), a leukocyte-restricted integrin receptor, mediates neutrophil/monocyte adhesion to vascular endothelium and phagocytosis of complement-opsonized particles. Recent studies have shown that Mac-1 also functions as a receptor for fibrinogen in a reaction linked to fibrin deposition on the monocyte surface. In this study, we have used extended proteolytic digestion of fibrinogen to identify the region of this molecule that interacts with Mac-1. We found that an Mr approximately 30,000 plasmic fragment D of fibrinogen (D30) produced dose-dependent inhibition (IC50 = 1.6 microM) of the interaction of intact 125I-fibrinogen with stimulated neutrophils and monocytes. 125I-D30 bound saturably to these cells with specific association of 136,200 +/- 15,000 molecules/cell in a reaction inhibited by OKM1 and M1/70, monoclonal antibodies specific for the alpha subunit of Mac-1. Direct microsequence analysis and an epitope-mapped monoclonal antibody showed that D30 lacks the COOH-terminal dodecapeptide of the gamma chain as well as the Arg-Gly-Asp sequences in the A alpha chain. We conclude that fibrinogen interacts with the leukocyte integrin Mac-1 through a novel recognition site that is not shared with other known integrins that function as fibrinogen receptors.     

The role of sex steroids on cellular events involved in vascular disease

In this work we checked the hypothesis whether estrone, progesterone, and testosterone are able to modulate the interactions between platelets, monocytes, and endothelial cells either under basal or inflammatory conditions. Using adhesion assays we demonstrated that pretreatment of endothelial cells with estrone, progesterone, or testosterone prevented monocytes and platelets adhesion induced by the proinflammatory agent bacterial lipopolysaccharide. The hormones reduced the expression of mRNA of ICAM-1, VCAM-1, and P-selectin, endothelial surface proteins that mediate monocytes and platelets adhesion respectively. Integrins are the main leukocyte proteins that allow firm adhesion. Using flow cytometry we showed that estrone treatment of monocytes reduced CD11b and CD11c expression, either under basal or injury (lipopolysaccharide) conditions. The three steroids inhibited platelet aggregation in a nitric oxide dependent manner. Platelet function was not affected by the steroid treatment. The molecular mechanisms of action exerted by the steroids included the participation of the intracellular signaling pathways PKC, MAPK, and PI3K, which selectively and differentially mediate the stimulation of nitric oxide release. We evidence that estrone, progesterone, and testosterone modulate monocyte and platelet adhesion to endothelial cells, events that play a major role in the initiation and progression of vascular lesions. The steroid action was evidenced under basal or inflammatory conditions. The mechanisms of action exerted by the steroids included stimulation of nitric oxide production and the participation of PKC, MAPK, and PI3K systems.     

Chlamydia pneumoniae-infected monocytes exhibit increased adherence to human aortic endothelial cells

Interactions between monocytes and endothelial cells play an important role in the pathogenesis of atherosclerosis, and monocyte adhesion to arterial endothelium is one of the earliest events in atherogenesis. Work presented in this study examined human monocyte adherence to primary human aortic endothelial cells following monocyte infection with Chlamydia pneumoniae, an intracellular pathogen associated with atherosclerosis by a variety of sero-epidemiological, pathological and functional studies. Infected monocytes exhibited enhanced adhesion to aortic endothelial cells in a time- and dose-dependent manner. Pre-treatment of C. pneumoniae with heat did not effect the organism's capacity to enhance monocyte adhesion, suggesting that heat-stable chlamydial antigens such as chlamydial lipopolysaccharide (cLPS) mediated monocyte adherence. Indeed, treatment of monocytes with cLPS was sufficient to increase monocyte adherence to endothelial cells, and increased adherence of infected or cLPS-treated monocytes could be inhibited by the LPS antagonist lipid X. Moreover, C. pneumoniae-induced adherence could be inhibited by incubating monocytes with a mAb specific to the human beta 2-integrin chain, suggesting that enhanced adherence resulted from increased expression of these adhesion molecules. These data show that C. pneumoniae can enhance the capacity of monocytes to adhere to primary human aortic endothelial cells. The enhanced adherence exhibited by infected monocytes may increase monocyte residence time in vascular sites with reduced wall shear stress and promote entry of infected cells into lesion-prone locations.     

Mechanism of cigarette smoke condensate induced adhesion of human monocytes to cultured endothelial cells

Cigarette smoking is ranked among the leading risk factors in the etiology of atherosclerotic vascular disease. The mechanisms, however, that link cigarette smoking to increased incidence of atherosclerosis are not understood. The adherence of circulating monocytes to the endothelium, migration into the subendothelium, and subsequent formation of foam cells are principal initial events in the development of atherosclerosis. We therefore determined whether cigarette smoke caused increased adherence of monocytes to endothelial cells and the cellular mechanism of this increased adherence. Cigrette smoke condensate (CSC), the particulate fraction of cigarette smoke derived from 2R1 standard research cigarettes, at a concentration of 25–30 μg/ml (average yield of CSC is 26.1 mg/cigarette), augmented (70–90%) basal adherence of human peripheral blood monocytes to a cultured monolayer of endothelial cells derived from bovine aorta (BAEC) and human umbilical vein (HUVEC). There was a concomitant increase in the expression of CD11b ligand on the surface of monocytes as determined by flow cytometry, utilizing FITC conjugated Mab MO-1 (CD11b). However, nicotine (1–15 μg/ml) and cadmium sulfate (10 μg/ml), constituents of CSC, individually or in combination had no effect either on CD11b expression or adherence of monocytes to endothelial cells. Treatment of HUVEC with CSC for 60 min also resulted in an increased expression of ICAM-1 and ELAM-1 as determined by mean fluorescence intensity of ICAM-1 and ELAM-1 labeled cells in flow cytometric analysis. The CSC induced expression of CD11b in monocytes was optimal at 25–30 min and was inhibited by protein kinase C inhibitors, staurosporine and H-7, and also by baicalein, a lipoxygenase inhibitor. Similarly, CSC induced ICAM-1 and ELAM-1 expression in HUVEC was inhibited by protein kinase C inhibitors. CSC stimulated the adherence of human monocytes but not the monocytic cell lines HL-60, U937, and THP-1 to endothelial cells. The CSC stimulated adherence of human monocytes was inhibited (80%) by MAb to CD11b and 50% by Mab to ICAM-1 and ELAM-1. These results suggest that cigarettee smoke particulate constituents activate protein kinase C, leading to increased surface expression of adhesive ligand CD11b on peripheral blood monocytes and counter receptor(s) ICAM-1 and ELAM-1 in endothelial cells. The expression of ligand and counter receptor leads to potentiated adherence of monocytes to endothelial cells, an initial event in the pathogenesis of cigarette smoke induced inflammatory response in the vessel wall. © 1994 Wiley-Liss, Inc.     

CD14 contributes to the adherence of human monocytes to cytokine-stimulated endothelial cells.

Monocyte adherence to endothelial cells (EC) is selectively increased during inflammation. The mechanisms underlying monocyte-EC interaction indicated the involvement of surface-adhesion molecules on monocytes and EC. In earlier studies we noticed that the monocyte-specific mAb, designated mAb 63D3, in contrast to mAb against the beta 2-integrin molecules, inhibited the monocyte binding to monolayers of rIL-1 alpha-stimulated venous EC. The aim of the present study was to further characterize the Ag recognized by mAb 63D3 and to investigate the specific contribution of this Ag to the adherence of monocytes to cultured human macrovascular venous or arterial EC. Flow cytometric analysis demonstrated that the 63D3 Ag is expressed exclusively on the surface of peripheral blood monocytes. SDS-PAGE analysis of mAb 63D3 immunoprecipitates of 125I-labeled human monocyte surface proteins revealed that the target Ag for mAb 63D3 is a 52- to 55-kDa molecule identical to the myeloid differentiation protein CD14. Stimulation of EC with rIL-1 alpha or rTNF-alpha for 4 or 24 h or rIFN-gamma for 24 h increased (p less than 0.005) the number of monocytes bound to both types of EC. This cytokine-induced increase in monocyte adherence was significantly (p less than 0.0005) inhibited when the monocytes were coated with various mAb against CD14. The binding of monocytes to nonstimulated venous or arterial EC was not inhibited by anti-CD14 mAb. Our results lead to the conclusion that CD14 molecules, which on basis of their structure and m.w. are not related to the beta 2-integrin family of heterodimeric leukocyte adhesion molecules, participate in the binding of monocytes to cytokine-stimulated EC.     

Activated leukocyte cell adhesion molecule is a component of the endothelial junction involved in transendothelial monocyte migration

Transendothelial leukocyte migration is a major aspect of the innate immune response. It is essential in repair and regeneration of damaged tissues and is regulated by multiple cell adhesion molecules (CAMs) including members of the immunoglobulin (Ig) superfamily. Activated leukocyte cell adhesion molecule (ALCAM/CD166) is an Ig CAM expressed by activated monocytes and endothelial cells. Hitherto, the functional relevance of ALCAM expression by endothelial cells and activated monocytes remained unknown. In this report, we demonstrate soluble recombinant human ALCAM significantly inhibited the rate of transendothelial migration of monocyte cell lines. Direct involvement of ALCAM in transendothelial migration was evident from the robust inhibition of this process by ALCAM blocking antibodies. However, soluble recombinant ALCAM had no impact on monocyte migration or adhesion to endothelium. Localization of ALCAM specifically at cell-cell junctions in endothelial cells supported its role in transendothelial migration. This study is the first to localize ALCAM to endothelial cell junctions and demonstrate a functional relevance for co-expression of ALCAM by activated monocytes and endothelial cells.     

Alpha and beta subunits of the LFA-1 membrane molecule are involved in human monocyte-endothelial cell adhesion

Human monocyte adhesion to vascular endothelium is an important transitional event in mononuclear phagocyte development. The molecular mechanism involved in monocyte adhesion to endothelial cells was studied using purified human monocytes and a panel of monoclonal antibodies (MAb). The purified human monocytes were phenotypically characterized and expressed relatively low levels of HLA class II antigens. The monocytes were labeled with Indium-111 to provide high specific activity and a sensitive measure of adhesion. Using this radionuclide adhesion assay, monocytes demonstrated consistent and reproducible adhesion to a confluent monolayer of human umbilical vein-derived endothelial cells. To identify the cell surface molecules involved in human monocyte-endothelial cell adhesion, 15 MAb to 11 monocyte surface structures were used to attempt to inhibit adhesion. MAb recognizing 10 monocyte cell surface molecules did not inhibit adhesion. In contrast, MAb recognizing the alpha and beta subunits of LFA-1 (lymphocyte function-associated) significantly inhibited monocyte adhesion to endothelial cells. Monocyte adhesion was comparably inhibited by F(ab')2 and intact MAb. Significant inhibition was observed at 5 micrograms/ml of anti-LFA-1 MAb. These results indicate that the alpha and beta subunits of the LFA-1 membrane molecule are involved in human monocyte-endothelial cell adhesions.     

Leukocyte adhesion to endothelial cells

The adhesion of leukocytes to endothelium is a physiological phenomenon which is the first step for leukocyte emigration. The adhesion can be dramatically increased in pathological situations such as inflammation and vascular diseases. The molecular basis of leukocyte-endothelium interaction has been largely investigated in the last ten years. Using monoclonal antibodies it is possible to characterize the leukocyte adhesion molecule (LeuCAM) also named CD11/CD18 complex. These molecules responsible for leukocyte adhesion are heterodimers consisting of a common beta subunit and different subunit CD11a/CD18 corresponding to LFA-1; CD11b/CD18 to Mac1/Mol; CD11c/CD18 to GP150-95. Beside these receptors, other leukocyte structures such as the fibronectin receptors are involved in the adhesive process. On the endothelial cell side specialized structures implicated in leukocyte adhesion have been identified. Structures like Intercellular Adhesion Molecule (ICAM) are expressed on endothelial cells in the absence of stimulation, while other receptors Endothelial Leukocyte Adhesion Molecule (ELAM) are only detectable on activated endothelial cells. Cytokines such as IL-1 induced the expression of ELAM, increased the number of ICAM and Human Leukocyte Antigens (HLA) DR, DP, DQ. In various pathological circumstances, namely extracorporeal circulation, Acute Respiratory Distress Syndrome (ARDS), hypercholesterolemia and diabetes mellitus increased leukocyte adhesion has been reported and is potentially responsible for vascular damage. Therefore, the modulation of leukocyte-endothelial cell interactions is a possible target for antithrombotic and antiatherosclerotic therapy.     

The P2Y2 nucleotide receptor mediates UTP-induced vascular cell adhesion molecule-1 expression in coronary artery endothelial cells

P2Y2 receptor up-regulation and activation induces intimal hyperplasia and monocyte/macrophage infiltration in the collared rabbit carotid artery model of vascular injury, suggesting a potential role for P2Y2 receptors in monocyte recruitment by vascular endothelium. In this study, we addressed the hypothesis that activation of P2Y2 receptors by extracellular nucleotides modulates the expression of adhesion molecules on vascular endothelial cells that are important for monocyte recruitment. Results indicated that the equipotent P2Y2 receptor agonists UTP or ATP (1-100 microm) stimulated the expression of vascular cell adhesion molecule-1 (VCAM-1) in human coronary artery endothelial cells (HCAEC) in a time- and dose-dependent manner. P2Y2 antisense oligonucleotides inhibited VCAM-1 expression induced by UTP but not by tumor necrosis factor-alpha. Furthermore, UTP induced VCAM-1 expression in human 1321N1 astrocytoma cell transfectants expressing the recombinant P2Y2 receptor, whereas vector-transfected control cells did not respond to UTP. The effect of UTP on VCAM-1 expression in HCAEC was prevented by depletion of intracellular calcium stores with thapsigargin or by inhibition of p38 mitogen-activated protein kinase or Rho kinase, but was not affected by inhibitors of the mitogen-activated protein/extracellular signal-regulated kinase pathway (i.e. MEK1/2). Consistent with a role for VCAM-1 in the recruitment of monocytes, UTP or ATP increased the adherence of monocytic U937 cells to HCAEC, an effect that was inhibited by anti-VCAM-1 antibodies. These findings suggest a novel role for the P2Y2 receptor in the p38- and Rho kinase-dependent expression of VCAM-1 that mediates the recruitment of monocytes by vascular endothelium associated with the development of atherosclerosis.     

Human cytomegalovirus (HCMV) infection of endothelial cells promotes naive monocyte extravasation and transfer of productive virus to enhance hematogenous dissemination of HCMV

Human cytomegalovirus (HCMV) pathogenesis is dependent on the hematogenous spread of the virus to host tissue. While data suggest that infected monocytes are required for viral dissemination from the blood to the host organs, infected endothelial cells are also thought to contribute to this key step in viral pathogenesis. We show here that HCMV infection of endothelial cells increased the recruitment and transendothelial migration of monocytes. Infection of endothelial cells promoted the increased surface expression of cell adhesion molecules (intercellular cell adhesion molecule 1, vascular cell adhesion molecule 1, E-selectin, and platelet endothelial cell adhesion molecule 1), which were necessary for the recruitment of na?ve monocytes to the apical surface of the endothelium and for the migration of these monocytes through the endothelial cell layer. As a mechanism to account for the increased monocyte migration, we showed that HCMV infection of endothelial cells increased the permeability of the endothelium. The cellular changes contributing to the increased permeability and increased na?ve monocyte transendothelial migration include the disruption of actin stress fiber formation and the decreased expression of lateral junction proteins (occludin and vascular endothelial cadherin). Finally, we showed that the migrating monocytes were productively infected with the virus, documenting that the virus was transferred to the migrating monocyte during passage through the lateral junctions. Together, our results provide evidence for an active role of the infected endothelium in HCMV dissemination and pathogenesis.     

Human Thy-1 (CD90) on activated endothelial cells is a counterreceptor for the leukocyte integrin Mac-1 (CD11b/CD18)

Leukocyte recruitment in response to inflammatory signals is in part governed by interactions between endothelial cell receptors belonging to the Ig superfamily and leukocyte integrins. In our previous work, the human Ig superfamily glycoprotein Thy-1 (CD90) was identified as an activation-associated cell adhesion molecule on human dermal microvascular endothelial cells. Furthermore, the interaction of Thy-1 with a corresponding ligand on monocytes and polymorphonuclear cells was shown to be involved in the adhesion of these leukocytes to activated Thy-1-expressing endothelial cells. In this study, we have identified the specific interaction between human Thy-1 and the leukocyte integrin Mac-1 (CD11b/CD18; alphaMbeta2) both in cellular systems and in purified form. Monocytes and polymorphonuclear cells were shown to adhere to transfectants expressing human Thy-1 as well as to primary Thy-1-expressing human dermal microvascular endothelial cells. Furthermore, leukocyte adhesion to activated endothelium as well as the subsequent transendothelial migration was mediated by the interaction between Thy-1 and Mac-1. This additional pathway in leukocyte-endothelium interaction may play an important role in the regulation of leukocyte recruitment to sites of inflammation.