[Protective effect of FAK inhibitor PF-562271 against human umbilical vein endothelial cell injury induced by aging platelets].

OBJECTIVE: To investigate the protective effect of PF-562271, a FAK inhibitor, against aging platelet-induced injury in human umbilical vein endothelial cells (HUVECs). METHODS: Cultured HUVECs were treated with vehicle, lipopolysaccharide (LPS), LPS+aging platelets, or LPS+aging platelets+PF-562271. The changes in protein expressions of FAK, pFAK and PECAM-1 in the treated cells were detected using Western blotting and immunofluorescence assay, and the level of reactive oxygen species (ROS) was detected with flow cytometry. The changes of barrier function of the cells were assessed with cell permeability test and transendothelial cell resistance test. RT-qPCR was used to analyze mRNA expressions of inflammatory factors, and pro-inflammatory cytokine levels in the culture supernatants was determined with enzyme-linked immunosorbent assay. Immunofluorescence assay was used to examine the effect of the ROS inhibitor vitamin C on PECAM-1 expression in the cells with different treatments. RESULTS: Treatment of HUVECs with LPS and aging platelets significantly increased cellular protein expressions of FAK, pFAK and PECAM-1, which were effectively lowered by addition of PF-562271 (P < 0.05). LPS and aged platelets obviously enhanced ROS production in the cells, which was inhibited by the addition of PF-562271 (P < 0.001). PF-562271 significantly alleviated the damage of endothelial cell barrier function of the cells caused by LPS and aging platelets (P < 0.01). The expressions of TNF-α, IL-6 and IL-8 in HUVECs increased significantly after exposure to LPS and aging platelets, and were obviously lowered after treatment with PF-562271 (P < 0.05). Treatment with vitamin C significantly decreased the expression of PECAM-1 protein in the cells (P < 0.01). CONCLUSION: The FAK inhibitor PF-562271 alleviates endothelial cell damage induced by LPS and aging platelets by lowering cellular oxidative stress levels and reducing inflammatory responses.

Platelet endothelial cell adhesion molecule-1 inhibits platelet response to thrombin and von Willebrand factor by regulating the internalization of glycoprotein Ib via AKT/glycogen synthase kinase-3/dynamin and integrin αIIbß3.

OBJECTIVE: Platelet endothelial cell adhesion molecule-1 (PECAM-1) regulates platelet response to multiple agonists. How this immunoreceptor tyrosine-based inhibitory motif-containing receptor inhibits G protein-coupled receptor-mediated thrombin-induced activation of platelets is unknown. APPROACH AND RESULTS: Here, we show that the activation of PECAM-1 inhibits fibrinogen binding to integrin αIIbß3 and P-selectin surface expression in response to thrombin (0.1-3 U/mL) but not thrombin receptor-activating peptides SFLLRN (3×10(-7)-1×10(-5) mol/L) and GYPGQV (3×10(-6)-1×10(-4) mol/L). We hypothesized a role for PECAM-1 in reducing the tethering of thrombin to glycoprotein Ibα (GPIbα) on the platelet surface. We show that PECAM-1 signaling regulates the binding of fluorescein isothiocyanate-labeled thrombin to the platelet surface and reduces the levels of cell surface GPIbα by promoting its internalization, while concomitantly reducing the binding of platelets to von Willebrand factor under flow in vitro. PECAM-1-mediated internalization of GPIbα was reduced in the presence of both EGTA and cytochalasin D or latrunculin, but not either individually, and was reduced in mice in which tyrosines 747 and 759 of the cytoplasmic tail of ß3 integrin were mutated to phenylalanine. Furthermore, PECAM-1 cross-linking led to a significant reduction in the phosphorylation of glycogen synthase kinase-3ß Ser(9), but interestingly an increase in glycogen synthase kinase-3α pSer(21). PECAM-1-mediated internalization of GPIbα was reduced by inhibitors of dynamin (Dynasore) and glycogen synthase kinase-3 (CHIR99021), an effect that was enhanced in the presence of EGTA. CONCLUSIONS: PECAM-1 mediates internalization of GPIbα in platelets through dual AKT/protein kinase B/glycogen synthase kinase-3/dynamin-dependent and αIIbß3-dependent mechanisms. These findings expand our understanding of how PECAM-1 regulates nonimmunoreceptor signaling pathways and helps to explains how PECAM-1 regulates thrombosis.

Effect of intracavernous administration of angiopoietin-4 on erectile function in the streptozotocin-induced diabetic mouse.

INTRODUCTION: Erectile dysfunction (ED) is a highly prevalent complication of diabetes, and the severity of endothelial dysfunction is one of the most important factors in reduced responsiveness to oral phosphodiesterase type 5 inhibitors. AIM: To study the effects of human angiopoietin-4 (Ang-4) protein on erectile function in diabetic mice. METHODS: Diabetes was induced by intraperitoneal injection of streptozotocin into 8-week-old C57BL/6J male mice. At 8 weeks after the induction of diabetes, the animals were divided into four groups: control nondiabetic mice and diabetic mice receiving two successive intracavernous injections of phosphate buffered saline (days -3 and 0), a single intracavernous injection of Ang-4 protein (day 0), or two successive intracavernous injections of Ang-4 protein (days -3 and 0). MAIN OUTCOME MEASURES: One week after treatment, we measured erectile function by electrical stimulation of the cavernous nerve. The penis was harvested and stained with hydroethidine or antibodies to Ang-4, platelet/endothelial cell adhesion molecule-1, and phosphorylated endothelial nitric oxide synthase (eNOS). We also determined the differential expression of Ang-4 in cavernous tissue in the control and diabetic mice. The effect of Ang-4 protein on the phosphorylation of Tie-2, Akt, and eNOS was determined in human umbilical vein endothelial cells (HUVECs) by Western blot. RESULTS: The cavernous expression of Ang-4 was downregulated in diabetic mice; Ang-4 was mainly expressed in endothelial cells. Local delivery of Ang-4 protein significantly increased cavernous endothelial content, induced eNOS phosphorylation, and decreased the generation of superoxide anion and apoptosis in diabetic mice. Ang-4 protein strongly increased the phosphorylation of Tie-2, Akt, and eNOS in HUVECs. Repeated intracavernous injections of Ang-4 induced significant restoration of erectile function in diabetic mice (87% of control values), whereas a single intracavernous injection of Ang-4 protein elicited modest improvement. CONCLUSIONS: Cavernous endothelial regeneration by use of Ang-4 protein may have potential for the treatment of vascular disease-induced ED, such as diabetic ED.

Platelet endothelial cell adhesion molecule targeted oxidant-resistant mutant thrombomodulin fusion protein with enhanced potency in vitro and in vivo.

Thrombomodulin (TM) is a glycoprotein normally present in the membrane of endothelial cells that binds thrombin and changes its substrate specificity to produce activated protein C (APC) that has antithrombotic and anti-inflammatory features. To compensate for loss of endogenous TM in pathology, we have fused recombinant TM with single chain variable fragment (scFv) of an antibody to mouse platelet endothelial cell adhesion molecule-1 (PECAM). This fusion, anti-PECAM scFv/TM, anchors on the endothelium, stimulates APC production, and provides therapeutic benefits superior to sTM in animal models of acute thrombosis and inflammation. However, in conditions of oxidative stress typical of vascular inflammation, TM is inactivated via oxidation of the methionine 388 (M388) residue. Capitalizing on the reports that M388L mutation renders TM resistant to oxidative inactivation, in this study we designed a mutant anti-PECAM scFv/TM M388L. This mutant has the same APC-producing capacity and binding to target cells, yet, in contrast to wild-type fusion, it retains APC-producing activity in an oxidizing environment in vitro and in vivo. Therefore, oxidant resistant mutant anti-PECAM scFv/TM M388L is a preferable targeted biotherapeutic to compensate for loss of antithrombotic and anti-inflammatory TM functions in the context of vascular oxidative stress.

A CD31-derived peptide prevents angiotensin II-induced atherosclerosis progression and aneurysm formation.

AIMS: The loss of the inhibitory receptor CD31 on peripheral T lymphocytes is associated with the incidence of atherosclerotic complications such as abdominal aortic aneurysms (AAA) in patients and plaque thrombosis in mice. However, we have recently discovered that a small fragment of extracellular CD31 remains expressed on the surface of the apparently 'CD31-negative' T-cells and that it is possible to restore the CD31-mediated T-cell inhibition in vivo by using a synthetic CD31-derived peptide. Here, we wanted to evaluate the therapeutic potential of the peptide in an experimental model of accelerated atherosclerosis and AAA formation. METHODS AND RESULTS: The effect of the murine CD31-derived peptide (aa 551-574, 1.5 mg/kg/day, sc) was evaluated on the extent of atherosclerotic plaques and the incidence of AAA in 28-week-old apolipoprotein E knockout mice (male, n ≥ 8/group) submitted to chronic angiotensin II infusion. The therapeutic mechanisms of the peptide were assessed by evaluating its effect on immune cell functions in vivo and in vitro. The prevalence of angiotensin II-induced AAA correlated with the loss of extracellular CD31 on T-cells. CD31 peptide treatment reduced both aneurysm formation and plaque size (P < 0.05 vs. control). Protection was associated with reduced perivascular leucocyte infiltration and T-cell activation in vivo. Functional in vitro studies showed that the peptide is able to suppress both T-cell and macrophage activation. CONCLUSION: CD31 peptides could represent a new class of drugs intended to prevent the inflammatory cell processes, such as those underlying progression of atherosclerosis and development of AAA.

Nitric oxide reduces T lymphocyte adhesion to human brain microvessel endothelial cells via a cGMP-dependent pathway.

The entry of lymphocytes into the brain is normally limited by the blood-brain barrier, however, during inflammation prominent lymphocytic infiltration occurs. In this study, we investigated the effects of nitric oxide (NO) on the adhesion of T cells to cultured human brain microvessel endothelial cells. T cell adhesion to unstimulated or tumor necrosis factor-alpha (TNF-alpha)-treated cells was quantified by counting the number of lymphocytes bound to the monolayer by light microscopy. TNF-alpha increased T cell adhesion in a time-dependent manner. Incubation of monolayers with NO donors decreased adhesion. This effect was blocked by a guanylyl cyclase inhibitor and mimicked by a cGMP agonist, and was thus dependent on the generation of cGMP. NO did not modulate adhesion molecule expression in the endothelial cells, suggesting an action on the T cells. Pre-treatment of T cells with NO or a cGMP agonist decreased binding to recombinant endothelial adhesion molecules. These findings suggest that NO can modulate the adhesion of T cells to human brain microvessel endothelial cells via a cGMP-dependent mechanism, and may thus regulate lymphocyte traffic during central nervous system inflammation.

PECAM-1-dependent neutrophil transmigration is independent of monolayer PECAM-1 signaling or localization.

Platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31), a tyrosine phosphoprotein highly expressed on endothelial cells and leukocytes, is an important component in the regulation of neutrophil transendothelial migration. Engagement of endothelial PECAM-1 activates tyrosine phosphorylation events and evokes prolonged calcium transients, while homophilic engagement of neutrophil PECAM-1 activates leukocyte beta-integrins. Although PECAM-1 modulates polymorphoneutrophil transmigration via homophilic PECAM-1-PECAM-1 interaction, the mechanisms underlying endothelial PECAM-1 function are unknown. Proposed mechanisms include (1) formation of a haptotactic gradient that "guides" neutrophils to the cell-cell border, (2) service as a "passive ligand" for neutrophil PECAM-1, ultimately mediating activation of neutrophil beta integrins, (3) regulation of endothelial calcium influx, and (4) mediation of SH2 protein association, and/or (5) catenin and non-SH2 protein interaction. Utilizing PECAM-1-null "model" endothelial cells (REN cells), we developed a neutrophil transmigration system to study PECAM-1 mutations that specifically disrupt PECAM-1-dependent signaling and/or PECAM-1 cell localization. We report that interleukin-1 beta (IL-1 beta) elicits PECAM-1-dependent transmigration that requires homophilic PECAM-PECAM-1 engagement, but not heterophilic neutrophil PECAM-1 interactions, and is intercellular adhesion molecule-1 dependent. Conversely, whereas IL-8 and leukotriene-B(4)-mediated transmigration is PECAM-1-independent, PECAM-1 and IL-8-dependent transmigration represent separable and additive components of cytokine-induced transmigration. Surprisingly, neither monolayer PECAM-1-regulated calcium signaling, cell border localization, nor the PECAM-1 cytoplasmic domain was required for monolayer PECAM-1 regulation of neutrophil transmigration. We conclude that monolayer (endothelial cell) PECAM-1 functions as a passive homophilic ligand for neutrophil PECAM-1, which after engagement leads to neutrophil signal transduction, integrin activation, and ultimately transmigration in a stimulus-specific manner.

Platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31) acts as a regulator of B-cell development, B-cell antigen receptor (BCR)-mediated activation, and autoimmune disease.

Platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31) is an immunoglobulin-immunoreceptor tyrosine-based inhibitory motif (Ig-ITIM) superfamily member that recruits and activates protein-tyrosine phosphatases, SHP-1 and SHP-2, through its intrinsic ITIMs. PECAM-1-deficient (PECAM-1(-/-) ) mice exhibit a hyperresponsive B-cell phenotype, increased numbers of B-1 cells, reduced B-2 cells, and develop autoantibodies. In the periphery, there are reduced mature recirculating B-2 cells and increased B-1a cells within the peritoneal cavity. In addition, PECAM-1(-/-) B cells display hyperproliferative responses to lipopolysaccharide and anti-IgM stimulation and showed enhanced kinetics in their intracellular Ca(++) response following IgM cross-linking. PECAM-1(-/-) mice showed increased serum levels of IgM with elevated IgG isotypes and IgA antidinitrophenol antibody in response to the T-independent antigen, dinitrophenol-Ficoll. Finally, PECAM-1(-/-) mice developed antinuclear antibodies and lupuslike autoimmune disease with age.

Macrophage colony stimulating factor modulates the development of hematopoiesis by stimulating the differentiation of endothelial cells in the AGM region.

Definitive hematopoietic stem cells arise in the aorta-gonad-mesonephros (AGM) region from hemangioblasts, common precursors for hematopoietic and endothelial cells. Previously, we showed that multipotential hematopoietic progenitors and endothelial cells were massively produced in primary culture of the AGM region in the presence of oncostatin M. Here we describe a role for macrophage-colony-stimulating factor (M-CSF) in the development of hematopoietic and endothelial cells in AGM culture. The number of hematopoietic progenitors including multipotential cells was significantly increased in the AGM culture of op/op embryos. The addition of M-CSF to op/op AGM culture decreased colony-forming unit (CFU)-GEMM, granulocyte macrophage-CFU, and erythroid-CFU, but it increased CFU-M. On the other hand, the number of cells expressing endothelial markers, vascular endothelial-cadherin, intercellular adhesion molecule 2, and Flk-1 was reduced in op/op AGM culture. The M-CSF receptor was expressed in PCLP1(+)CD45(-) cells, the precursors of endothelial cells, and M-CSF up-regulated the expression of more mature endothelial cell markers-VCAM-1, PECAM-1, and E-selectin-in PCLP1(+)CD45(-) cells. These results suggest that M-CSF modulates the development of hematopoiesis by stimulating the differentiation of PCLP-1(+)CD45(-) cells to endothelial cells in the AGM region.

Expression of IL-6 and IL-6 receptors by circulating clonotypic B cells in multiple myeloma: potential for autocrine and paracrine networks.

OBJECTIVE: To investigate the participation of clonotypic MM B cells in the IL-6 network in patients with multiple myeloma. METHODS: CD19(+) B cells from 45 patients with multiple myeloma and from 18 healthy donors were sorted and their expression of IL-6, IL-6 receptor (CD126) characterized by flow cytometry, in situ RT-PCR, and ELISA measurement of IL-6 and soluble IL-6R. Expression of CD31 was detected by flow cytometry. RESULTS: Interleukin-6 (IL-6) is a pleiotropic cytokine often overexpressed in multiple myeloma (MM). IL-6 induces growth and inhibits apoptosis of MM plasma cells, and upregulates the activity of osteoclasts. MM plasma cells, the most mature component of the MM clone, secrete IL-6 and induce IL-6 production from other cell types. However, the MM clone also includes circulating clonotypic B lymphocytes. Using ELISA and in situ RT-PCR we demonstrate here that, unlike the healthy control B cells, MM B cells express IL-6 mRNA and secrete IL-6 protein. In vitro, MM B cells were the major producers of IL-6 in peripheral blood mononuclear cells. On average, 50% of MM B cells express the IL-6 receptor (IL-6R, CD126), suggestive of autocrine stimulation. They also express CD31, potentially facilitating their paracrine interactions with osteoclast precursors. CONCLUSION: Secretion of IL-6 by circulating clonotypic B cells in MM may contribute to the autocrine and paracrine cytokine networks that maintain the malignant clone and are responsible for disruption of normal bone metabolism in this incurable disease.

Binding of Plasmodium falciparum-infected erythrocytes to soluble platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31): frequent recognition by clinical isolates.

Platelet-endothelial cell adhesion molecule-1 or CD31 (PECAM-1/CD31) is a receptor recognized by Plasmodium falciparum-parasitized erythrocytes (pRBCs). Fluorescence-labeled soluble recombinant PECAM-1/CD31 (sPECAM-1/CD31) is shown to bind to the surface of P. falciparum-infected erythrocytes on up to 70% of the cells. Binding is blocked by the addition of the unlabeled receptor in a dose-dependent fashion, but not by unrelated receptor-proteins. A significant correlation was found between the binding of sPECAM-1/CD31 to pRBCs and the binding to transfected L cells expressing the receptor as seen with six different P. falciparum lines or clones. Panning of cultures on PECAM-1/CD31 transfected L cells was paralleled by an increase in the binding of sPECAM-1/CD31. The pRBCs of 54% of fresh patient-isolates bound sPECAM-1/CD31 with a mean rate of 12.9% (range = 1.1-44%). The data suggest that PECAM-1/CD31 is a common receptor recognized by wild isolates and that the soluble PECAM-1/CD31 suspension assay is a sensitive and reliable way to study PECAM-1/CD31 binding.

Platelet endothelial cell adhesion molecule-1 serves as an inhibitory receptor that modulates platelet responses to collagen.

Platelet responses to collagen are mediated by the combined actions of the integrin alpha2beta1, which serves as a major collagen-binding receptor, and the GPVI/FcRgamma-chain complex, which transmits collagen-specific activation signals into the cell interior through the action of an immunoreceptor tyrosine-based activation motif within the cytoplasmic domain of the FcRgamma-chain. Despite much progress in identifying components of the signaling pathway responsible for collagen-induced platelet activation, virtually nothing is known about the regulatory elements that modulate this important hemostatic event. PECAM-1, a recently recognized member of the inhibitory receptor family, contains a functional immunoreceptor tyrosine-based inhibitory motif within its cytoplasmic domain that, when tyrosine phosphorylated, recruits and activates the protein-tyrosine phosphatase, SHP-2. To test the hypothesis that PECAM-1 functions to regulate GPVI/FcRgamma-chain-mediated platelet activation, the responses of wild-type versus PECAM-1-deficient murine platelets to GPVI-specific agonists were compared. Four distinct GPVI/FcRgamma-chain-dependent responses were found to be significantly exaggerated in platelets derived from PECAM-1-deficient mice, including Mg++-independent adhesion to immobilized fibrillar collagen, collagen-induced platelet aggregation, platelet aggregation induced by the GPVI-specific agonist collagen-related peptide, and GPVI/FcRgamma-chain-induced dense granule secretion. Together, these data provide compelling evidence that PECAM-1 modulates platelet responses to collagen, and they implicate this novel member of the inhibitory receptor family in the regulation of primary hemostasis.

Platelet-endothelial cell adhesion molecule-1 (PECAM-1)-deficient mice demonstrate a transient and cytokine-specific role for PECAM-1 in leukocyte migration through the perivascular basement membrane.

Studies with neutralizing antibodies have indicated roles for platelet-endothelial cell adhesion molecule-1 (PECAM-1) in leukocyte migration through the endothelium and the perivascular basement membrane. Because some of these findings have been contentious, this study aimed to explore the role of PECAM-1 in leukocyte migration by analyzing leukocyte responses in interleukin 1beta (IL-1beta)- and tumor necrosis factor-alpha (TNFalpha)-activated cremasteric venules of PECAM-1-deficient mice using intravital and electron microscopy. Although no differences in levels of leukocyte rolling flux or firm adhesion were observed, a delay in leukocyte transmigration in response to IL-1beta, but not TNFalpha, was detected in PECAM-1-deficient mice. Electron microscopy indicated that this delay occurred at the level of perivascular basement membrane. To address the cytokine specificity of PECAM-1 dependence, in vitro experiments demonstrated that TNFalpha, but not IL-1beta, could induce rapid adhesion of murine neutrophils to protein-coated surfaces, suggesting that TNFalpha elicited leukocyte transmigration in wild-type mice via direct stimulation of leukocytes. In summary, the results suggest a regulatory role for PECAM-1 in leukocyte migration through the perivascular basement membrane, a role that appears to be cytokine-specific and associated with the ability of the cytokine to stimulate rapid neutrophil adhesion.

Lack of in vivo function of CD31 in vascular thrombosis.

A murine model of endothelial cell injury-based vascular thrombosis was used to test the role of platelet-endothelial cell adhesion molecule-1 (PECAM-1, CD31) in blood cell aggregate formation and vessel occlusion in vivo. Photochemically-induced thrombus formation was analyzed in detail using intravital fluorescence microscopy of individual microvessels in cremaster muscle preparations of CD31-deficient and wildtype mice. In venules, epi-illumination induced rapid thrombus formation with first platelet deposition after 0.56 +/- 0.11 min and complete vessel occlusion within 5.05 +/- 0.45 min. In arterioles, thrombus formation was markedly delayed with first platelet deposition after 3.03 +/- 0.47 min and complete vessel occlusion within 10.04 +/- 1.26 min. Kinetics of thrombus formation in both venules (first platelet deposition: 0.52 +/- 0.1 min; vessel occlusion: 5.03 +/- 0.52 min) and arterioles (first platelet deposition: 3.06 +/- 0.68 min; vessel occlusion: 10.02 +/- 1.38 min) of CD31-deficient mice was found almost identical compared with that in wildtype animals. Tail bleeding time was 233 +/- 24 s in wildtype and 243 +/- 32 s in CD31-deficient mice. Moreover, CD31-deficient and wildtype mice revealed comparable interaction of leukocytes to endothelium. This study shows for the first time in vivo that CD31 is not critically involved in blood cell thrombus formation upon endothelial cell injury.

Chemotactic properties of ICAM-1 and PECAM-1 on neutrophil granulocytes in ulcerative colitis: effects of prednisolone and mesalazine.

BACKGROUND: ICAM-1 seems to exhibit effects other than passive leucocyte/endothelial interaction. AIM: To investigate the attracting properties of selected adhesion molecules, assessing the influence of the two major anti-inflammatory drugs in ulcerative colitis, prednisolone and mesalazine. METHODS: Circulating neutrophils (11 ulcerative colitis, 15 controls) were assessed in microchemotaxis chambers by the leading front technique, using physiologically relevant concentrations of ICAM-1 (0.005-5000 pM), PECAM-1 (0.001-1000 nM), and P-selectin (0.01-100 nM). Neutrophils pre-incubated with prednisolone (10(-8)-10(-4) M) or mesalazine (0.65-10. 4 nM) were assessed towards ICAM-1. RESULTS: Migration of neutrophils towards ICAM-1 showed a bell-shaped curve with a maximum at 5 pM (migration: 37.7 microm; P<0.001), whereas PECAM-1 attracted neutrophils equally in the range of 0.1-10 nM (25.0 microm; P<0.001). P-selectin had no cell-attracting effect. No differences were detected between cells from ulcerative colitis patients and controls. Pre-treatment with prednisolone decreased the cell attracting effect of ICAM-1 in a dose-dependent manner to 72% of the basal migration (P<0.001). Conversely, prednisolone showed a pro-chemokinetic effect by increasing the spontaneous locomotion of neutrophils by 40% (P<0.001). CONCLUSIONS: Specific chemotactic properties were observed for ICAM-1 and PECAM-1. Prednisolone exhibited a dual effect in inhibiting the ICAM-1-mediated migration and stimulating the general locomotion of neutrophils.

Soluble domain 1 of platelet-endothelial cell adhesion molecule (PECAM) is sufficient to block transendothelial migration in vitro and in vivo.

The inflammatory response involves sequential adhesive interactions between cell adhesion molecules of leukocytes and the endothelium. Unlike the several adhesive steps that precede it, transendothelial migration (diapedesis), the step in which leukocytes migrate between apposed endothelial cells, appears to involve primarily one adhesion molecule, platelet-endothelial cell adhesion molecule (PECAM, CD31). Therefore, we have focused on PECAM as a target for antiinflammatory therapy. We demonstrate that soluble chimeras made of the entire extracellular portion of PECAM, or of only the first immunoglobulin domain of PECAM, fused to the Fc portion of IgG, block diapedesis in vitro and in vivo. Furthermore, the truncated form of the PECAM-IgG chimera does not bind stably to its cellular ligand. This raises the possibility of selective anti-PECAM therapies that would not have the untoward opsonic or cell-activating properties of antibodies directed against PECAM.

Antibody mediated ligation of platelet/endothelial cell adhesion molecule-1 (PECAM-1) on neutrophils enhances adhesion to cultured human dermal microvascular endothelial cells.

We investigated the effects on leukocyte adhesion to cultured human dermal microvascular endothelial (DMvE) cell by antibody mediated ligation of platelet/endothelial cell adhesion molecule-1 (PECAM-1) on neutrophils. Preincubation of neutrophils with anti-PECAM-1 antibody markedly enhanced their adhesion to DMvE cells, while the adhesion-increasing action was cancelled out by anti-CR3 or anti-LFA-1 beta (CD18) antibody. These findings suggest that ligation of PECAM-1 on neutrophils increases their adhesion to cultured DMvE cells depending on the beta 2 integrin family, and PECAM-1 may have a crucial role in dermal inflammation.