Cell Adhesion Molecule PECAM-1 Research Articles

Impaired production of platelet-endothelial cell adhesion molecules as an early diagnostic marker of heart transplant rejection

Reveal the involvement of platelet-endothelial cell adhesion molecules PECAM-1 in the transplanted heart rejection pathogenesis and determine the CD31 marker significance for biopsy diagnostics of the process form and degree. Sections from endomyocardial biopsies of 56 heart transplant recipients were stained with hematoxylin and eosin. The streptavidin-biotin method was used to determine the expression of T-lymphocytes (CD3), B-lymphocytes (CD20), macrophages (CD68) and the C4d component of complement to determine the form and degree of graft rejection. Additionally, the expression of platelet-endothelial cell adhesion molecules PECAM-1 (CD31) was detected. Using computer morphometry in digital images, the area of pathological changes and the area of CD31 expression was measured with the calculation of the staining area coefficient. The highest levels of PECAM-1 expression were found in the absence of a heart transplant rejection. The degree of rejection of 1R is characterized by a decrease in expression by 1.3 times, when there are no significant signs of necrosis in the myocardium, the area of which increases sharply at degree 2R by 163.7 times, and at 3R by 570.7 times compared with 1R. The process proceeds in parallel with a further decrease in the level of CD31 expression and is accompanied by the development of hemorrhagic manifestations. The intensity of hemorrhages in the myocardium increases by 7.3 times with grade 3R compared with 1R. Expression of PECAM-1 reflects the state of the vascular bed of the heart transplant. Its decrease can be considered as an early pathomorphological marker of transplanted heart rejection. The expression of CD31 continues to decrease with increasing severity of rejection and is accompanied by the progressive development of necrosis and hemorrhages in the graft heart muscle.

Vascular channels formed by subpopulations of PECAM1+ melanoma cells.

Targeting the vasculature remains a promising approach for treating solid tumors; however, the mechanisms of tumor neovascularization are diverse and complex. Here we uncover a new subpopulation of melanoma cells that express the vascular cell adhesion molecule PECAM1, but not VEGFR-2, and participate in a PECAM1-dependent form of vasculogenic mimicry (VM). Clonally-derived PECAM1+ tumor cells coalesce to form PECAM1-dependent networks in vitro and they generate well-perfused, VEGF-independent channels in mice. The neural crest specifier AP-2α is diminished in PECAM1+ melanoma cells and is a transcriptional repressor of PECAM1. Reintroduction of AP-2α into PECAM1+ tumor cells represses PECAM1 and abolishes tube-forming ability whereas AP-2α knockdown in PECAM1− tumor cells up-regulates PECAM1 expression and promotes tube formation. Thus, VM-competent subpopulations, rather than all cells within a tumor, may instigate VM, supplant host-derived endothelium, and form PECAM1-dependent conduits that are not diminished by neutralizing VEGF.

Angiopoietins-1 and -2 play opposing roles in endothelial sprouting of embryoid bodies in 3D culture and their receptor Tie-2 associates with the cell–cell adhesion molecule PECAM1

Angiopoietins 1 and 2, ligands for the receptor kinase Tie-2, have been proposed to play critical but opposing roles in vascular development. Since signaling by Tie-2 is likely affected by other endothelial cell receptors such as Flk-1, the receptor for VEGF, and cell–cell adhesion receptors PECAM1 and VE-cad, we explored their interactions in a 3D model of vasculogenesis. When murine embryoid bodies (EBs) were treated with VEGF in Matrigel in the presence or absence of Ang-1 or Ang-2 for eight days, Ang-1 abrogated vascular sprouting for treatments started at days 0 or 3. In contrast, Ang-2 greatly accelerated vascular sprouting compared to untreated EBs. These results were confirmed in a second model system where VEGF treated HUVECs were grown in Matrigel in the presence or absence of Ang-1 or Ang-2. Since vascular sprouting must be precisely controlled in the developing embryo, it is likely that cell–cell adhesion molecules play a role in sensing the density of vascular sprouts. In this respect, we have shown that PECAM1 and CEACAM1 play essential roles in vascular sprouting. We now show that PECAM1 is associated with Tie-2, becomes phosphorylated on its ITIMs, and recruits the inhibitory phosphatases SHP-1 and SHP-2. In addition, PECAM1 is associated with VE-cad and may similarly regulate its signaling via recruitment of SHP-1/2.

Selective Inhibitors of Kv11.1 Regulate IL-6 Expression by Macrophages in Response to TLR/IL-1R Ligands

The mechanism by which the platelet-endothelial cell adhesion molecule PECAM-1 regulates leukodiapedesis, vascular endothelial integrity, and proinflammatory cytokine expression in vivo is not known. We recently identified PECAM-1 as a negative regulator of Kv11.1, a specific voltage-gated potassium channel that functioned in human macrophages to reset a resting membrane potential following depolarization. We demonstrate here that dofetilide (DOF), a selective inhibitor of the Kv11.1 current, had a profound inhibitory effect on neutrophil recruitment in mice following TLR/IL-1R–elicited peritonitis or intrascrotal injection of IL-1β, but had no effect on responses seen with TNFα. Furthermore, inhibitors of Kv11.1 (DOF, E4031, and astemizole), but not Kv1.3 (margatoxin), suppressed the expression of IL-6 and MCP-1 cytokines by murine resident peritoneal macrophages, while again having no effect on TNFα. In contrast, IL-6 expression by peritoneal mesothelial cells was unaffected. Using murine P388 cells, which lack endogenous C/EBPβexpression and are unresponsive to LPS for the expression of both IL-6 and MCP-1, we observed that DOF inhibited LPS-induced expression of IL-6 mRNA following ectopic expression of wild-type C/EBPβ, but not a serine-64 point mutant. Finally, DOF inhibited the constitutive activation of cdk2 in murine peritoneal macrophages; cdk2 is known to phosphorylate C/EBPβ at serine-64. Taken together, our results implicate a potential role for Kv11.1 in regulating cdk2 and C/EBPβ activity, where robust transactivation of both IL-6 and MCP-1 transcription is known to be dependent on serine-64 of C/EBPβ. Our data might also explain the altered phenotypes displayed by PECAM-1 knockout mice in several disease models.

TRANSLATIONAL IMPACT

Atherosclerosis is a progressive, chronic inflammatory disease of large arteries in which lesions preferentially occur at vessel sites exposed to rapid changes in blood flow. Low density lipoprotein (LDL) deposition and leukocyte infiltration leads to plaque formation and narrowing of the artery (stenosis), which restricts blood flow and causes angina. Atherosclerosis can also induce thrombus formation if lipids from plaques come into contact with blood, and plaque associated weakening of artery walls can lead to eventual rupture (aneurysm).PECAM-1 is a membrane protein expressed on the surface of many cells in the blood, as well as the endothelial cells lining blood vessels. PECAM-1 participates in the adhesion cascade necessary for leukocyte extravasation and serves as a sensor responding to changes in mechanical shear force that result from changes in blood flow.In this report, the authors determine that PECAM-1 induces susceptibility to atherosclerosis. Using mice deficient in the cell adhesion molecule PECAM-1 (PECAM-1−/−), researchers found a 50–60% reduction of atherosclerotic lesions in the aortae of mice with a genetic susceptibility to atherosclerosis (ApoE−/−). Since some athlerosclerotic lesions are found in PECAM-1−/− animals, it is likely that the absence of PECAM-1 is not completely preventative, but delays the onset of pathology.PECAM-1 functions in processes of inflammation and in the sensing of shear stress related to blood flow. It is possible that one or both of these processes contribute to the development and progression of atherosclerosis, suggesting that PECAM-1 is a potential candidate for drug targeting to limit the disease process.

Short-term sPECAM-Fc treatment ameliorates EAE while chronic use hastens onset of symptoms

The homophilic cell adhesion molecule PECAM-1 is a major participant in the migration of leukocytes across endothelium. We examined the ability of a chimeric soluble PECAM-1 fused to human IgG-Fc to impair leukocyte entry through the blood–brain barrier and reduce CNS autoimmunity. sPECAM-Fc impaired migration of lymphocytes across brain endothelial monolayers and diminished the severity of EAE, an experimental model of MS, when administered at the onset of symptoms. However, in mice transgenic for sPECAM-Fc, the chronically elevated levels of sPECAM-Fc hastened onset of EAE disease without significantly changing clinical score severity. Our data suggest that short-term treatment of diseases like MS with sPECAM-Fc has therapeutic potential.

Ultrastructural and morphometric studies related to expression of the cell adhesion molecule PECAM-1/CD31 in developing rat lung.

It has recently been postulated that platelet/endothelial cell adhesion molecule-1 (PECAM-1/CD31) might play a role in vascular tube formation. To evaluate the role of PECAM-1/CD31 in the formation of the capillary network in vivo, we conducted an ultrastructural immunohistochemical evaluation of the localization of PECAM-1/CD31 and its developmentally regulated expression in the periphery of the lungs of fetal, newborn, and adult rats. PECAM-1/CD31 was present mainly on luminal surfaces and at the junctions between endothelial cells. Moreover, in fetal lung, products of the immunoreaction were also found on the abluminal surfaces of endothelial cells. To relate those findings to the developmental changes in the capillary area of the lung, we performed a morphometric study of electronmicrographs. The cross-sectional area of blood vessels at the periphery of the lungs was significantly greater in 15-19-day-old fetuses than in postpartum animals (p<0.0001). Disappearance of the expression of PECAM-1/CD31 on the abluminal endothelial surface paralleled the changes in the cross-sectional area of blood vessels that occurred during the perinatal period. (J Histochem Cytochem 48:1283-1289, 2000)

Arrest of Vascular Development Correlates with the Failure of Platelet Endothelial Factor-1 (PECAM-1) Dephosphorylation in Hyperglycemia-Induced Embryopathy • 366

The vascular system is the first organ system to develop in the embryo and its maldevelopment induced by hyperglycemia has serious consequences including embryonic and perinatal lethality. We previously reported that the state of phosphorylation/dephosphorylation of the vascular system specific cell adhesion molecule PECAM-1 correlates with development of vitelline vasculature. PECAM-1 is highly tyrosine phosphorylated prior to vessel formation, and dephosphorylated when the interconnecting circulation between the yolk sac and the embryo is established. We hypothesized that in diabetes-related embryopathy there is alteration of PECAM-1 phosphorylation state and this adversely affects an integrin mediated signal transduction pathway. We utilized an in vitro whole embryo culture system as a model for in situ vasculogenesis. Conceptuses were cultured between 7.5 and 9.5 days p.c. in control (glucose levels: 7.4-9.5 mMol) and in hyperglycemic (25 mMol) rat serum for 48 hr. An arrest of yolk sac vessel development at the primitive capillary plexus stage with aberrant arborization was present in the hyperglycemic condition. This yolk sac vasculopathy always accompanied a malformation of the embryos (malformation in axial rotation, open neural tube, heart defects) when conceptuses were harvested prior to the late headfold stage. A 3 hr period of hyperglycemia was sufficient to elicit yolk sac vasculopathy during this susceptible time of development. Exposure to the same high glucose environment in conceptuses past the late headfold stage was not sufficient to elicit yolk sac vasculopathy. Immunoprecipitation-Western blot analyses revealed that PECAM-1 remained highly tyrosine phosphorylated in the hyperglycemia-exposed malformed conceptuses at the end of organogenesis, in contrast to its dephosphorylated state in normally developing age matched controls. We also found that PECAM-1 in hyperglycemia-exposed conceptuses co-precipitates with the phosphatase SHPTP2. These data indicate that the cell adhesion molecule PECAM-1 is involved in vasculogenesis and its dephosphorylation is delayed/arrested in hyperglycemia-related embryopathy. Thus, modulation of PECAM-1 cytoplasmic tyrosine residues Y663 and Y686 which comprise part of PECAM-1's ITAM domain is postulated to have effect on signaling and/or adaptor molecule binding and subsequent transduction pathways influencing vasculo-and angiogenesis.

Expression of adhesion molecules on endothelial cells after contact with knitted Dacron

The aim of this study is to evaluate the expression of some adhesion molecules on the surface of endothelial cells cultured in contact with knitted Dacron ®. These molecules, as mediators of cell adhesion, could play a role in the modulation of adhesion on the biomaterials, therefore conditioning the response of tissues to implant. Twenty different cultures of human umbilical vein endothelial cells (HUVECs) were cultured in contact with knitted Dacron. Both HUVECs grown without the material and HUVECs incubated with endotoxin were used as control. After 24 h, the cell adhesion molecules PECAM-1, ELAM-1, ICAM-1 and VCAM-1 were evaluated on the cells by monoclonal antibodies and flow cytometry. After 24 h of contact with knitted Dacron, a significant decrease in the proportion of cells expressing PECAM-1 was observed, as well as a significant increase in the proportion of cells expressing ELAM-1. The contact with knitted Dacron did not induce significant variations of ICAM-1 and VACM-1. The incubation with endotoxin determined a significant increase in the proportion of ELAM-1-positive cells, a significant increase in ICAM-1 fluorescence intensity, and a significant increase both in fluorescence intensity and in the proportion of VCAM-1-positive cells. The results obtained with the endotoxin are in agreement with those reported in the literature. The ELAM-1 increase, observed after contact with knitted Dacron, could favour leucocyte adhesion, while the decrease in PECAM-1 expression could result from an inhibiting effect on the endothelial cell adhesion so as to hinder the mechanisms involved in the endothelialization of the material. The variations were interpreted as inhibiting endothelialization and favouring the leucocyte adhesion effect by knitted Dacron.

Biosynthesis and processing of the cell adhesion molecule PECAM-1 includes production of a soluble form.

PECAM-1 (CD31) is a 130-kDa glycoprotein found on platelets, endothelial cells, granulocytes, and monocytes, as well as on certain myelomonocytic cell lines. Recent studies have shown that PECAM-1 may be involved in activation of leukocyte integrins and may also be involved in adhesive interactions of circulating leukocytes and the vessel wall. In spite of the important functional role that PECAM-1 plays in these processes, little is known about the biosynthesis, processing, and turnover of PECAM-1 on the cell surface. We have studied the biosynthesis of PECAM-1 in the promonocytic cell line U937, and in endothelial cells, by pulse-chase labeling and immunoprecipitation. PECAM-1 was synthesized as a 110-kDa precursor form, which was processed into the 130-kDa mature form within 1-3 h, during which time it began to move to the cell surface. The protein disappeared from the cell surface in both cell types about 48 h after labeling. A soluble form of PECAM-1, which is 5-10 kDa smaller than cell-associated PECAM-1 and contains the cytoplasmic tail, was observed in the culture media of HUVECs and phorbol ester-treated U937 cells. This form of soluble PECAM-1 is encoded by an alternatively spliced mRNA from which the exon containing the transmembrane domain has been removed. Soluble PECAM-1 was also detected in normal human plasma at levels of 10-25 ng/ml. Two isoforms of plasma PECAM-1, which differed in the presence of the cytoplasmic tail, were observed by Western blot analysis. In parallel with soluble forms of other cell adhesion molecules, soluble PECAM-1 may play a role in modulating the inflammatory response.