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H M DeLisser

Publications and source records attributed to H M DeLisser.

At least 19 recordsLinked to original sources
PubMed2000

Antibodies against the first Ig-like domain of human platelet endothelial cell adhesion molecule-1 (PECAM-1) that inhibit PECAM-1-dependent homophilic adhesion block in vivo neutrophil recruitment.

Platelet endothelial cell adhesion molecule (PECAM-1), a member of the Ig superfamily, is found on endothelial cells and neutrophils and has been shown to be involved in the migration of leukocytes across the endothelium. Adhesion is mediated, at least in part, through binding interactions involving its first N-terminal Ig-like domain, but it is still unclear which sequences in this domain are required for in vivo function. Therefore, to identify functionally important regions of the first Ig-like domain of PECAM-1 that are required for the participation of PECAM-1 in in vivo neutrophil recruitment, a panel of mAbs against this region of PECAM-1 was generated and characterized in in vitro adhesion assays and in an in vivo model of cutaneous inflammation. It was observed that mAbs that disrupted PECAM-1-dependent homophilic adhesion in an L cell aggregation assay also blocked TNF-alpha-induced intradermal accumulation of neutrophils in a transmigration model using human skin transplanted onto SCID mice. Localization of the epitopes of these Abs indicated that these function-blocking Abs mapped to specific regions on either face of domain 1. This suggests that these regions of the first Ig-like domain may contain or be close to binding sites involved in PECAM-1-dependent homophilic adhesion, and thus may represent potential targets for the development of antiinflammatory reagents.

Amino Acid Sequence↗
PubMed2000

Bleomycin-induced pulmonary injury in mice deficient in SPARC.

SPARC (secreted protein, acidic and rich in cysteine) is a component of the matrix that appears to regulate tissue remodeling. There is evidence that it accumulates in the lung in the setting of pulmonary injury and fibrosis, but direct evidence of its involvement is only now emerging. We therefore investigated the development of pulmonary fibrosis induced by bleomycin administered either intratracheally or intraperitoneally in mice deficient in SPARC. Bleomycin (0.15 U/mouse) given intratracheally induced significantly more pulmonary fibrosis in mice deficient in SPARC compared with that in wild-type control mice, with the mutant mice demonstrating greater neutrophil accumulation in the lung. However, in wild-type and SPARC-deficient mice given intraperitoneal bleomycin (0.8 U/injection x 5 injections over 14 days), the pattern and severity of pulmonary fibrosis, as well as the levels of leukocyte recruitment, were similar in both strains of mice. These findings suggest that the involvement of SPARC in pulmonary injury is likely to be complex, dependent on several factors including the type, duration, and intensity of the insult. Furthermore, increased neutrophil accumulation in the peritoneal cavity was also observed in SPARC-null mice after acute chemical peritonitis. Together, these data suggest a possible role for SPARC in the recruitment of neutrophils to sites of acute inflammation.

Animals↗
PubMed2000

A role for hyaluronan in macrophage accumulation and collagen deposition after bleomycin-induced lung injury.

Elevated concentrations of hyaluronan (HA) are associated with the accumulation of macrophages in the lung after injury. We have investigated the role of HA in the inflammatory and fibrotic responses to lung injury using the intratracheal instillation of bleomycin in rats as a model. After bleomycin-induced lung injury, both HA content in bronchoalveolar lavage (BAL) and staining for HA in macrophages accumulating in injured areas of the lung were maximal at 4 d. Increased HA in BAL correlated with increased locomotion of isolated alveolar macrophages. HA-binding peptide was able to specifically block macrophage motility in vitro. Importantly, systemic administration of HA-binding peptide to rats before injury not only decreased alveolar macrophage motility and accumulation in the lung, but also reduced lung collagen alpha (I) messenger RNA and hydroxyproline contents. We propose a model in which HA plays a critical role in the inflammatory response and fibrotic consequences of acute lung injury.

Amino Acid Sequence↗
PubMed1999

Loss of endothelial surface expression of E-selectin in a patient with recurrent infections.

Neutrophil accumulation at sites of inflammation is mediated by specific groups of cell adhesion molecules including the beta2 (CD18) integrins on leukocytes and the selectins (P- and E-selectin on the endothelium and L-selectin on the leukocyte). This is supported by studies of patients with leukocyte adhesion deficiency syndromes whose leukocytes are genetically deficient in the expression of beta2 integrins or selectin carbohydrate ligands (eg, sialyl-Lewis(x)). However, inherited deficiency or dysfunction of endothelial cell adhesion molecules involved in leukocyte recruitment has not been previously described. In this report we describe a child with recurrent infections and clinical evidence of impaired pus formation reminiscent of a leukocyte adhesion deficiency syndrome, but whose neutrophils were functionally normal and expressed normal levels of CD18, L-selectin, and sialyl-Lewis(x). In contrast, immunohistochemical staining of inflamed tissue from the patient showed the absence of E-selectin from the endothelium, although E-selectin mRNA was present. However, E-selectin protein was expressed as significantly elevated levels of circulating soluble E-selectin were detected, the molecular size of which was consistent with a proteolytically cleaved form of E-selectin. Gene sequencing failed to show evidence of a secreted mutant variant. These data represent, to our knowledge, the first description of a potentially inherited dysfunction of an endothelial cell adhesion molecule involved in leukocyte recruitment and provide additional human evidence of the importance of endothelial selectins in the inflammatory response.

Bacterial Infections↗
PubMed1999

Epitope mapping.

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Animals↗
PubMed1997

Tyrosine residue in exon 14 of the cytoplasmic domain of platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31) regulates ligand binding specificity.

Platelet/endothelial cell adhesion molecule (PECAM-1) is a cell adhesion molecule of the immunoglobulin superfamily that plays a role in a number of vascular processes including leukocyte transmigration through endothelium. The presence of a specific 19- amino acid exon within the cytoplasmic domain of PECAM-1 regulates the binding specificity of the molecule; specifically, isoforms containing exon 14 mediate heterophilic cell-cell aggregation while those variants missing exon 14 mediate homophilic cell-cell aggregation. To more precisely identify the region of exon 14 responsible for ligand specificity, a series of deletion mutants were created in which smaller regions of exon 14 were removed. After transfection into L cells, they were tested for their ability to mediate aggregation. For heterophilic aggregation to occur, a conserved 5-amino acid region (VYSEI in the murine sequence or VYSEV in the human sequence) in the mid-portion of the exon was required. A final construct, in which this tyrosine was mutated into a phenylalanine, aggregated in a homophilic manner when transfected into L cells. Inhibition of phosphatase activity by exposure of cells expressing wild type or mutant forms of PECAM-1 to sodium orthovanadate resulted in high levels of cytoplasmic tyrosine phosphorylation and led to a switch from heterophilic to homophilic aggregation. Our data thus indicate either loss of this tyrosine from exon 14 or its phosphorylation results in a change in ligand specificity from heterophilic to homophilic binding. Vascular cells could thus determine whether PECAM-1 functions as a heterophilic or homophilic adhesion molecule by processes such as alternative splicing or by regulation of the balance between tyrosine phosphorylation or dephosphorylation. Defining the conditions under which these changes occur will be important in understanding the biology of PECAM-1 in transmigration, angiogenesis, development, and other processes in which this molecule plays a role.

Alternative Splicing↗
PubMed1997

Neutrophil platelet endothelial cell adhesion molecule-1 participates in neutrophil recruitment at inflammatory sites and is down-regulated after leukocyte extravasation.

Platelet endothelial cell adhesion molecule-1 (PECAM-1), a member of the Ig superfamily, is found on endothelial cells and neutrophils, and has been shown to be involved in the migration of leukocytes across the endothelium. Although studies have supported a role for endothelial PECAM-1 in this process, the participation of neutrophil PECAM-1 in vivo has not been unambiguously demonstrated. Therefore, to examine the involvement of neutrophil PECAM-1 in leukocyte recruitment, we studied the effect of a blocking Ab against murine PECAM-1 on neutrophil recruitment in an established model of murine peritonitis and in a murine model for studying leukocyte-endothelial interactions involving the human vasculature. These studies not only confirmed that neutrophil PECAM-1 is important in the accumulation of neutrophils at inflammatory sites, but that extravasated neutrophils displayed decreased surface expression of PECAM-1. In vitro, the surface expression of murine neutrophil PECAM-1 was not decreased significantly by inflammatory mediators, but was reduced after transendothelial migration. These studies, consistent with previous in vitro observations, confirm that neutrophil PECAM-1, as well as endothelial PECAM-1, is involved in the recruitment of neutrophils into inflammatory sites in vivo, and suggest that the expression of neutrophil PECAM-1 is down-regulated after extravasation into inflamed tissues possibly as a result of engagement of its ligand.

Animals↗
PubMed1997

Involvement of endothelial PECAM-1/CD31 in angiogenesis.

The adhesive interactions of endothelial cells with each other and the adhesion receptors that mediate these interactions are probably of fundamental importance to the process of angiogenesis. We therefore studied the effect of inhibiting the function of the endothelial cell-cell adhesion molecule, PECAM-1/ CD31, in rat and murine models of angiogenesis. A polyclonal antibody to human PECAM-1, which cross-reacts with rat PECAM-1, was found to block in vitro tube formation by rat capillary endothelial cells and cytokine-induced rat corneal neovascularization. In mice, two monoclonal antibodies against murine PECAM-1 prevented vessel growth into subcutaneously implanted gels supplemented with basic fibroblast growth factor (bFGF). Taken together these findings provide evidence that PECAM-1 is involved in angiogenesis and suggest that the interactions of endothelial cell-cell adhesion molecules are important in the formation of new vessels.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed1997

Expression and function of endothelial cell alpha v integrin receptors in wound-induced human angiogenesis in human skin/SCID mice chimeras.

Accumulating evidence indicates that endothelial cell integrins that bind to the matrix proteins associated with inflammation and wound healing are involved in the process of angiogenesis. The integrins containing the alpha v subunit appear to be particularly important. To study the involvement of these receptors in human angiogenesis, a model of wound-associated human angiogenesis was established in human skin transplanted onto severe combined immunodeficient (SCID) mice. Using this model, we studied the expression of several alpha v integrins and tested the hypothesis that blockage of the alpha v beta 3 integrin would inhibit human angiogenesis during human wound healing. These studies revealed that the alpha v beta 3, alpha v beta 5, and alpha v beta 6 integrins are up-regulated briefly during wound angiogenesis with different patterns of expression and that inhibition of the alpha v beta 3 integrin blocked new vessel formation during human wound healing.

Animals↗
PubMed1996

Platelet endothelial cell adhesion molecule-1 (PECAM-1) homophilic adhesion is mediated by immunoglobulin-like domains 1 and 2 and depends on the cytoplasmic domain and the level of surface expression.

PECAM-1/CD31 is vascular cell adhesion and signaling molecule of the Ig superfamily that plays a role in neutrophil recruitment at inflammatory sites and may be involved the release of leukocytes from the bone marrow and in cardiovascular development. The interactions of PECAM-1 with its ligands are complex in that it is able to bind both with itself (homophilic adhesion) or with non-PECAM-1 ligands (heterophilic adhesion). Although the factors that regulate ligand binding are not fully understood, these interactions are regulated in part by its large cytoplasmic domain, a region of 118 amino acids encoded by 8 exons of its gene (exons 9-16). The purpose of this work was to better define the mechanisms of PECAM-1-dependent homophilic adhesion by analyzing the binding interactions of L-cells expressing full-length and selectively mutated forms of human, murine, and human/murine chimeric PECAM-1 molecules in an established aggregation assay. These studies demonstrate that 1) the minimal length of the cytoplasmic domain required for cellular aggregation is represented within the sequences encoded by exons 9 and 10, 2) removal or addition of the sequences encoded by exon 14 from the cytoplasmic domain can determine whether the mechanism of aggregation is a heterophilic calcium-dependent process or a homophilic calcium-independent process, 3) high levels of surface expression of PECAM-1 on the cell surface change the mechanism of aggregation from heterophilic to homophilic, and 4) PECAM-1-dependent homophilic binding appears to involve the direct interaction of only the first two extracellular Ig-like domains. These data suggest that PECAM-1-ligand interactions can be regulated through multiple pathways including alterations of the cytoplasmic domain and the level of surface expression.

Animals↗
PubMed1996

Individually distinct Ig homology domains in PECAM-1 regulate homophilic binding and modulate receptor affinity.

PECAM-1 (CD31) is a 130-kDa member of the immunoglobulin (Ig) gene superfamily that is constitutively expressed at high concentration at endothelial cell intercellular junctions and at moderate density on the surface of circulating leukocytes and platelets. Recent in vitro and in vivo studies have shown the PECAM-1 plays a central role in mediating the extravasation of leukocytes from the vessel wall in response to inflammatory mediators. To study the binding characteristics of PECAM-1, phospholipid vesicles were prepared and examined by flow cytometry and immunofluorescence microscopy for their ability to associate with each other and with cells. Proteoliposomes containing high concentrations of PECAM-1 interacted homophilically with each other, forming large self-aggregates. PECAM-1 proteoliposomes, as well as soluble bivalent PECAM-1 in the form of a PECAM-1/IgG immunoadhesin, associated homophilically with cells expressing human, but not murine, PECAM-1. This binding could be completely inhibited by monoclonal antibody Fab fragments specific for Ig homology Domain 1 or Domains 1 + 2. Binding studies using cells expressing human PECAM-1 deletion mutants and murine/human chimeras confirmed that both Ig Domains 1 and 2 were both necessary and sufficient for homophilic binding. In contrast, engagement of membrane-proximal Domain 6 with monoclonal antibody Fab fragments had the opposite effect and augmented the binding of PECAM-1 proteoliposomes to cells. Thus, PECAM-1, like certain integrins, appears to be capable of antibody-induced conformational changes that alter affinity for its ligand. Similar changes induced by physiologic stimuli could be important in regulating the function of PECAM-1 in vascular cells.

Animals↗
PubMed1995

Alternative splicing of a specific cytoplasmic exon alters the binding characteristics of murine platelet/endothelial cell adhesion molecule-1 (PECAM-1).

Platelet/endothelial cell adhesion molecule-1 (PECAM-1, CD31) is a membrane glycoprotein expressed on endothelial cells, platelets, and leukocytes. Analysis of PECAM-1 expression in the developing mouse embryo has revealed the presence of multiple isoforms of murine PECAM-1 (muPECAM-1) that appeared to result from the alternative splicing of exons encoding cytoplasmic domain sequences (exons 10-16) (Baldwin, H. S., Shen, H. M., Yan, H., DeLisser, H. M., Chung, A., Mickanin, C., Trask, T., Kirschbaum, N. E. Newman, P. J., Albelda, S., and Buck, C. A. (1994) Development 120, 2539-2553). To investigate the functional consequences of alternatively spliced muPECAM-1 cytoplasmic domains, L-cells were transfected with cDNA for each variant and their ability to promote cell aggregation was compared. In this assay, full-length muPECAM-1 and all three isoforms containing exon 14 behaved like human PECAM-1 in that they mediated calcium- and heparin-dependent heterophilic aggregation. In contrast, three muPECAM-1 variants, all missing exon 14, mediated calcium- and heparin-independent homophilic aggregation. Exon 14 thus appears to modulate the ligand and adhesive interactions of the extracellular domain of PECAM-1. These findings suggest that alternative splicing may represent a mode of regulating the adhesive function of PECAM-1 in vivo and provides direct evidence that alternative splicing involving the cytoplasmic domain affects the ligand specificity and binding properties of a cell adhesion receptor.

Alternative Splicing↗
PubMed1995

IFN-gamma and TNF-alpha induce redistribution of PECAM-1 (CD31) on human endothelial cells.

Platelet endothelial adhesion molecule-1 (PECAM-1/CD31) is a glycoprotein adhesion molecule of the Ig superfamily that is constitutively expressed on leukocytes, platelets, and endothelial cells where it concentrates at the intercellular borders of adjacent cells. Recent studies have confirmed that endothelial PECAM-1 is involved in the recruitment of neutrophils into inflammatory sites. However, the effects of inflammatory cytokines such as TNF-alpha and IFN-gamma on endothelial PECAM-1 expression are not known. We studied the effect of several inflammatory cytokines on human umbilical vein endothelial cell PECAM-1 expression. We found that TNF-alpha and IFN-gamma produced dose-dependent changes in the surface distribution of PECAM-1, with a loss of the typical staining of PECAM-1 at intercellular junctions. Because TNF-alpha and IFN-gamma did not alter PECAM-1 transcription or total surface PECAM-1, these changes in PECAM-1 localization are most consistent with a redistribution of the protein away from intercellular junctions. This cytokine-induced surface redistribution of PECAM-1 was associated with changes in PECAM-1 cytoskeletal association but did not involve the expression of different alternatively spliced variants of the molecule. Given the involvement of endothelial PECAM-1 in neutrophil recruitment, redistribution of PECAM-1 may serve as a mechanism for regulating the transmigration of leukocytes across the vascular endothelium.

Alternative Splicing↗
PubMed1995

Expression of the beta 6 integrin subunit in development, neoplasia and tissue repair suggests a role in epithelial remodeling.

The alpha v beta 6 integrin was identified in cultured epithelial cells and functions as a fibronectin receptor. We have now used monoclonal antibodies to determine in vivo expression patterns of the beta 6 subunit in normal and pathological human or primate tissues, and during experimental wound healing or induced lung injury. The results indicate that beta 6 expression is restricted to epithelia and is up-regulated in parallel with morphogenetic events, tumorigenesis, and epithelial repair. During development of the kidney, lung, and skin, we found that beta 6 is expressed by specific types of epithelial cells, whereas it is mostly undetectable in normal adult kidney, lung and skin. In contrast, we detected high-level expression in several types of carcinoma. For example, beta 6 is almost invariably neo-expressed in squamous cell carcinomas derived from the oral mucosa, often focally localized at the infiltrating edges of tumor islands. Expression of beta 6 is also upregulated in migrating keratinocytes at the wound edge during experimental epidermal wound healing. Similarly, beta 6 expression is induced in type II alveolar epithelial cells during lung injury caused by injection of live bacteria. We also observed beta 6 expression in adult lungs and kidneys at focal sites of subclinical inflammation, as well as in a variety of clinical specimens from patients with chronic or acute inflammation of the lungs or kidneys. From these findings and earlier results, we hypothesize that alpha v beta 6 affects cell spreading, migration and growth during reorganization of epithelia in development, tissue repair, and neoplasia.

Animals↗
PubMed1995

Localization of multiple functional domains on human PECAM-1 (CD31) by monoclonal antibody epitope mapping.

PECAM-1, a cell adhesion molecule of the immunoglobulin gene (Ig) superfamily, has been implicated in white cell transmigration, integrin activation on lymphocytes, and cell-cell adhesion. The purpose of this investigation was to identify specific regions of the PECAM-1 extracellular domain mediating these functions by identifying the location of epitopes of bioactive anti-PECAM-1 monoclonal antibodies. The binding regions of mAbs important in PECAM-1-mediated leukocyte transmigration (Hec 7.2 and 3D2) were mapped to N-terminal Ig-like domains. The epitopes of monoclonal antibodies that activated integrin function on lymphocytes were dispersed over the entire extracellular region, but those that had the strongest activating effect were preferentially localized to the N-terminus of the molecule. The binding regions of mAbs that blocked PECAM-1-mediated heterophilic L-cell aggregation were located either in Ig-like domain 2 (NIH31.4) or Ig-like domain 6 (4G6 and 1.2). Site-directed mutagenesis further pinpointed the epitope of the 4G6 mAb to a hexapeptide, CAVNEG, within Ig-like domain 6. These results demonstrate that PECAM-1 contains multiple functional domains. Regions within N-terminal Ig-like domains appear to be required for transmigration. In contrast, two distinct regions were implicated in L-cell mediated heterophilic aggregation.

Amino Acid Sequence↗
PubMed1994

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.

Alternative Splicing↗