Biomedical subjects

H C Yan

Publications and source records attributed to H C Yan.

At least 19 recordsLinked to original sources

PubMed2001

Cytokine up-regulation in ischaemic/reperfused lungs perfused with University of Wisconsin solution and normal saline.

Ischaemia/reperfusion (I/R) lung injury using University of Wisconsin solution (UW) as perfusate has not been well studied. Isolated rat lungs were challenged with various periods of ischaemia and/or reperfusion. Haemodynamics, lung weight gain (LWG), capillary filtration coefficient (K(fc)), tissue pathology, the concentrations of cytokines in the perfusate, and mRNAs for the various cytokines in the lung tissues were measured. I/R induced a permeability type of pulmonary oedema, as reflected by increases in LWG and K(fc). LWG and K(fc) in the I(45)R(60)(UW) group (45 min of ischaemia followed by 60 min of reperfusion with UW) were only 2% and 5% respectively of those in the I(45)R(60)(NS) group (where NS is normal saline). LWG and K(fc) in the UW group had both increased by 180 min, to values similar to those in the I(45)R(60)(NS) group. However, these findings show that UW was remarkably effective at preventing LWG after 60 min of reperfusion, and was more than 3-fold more effective than NS in delaying LWG. For longer ischaemic times only, or the same period of ischaemia followed by longer reperfusion periods, greater lung injury occurred. I/R lung injury also induced increased concentrations of tumour necrosis factor-alpha (TNF-alpha), interleukin 1 and interleukin 6 in the perfusate, and increased the mRNAs for these cytokines in lung tissue. A significant correlation was obtained between TNF-alpha concentration and LWG. TNF-alpha production in the I(45)R(60)(UW) group was only 7% of that in the I(45)R(60)(NS) group. However, TNF-alpha mRNA expression in the I(45)R(60)(UW) group was 80% of that in the I(45)R(60)(NS) group. This indicates that transcription/translation do not correlate well with cytokine production, and also suggests that one reason for the effectiveness of UW in delaying LWG may be because it delays TNF-alpha production. In summary, ischaemia or I/R caused a permeability-type pulmonary oedema that was associated with leucocyte infiltration and the up-regulation of various cytokines, regardless of the perfusion fluid. Except for pulmonary hypertension, less severe I/R lung injury and delayed cytokine production in lungs perfused with UW, the pattern of injury associated with I/R challenge was similar to that in lungs perfused with NS. We propose that more or long-acting protective agents are required as additives in order to modify UW to produce an optimal preservation solution.

Adenosine↗

PubMed2000

Dexamethasone and pentastarch produce additive attenuation of ischaemia/reperfusion lung injury.

The choice of an intravenous solution for the attenuation of ischaemia/reperfusion (I/R) lung injury is still a difficult one. Although 10% (w/v) pentastarch has been used in ICU settings, its use in I/R lung injury has not been well explored. We hypothesized that this synthetic colloid substance, which maintains colloid osmotic pressure and potentially 'seals' capillary leaks, in combination with an anti-inflammatory agent (i.e. dexamethasone), would ameliorate I/R lung injury. After 60 min of lung ischaemia in an isolated rat lung model, lungs were reperfused for 60 min in a closed circulating system with one of the following solutions: (1) NS (0.9% normal saline), (2) NS+Dex (dexamethasone), (3) NS+Penta (pentastarch), or (4) NS+Penta+Dex. Haemodynamic changes, lung weight gain (LWG), capillary filtration coefficient (K(fc)) and lung pathology were analysed. Results showed significantly lower values of K(fc) and LWG in pentastarch- or dexamethasone-perfused groups as compared with those in the NS group. Dexamethasone as an additive to NS+Penta further decreased K(fc) and LWG. Histopathological studies showed similar decreases in injury profiles. We conclude that reperfusion with dexamethasone and pentastarch can attenuate I/R lung injury, and that dexamethasone and pentastarch have additive effects. Our data thus suggest that the combination of a colloid substance with 'sealing effects' and an anti-inflammatory agent may provide a better reperfusion solution for patients with I/R lung injury or for lungs stored for transplant.

Animals↗

PubMed2000

Effect of zinc intake on fetal and infant growth among Chinese pregnant and lactating women.

The relationship between maternal nutrient intake and fetal size or infant growth was studied in 1956 pregnant women, 599 parturients and 1043 lactating women, 318 non-pregnant women included as controls. The study was conducted in eight regions that were representative of all geographical areas of China. The diet was comprised primarily of cereal products with 70% to 85% of the zinc intake derived from plant sources. Women in the third trimester of pregnancy, parturients and lactating women consumed more food than non-pregnant women or women in the first two trimesters of pregnancy. Total energy, protein and iron intakes met the recommended allowances for each stage of reproduction. Calcium and zinc intakes, however, were 50% and 47% of the amount recommended, respectively. Only 7.2% of the women exceeded two-thirds of the recommended zinc intake. The mean intake of zinc was 6.5 mg to 9.0 mg each day among all the subjects. Correlation and stepwise regression analysis showed that maternal zinc intake was a predictor factor for fetal dimensions and birthweight. The results of this study show that fetal growth and birthweight are directly related to maternal zinc intake among Chinese women, and that there is no relationship between maternal zinc intake during lactation and infant height, weight, or weight gain from birth.

Adult↗

PubMed2000

Use of anti-(tumour necrosis factor-alpha) antibody or 3-deaza-adenosine as additives to promote protection by University of Wisconsin solution in ischaemia/reperfusion injury.

Experimental interventions that reduce ischaemia/reperfusion (I/R) lung injury can be used to improve the properties of preservation solutions. We attempted to increase the attenuation of I/R injury by University of Wisconsin solution (UW) by adding an antibody against tumour necrosis factor-alpha (TNF-alpha), to neutralize TNF-alpha, and/or by adding 3-deaza-adenosine (c3-Ado), to inhibit leucocyte adhesion and the biosynthesis of ICAM-1 (intercellular cell-adhesion molecule 1). We examined I/R injury using an isolated rat lung model. Six different solutions were perfused individually, followed by evaluation of I/R injury: (1) 0.9% NaCl (normal saline; NS), (2) NS+anti-TNF-alpha antibody, (3) UW alone, (4) UW+anti-TNF-alpha, (5) UW+c3-Ado and (6) UW+anti-TNF-alpha+c3-Ado. Haemodynamic changes, lung weight gain, capillary filtration coefficient, TNF-alpha levels and lung pathology were analysed in order to evaluate I/R injury. Compared with lungs perfused with NS, lungs treated with NS+anti-TNF-alpha showed less I/R injury. The addition of anti-TNF-alpha and/or c3-Ado to UW reduced I/R injury compared with unmodified UW. Among the six solutions tested, UW containing anti-TNF-alpha antibody reduced I/R injury to the greatest extent. We conclude that addition of anti-TNF-alpha antibody or c3-Ado protects against I/R lung injury when using UW. Further investigation of the improved properties of modified UWs would be beneficial with regard to lung transplantation research.

Adenosine↗

PubMed1999

Hypothermia and prostaglandin E(1) produce synergistic attenuation of ischemia-reperfusion lung injury.

Current methods of preserving lung tissue for transplantation are inadequate. In this study, we tested whether the combination of hypothermia plus prostaglandin E(1) (PGE(1)) treatment would have synergistic attenuation on ischemia-reperfusion (I/R) lung injury. Isolated rat lung experiments with ischemia for 1 h then reperfusion for 1 h, were conducted using six different perfusates: (1) University of Wisconsin solution (UW) at 30 degrees C (n = 5), (2) UW at 22 degrees C (n = 5), (3) UW at 10 degrees C (n = 4), (4) UW+PGE(1) at 30 degrees C (n = 4), (5) UW+PGE(1) at 22 degrees C (n = 4), and (6) UW+PGE(1) at 10 degrees C (n = 4). Hemodynamic changes, lung weight gain, capillary filtration coefficients, and lung pathology were analyzed to evaluate the I/R injury. Compared with 30 degrees C UW, animals treated with 22 degrees C UW and 10 degrees C UW had less I/R lung injury, with the groups receiving 22 degrees C UW showing superior results to group receiving 10 degrees C UW. The addition of PGE(1) to UW solution produced more attenuation of I/R injury than did UW alone. Among the six groups, 10 degrees C UW+PGE(1) produced the most reduction of I/R injury. This study has shown that hypothermia can attenuate I/R injury with the optimal flushing temperature being near 22 degrees C. PGE(1) also has a protective effect on I/R. Furthermore, hypothermia and PGE(1) have synergistic attenuation of I/R lung injury. We propose that pulmonary artery flushed with cooling UW+PGE(1) might improve lung preservation and improve results in lung transplantation.

Adenosine↗

PubMed1998

Protective agents used as additives in University of Wisconsin solution to promote protection against ischaemia-reperfusion injury in rat lung.

1. An intervention to reduce ischaemia-reperfusion lung injury will be an important advance in transplant medicine. Although the mechanisms associated with producing ischaemia-reperfusion endothelial injury have not been completely elucidated, many of the injury mediators have been studied in detail. While no single pharmacological therapy is likely to be totally effective in eliminating this complex injury, we have developed a mixture of agents that are known to block pathways involved in producing ischaemia-reperfusion-associated lung vascular injury.2. The present study modified University of Wisconsin solution (UW) by adding one of the protective agents prostaglandin E1 (PGE1), dexamethasone (Dex) or dibutyryl cAMP (Bt2-cAMP), or a combination of these, to the perfusate of rat lungs exposed to 4 h of cold ischaemia followed by 1 h of reperfusion. Nine modified UW solutions were studied: (1) UW+Dex, (2) UW+PGE1, (3) UW+Bt2-cAMP, (4) UW+Dexx3, (5) UW+PGE1x3, (6) UW+Bt2-cAMPx3, (7) UW+Dex+PGE1, (8) UW+Dex+Bt2-cAMP, (9) UW+PGE1+Bt2-cAMP. These solutions were utilized in individual experiments to assess haemodynamic changes, lung weight gain, the capillary filtration coefficient (Kfc) and pathology in all lungs.3. The results indicate that lung weight gain and Kfc values were significantly lower than with UW alone in groups 1, 2 and 3, which contained only one additional protective agent. In groups 4, 5 and 6, which contain three times the concentration of each protective agent, both Kfc and lung weight gain were similar to those measured in groups 1, 2 and 3, i.e. lungs were protected but the protection was not dose dependent. In groups 7, 8 and 9, which contained two protective agents, lung weight gain and Kfc were greatly reduced compared with UW alone. Histopathological studies showed similar decreases in the injury profiles of lungs.4. Although UW contains several antioxidant protective agents such as allopurinol and glutathione, it did not provide effective protection in our ischaemia-reperfusion lung injury model. UW modified with an additive of PGE1, Dex or Bt2-cAMP attenuated ischaemia-reperfusion injury. Furthermore, UW containing two of these protective agents augmented the protection. Among the modified solutions, it appears that UW+PGE1+Bt2-cAMP protects the lungs to a greater extent than all other solutions used in our study. We suggest that preservation solutions containing PGE1-Bt2-cAMP will provide additional protective effects to organs stored for transplantation.

Adenosine↗

PubMed1997

PGE1, dexamethasone, U-74389G, or Bt2-cAMP as an additive to promote protection by UW solution in I/R injury.

A method to reduce ischemia-reperfusion (I/R) injury can be an important criterion to improve the preservation solution. Although University of Wisconsin solution (UW) works as a lung preservation solution, its attenuation effect on I/R injury has not been investigated. We attempted to determine whether, by adding various protective agents, modified UW solutions will enhance the I/R attenuation by UW. We examined the I/R injury in an isolated rat lung model. Various solutions, e.g., physiological salt solution (PSS), UW, and modified UW solutions containing various protective agents such as prostaglandin E1, dexamethasone, U-74389G, or dibutyryl adenosine 3',5'-cyclic monophosphate were perfused individually to evaluate the I/R injury. Isolated rat lung experiments, with ischemia for 45 min, then reperfusion for 60 min, were conducted in a closed circulating system. Hemodynamic changes, lung weight gain (LWG), capillary filtration coefficient (Kfc), protein content of lavage fluid, concentration of cytokines, and lung histopathology were analyzed. Results showed that the acute I/R lung injury with immediate permeability pulmonary edema was associated with an increase in tumor necrosis factor-alpha (TNF-alpha) production. A significant correlation existed between TNF-alpha and Kfc (r = 0.8, P < 0.0001) and TNF-alpha and LWG (r = 0. 9, P < 0.0001), indicating that TNF-alpha is an important cytokine modulating early I/R injury. Significantly lower levels of Kfc, LWG, TNF-alpha, and protein concentration of lung lavage (P < 0.05) were found in the UW-perfused group than in the control group perfused with PSS. Modified UW promoted the protective effect of UW to further decrease Kfc, LWG, and TNF-alpha (P < 0.05). Histopathological observations also substantiated this evidence. In the UW+U-74389G group, bronchial alveolar lavage fluid contained lowest protein concentration. We conclude that the UW solution attenuates I/R injury of rat lung and that the modified UW solutions further enhance the effect of UW in reducing I/R injury. Among modified solutions, UW+U-74389G is the best. Further investigation of the improved effects of the modified UW solutions would be beneficial in lung transplantation.

Adenosine↗

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

The role of selectins and CD18 in leukotriene B4-mediated white blood cell emigration in human skin grafts transplanted on SCID mice.

The purpose of this study was to examine the role of selectins and CD18 cell adhesion molecules (CAMs) in inflammation induced by injection of leukotriene B4 (LTB4) into human skin. To accomplish this, the expression of CAMs and the ability of specific antibodies against CAMs to block white blood cell (WBC) transmigration were studied in an in vivo model consisting of human skin transplanted onto mice with the severe combined immune deficiency (SCID) mutation. The results indicate that LTB4-induced WBC transmigration in the human/SCID model is rapid and pronounced; however, it is not accompanied by a significant upregulation of the baseline expression of endothelial P-selectin, E-selectin, ICAM-1 or VCAM-1. An anti-murine CD18 mAb markedly inhibited white cell infiltration (89% inhibition) confirming the importance of beta 2 integrins in the process. The role of selectins was also examined. MEL-14, a bioactive antibody against murine L-selectin inhibited transmigration by 66%. A significant, but smaller, effect (39% inhibition) was observed by blocking E-selectin function. These results indicate that LTB4-induced inflammation does not require upregulation of endothelial CAM expression and, in contrast to TNF alpha-induced transmigration, is only partially blocked by anti-E-selectin antibodies.

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

Pentoxifylline attenuates acute lung injury induced by microemboli.

Pentoxifylline (PTX), a methylxanthine derivative, effectively prevents acute lung injury in different animal models. To investigate whether PTX would attenuate acute lung injury induced by microemboli resulting from treatment with calcium chloride (CaCl2) suspension, an isolated blood-perfused rat lung model was used. Pretreatment with PTX prevented the increase in pulmonary arterial pressure (PAP), lung weight gain and protein concentration in the lavage fluid after CaCl2 treatment.

Acute Disease↗

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

Modulation of adhesion molecules by cytokines in vivo using human/severe combined immunodeficient (SCID) mouse chimeras.

Endothelial cell-leukocyte interactions involve multiple cell adhesion molecules acting in a programmed and sequential manner to create a leukocyte-endothelial cell adhesion cascade. To understand this process fully, in vivo models are needed. To accomplish this, we have transplanted pieces of normal human tissues onto immunodeficient mice to create chimeric animals. In one model, human skin is grafted and closely resembles normal skin histologically. The grafts retain their human vasculature and show low baseline expression of E-selectin, vascular cell adhesion molecule-1, and intercellular cell adhesion molecule-1. After intradermal injection of human cytokines, these cell adhesion molecules are markedly upregulated and an active inflammatory reaction ensues, with migration of murine leukocytes. Intravenous injection of an anti-human E-selectin antibody completely inhibits leukocyte accumulation induced by tumor necrosis factor-alpha but only partially inhibits leukotriene B4-induced inflammation. In a second model, human bronchus was successfully transplanted heterotopically into severe combined immunodeficient mice. Injection of tumor necrosis factor induced upregulation of E-selectin, intercellular cell adhesion molecule-1, and vascular cell adhesion molecule-1 in the submucosal microvessels, with slightly different kinetics than in the skin. In conclusion, human-severe combined immunodeficient chimeric mice represent a useful model system to study the regulation and function of human cell adhesion molecules in an in vivo setting.

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↗

PubMed1994

Leukocyte recruitment into human skin transplanted onto severe combined immunodeficient mice induced by TNF-alpha is dependent on E-selectin.

In order to study the in vivo role of E-selectin in human inflammation, we have developed a model in which human skin is transplanted onto severe combined immunodeficient (SCID) mice. The grafted skin closely resembles normal skin and retains its human vasculature. After intradermal injection of rTNF-alpha, human E-selectin was rapidly up-regulated on dermal microvessels, with significant expression (determined immunohistochemically) at 1 h postinjection and maximum expression at 2 h postinjection. To study the functional role of E-selectin, a murine Ab against human E-selectin (mAb HEL 3/2) was developed that inhibited the in vitro adhesion of both human U937 cells and murine 32D cells to TNF-alpha-stimulated human endothelial cells. After intradermal injection of TNF-alpha, large numbers of murine leukocytes migrated into the grafts within 2 h. Intravenous injection of the antihuman E-selectin mAb 3/2 completely inhibited murine white blood cell (WBC) transmigration into the skin grafts, but an isotype-matched control Ab that also bound to human endothelium had no effect. Antihuman E-selectin mAb 3/2 was also able to inhibit the migration of i.v. 51Cr-labeled human neutrophils. These findings demonstrate that E-selectin is important in early white blood cell adhesion events and is required for TNF-alpha-induced white blood cell transmigration in the human/SCID mouse chimeric model.

Animals↗

PubMed1994

Deletions in the cytoplasmic domain of platelet-endothelial cell adhesion molecule-1 (PECAM-1, CD31) result in changes in ligand binding properties.

Platelet/endothelial cell adhesion molecule-1 (PECAM-1, CD31) is a member of the immunoglobulin superfamily present on platelets, endothelial cells, and leukocytes that may function as a vascular cell adhesion molecule. The purpose of this study was to examine the role of the cytoplasmic domain in PECAM-1 function. To accomplish this, wild-type and mutated forms of PECAM-1 cDNA were transfected into murine fibroblasts and the functional characteristics of the cells analyzed. Wild-type PECAM-1 localized to the cell-cell borders of adjacently transfected cells and mediated heterophilic, calcium-dependent L-cell aggregation that was inhibitable by a polyclonal and two monoclonal anti-PECAM-1 antibodies. A mutant protein lacking the entire cytoplasmic domain did not support aggregation or move to cell-cell borders. In contrast, both forms of PECAM-1 with partially truncated cytoplasmic domains (missing either the COOH-terminal third or two thirds of the cytoplasmic domain) localized to cell-cell borders in 3T3 cells in a manner analogous to the distribution seen in cultured endothelial cells. L-cells expressing these mutants demonstrated homophilic, calcium-independent aggregation that was blocked by the polyclonal anti-PECAM-1 antibody, but not by the two bioactive monoclonal antibodies. Although changes in the cytoplasmic domain of other receptors have been shown to alter ligand-binding affinity, to our knowledge, PECAM-1 is the first example of a cell adhesion molecule where changes in the cytoplasmic domain result in a switch in the basic mechanism of adhesion leading to different ligand-binding specificity. Variations in the cytoplasmic domain could thus be a potential mechanism for regulating PECAM-1 activity in vivo.

3T3 Cells↗

PubMed1994

Platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31): alternatively spliced, functionally distinct isoforms expressed during mammalian cardiovascular development.

The establishment of the cardiovascular system represents an early, critical event essential for normal embryonic development. An important component of vascular ontogeny is the differentiation and development of the endothelial and endocardial cell populations. This involves, at least in part, the expression and function of specific cell surface receptors required to mediate cell-cell and cell-matrix adhesion. Platelet endothelial cell adhesion molecule-1 (PECAM-1, CD31) may well serve such a function. It is a member of the immunoglobulin superfamily expressed by the entire vascular endothelium in the adult. It is capable of mediating adhesion by a heterophilic mechanism requiring glycosaminoglycans, as well as by a homophilic, glycosaminoglycan independent, mechanism. It has been shown to regulate the expression of other adhesion molecules on naive T cells. This report documents by RT-PCR and immunohistochemical analysis the expression of PECAM-1 during early post implantation mouse embryo development. PECAM-1 was expressed by early endothelial precursors first within the yolk sac and subsequently within the embryo itself. Interestingly, embryonic PECAM-1 was expressed as multiple isoforms in which one or more clusters of polypeptides were missing from the cytoplasmic domain. The sequence and location of the deleted polypeptides corresponded to exons found in the human PECAM-1 gene. The alternatively spliced isoforms were capable of mediating cell-cell adhesion when transfected into L-cells. The isoforms differed, however, in their sensitivity to a panel of anti-PECAM-1 monoclonal antibodies. These data suggest that changes in the cytoplasmic domain of PECAM-1 may affect its function during cardiovascular development, and are consistent with our earlier report that systematic truncation of the cytoplasmic domain of human PECAM-1 resulted in changes in its ligand specificity, divalent cation and glycosaminoglycan dependence, as well as its susceptibility to adhesion blocking monoclonal antibodies. This is the first report of naturally occurring alternatively spliced forms of PECAM-1 having possible functional implications.

Amino Acid Sequence↗