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Novel cytokine-independent induction of endothelial adhesion molecules regulated by platelet/endothelial cell adhesion molecule (CD31).

Litwin M, Clark K, Noack L, Furze J, Berndt M, Albelda S, Vadas M, Gamble J - J. Cell Biol. (1997)

Bottom Line: In contrast, cells plated at low density displayed a 17-fold greater expression of E-selectin than did high density ECs with 57 +/- 4% (n = 5) positive for E-selectin expression 15 h after plating, and significant expression still evident 72 h after plating.In contrast, untransfected L cells or L cells transfected with an adhesion-defective domain 2 deletion PECAM-1 mutant failed to regulate E-selectin expression.In an in vitro model of wounding the wound front displayed an increase in the number of E-selectin-expressing cells, and also an increase in the intensity of expression of E-selectin positive cells compared to the nonwounded monolayer.

View Article: PubMed Central - PubMed

Affiliation: Division of Human Immunology, Hanson Centre for Cancer Research, Adelaide, South Australia.

ABSTRACT
Tumor necrosis factor-alpha, interleukin-1, and endotoxin stimulate the expression of vascular endothelial cell (EC) adhesion molecules. Here we describe a novel pathway of adhesion molecule induction that is independent of exogenous factors, but which is dependent on integrin signaling and cell-cell interactions. Cells plated onto gelatin, fibronectin, collagen or fibrinogen, or anti-integrin antibodies, expressed increased amounts of E-selectin, vascular cell adhesion molecule-1, and intercellular adhesion molecule-1. In contrast, ECs failed to express E-selectin when plated on poly-L-lysine or when plated on fibrinogen in the presence of attachment-inhibiting, cyclic Arg-Gly-Asp peptides. The duration and magnitude of adhesion molecule expression was dependent on EC density. Induction of E-selectin on ECs plated at confluent density was transient and returned to basal levels by 15 h after plating when only 7 +/- 2% (n = 5) of cells were positive. In contrast, cells plated at low density displayed a 17-fold greater expression of E-selectin than did high density ECs with 57 +/- 4% (n = 5) positive for E-selectin expression 15 h after plating, and significant expression still evident 72 h after plating. The confluency-dependent inhibition of expression of E-selectin was at least partly mediated through the cell junctional protein, platelet/endothelial cell adhesion molecule-1 (PECAM-1). Antibodies against PECAM-1, but not against VE-cadherin, increased E-selectin expression on confluent ECs. Co- culture of subconfluent ECs with PECAM-1- coated beads or with L cells transfected with full-length PECAM-1 or with a cytoplasmic truncation PECAM-1 mutant, inhibited E-selectin expression. In contrast, untransfected L cells or L cells transfected with an adhesion-defective domain 2 deletion PECAM-1 mutant failed to regulate E-selectin expression. In an in vitro model of wounding the wound front displayed an increase in the number of E-selectin-expressing cells, and also an increase in the intensity of expression of E-selectin positive cells compared to the nonwounded monolayer. Thus we propose that the EC junction, and in particular, the junctional molecule PECAM-1, is a powerful regulator of endothelial adhesiveness.

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Expression of  E-selectin, VCAM-1, and  ICAM-1 are confluency dependent. ECs plated at subconfluent density (stripes,  0.25 × 105 cells per cm2) and  confluent density (solid, 105  cells per cm2) were stained  20 h later for a, E-selectin, b,  VCAM-1, c, ICAM-1, or d,  PECAM-1. Expression is  shown as the MFI of 23, 11,  7, and 7 EC lines, respectively. Error bars represent  the SEM. Asterisks denote  values significantly different  from cells plated at cobblestone density (P < 0.03) by  paired t test.
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Figure 4: Expression of E-selectin, VCAM-1, and ICAM-1 are confluency dependent. ECs plated at subconfluent density (stripes, 0.25 × 105 cells per cm2) and confluent density (solid, 105 cells per cm2) were stained 20 h later for a, E-selectin, b, VCAM-1, c, ICAM-1, or d, PECAM-1. Expression is shown as the MFI of 23, 11, 7, and 7 EC lines, respectively. Error bars represent the SEM. Asterisks denote values significantly different from cells plated at cobblestone density (P < 0.03) by paired t test.

Mentions: VCAM-1 and ICAM-1 (as well as E-selectin) demonstrated EC density–dependent expression, but the expression of PECAM-1 (or CD31), a non-inducible adhesion protein was not altered (Fig. 4). Increased expression of VCAM-1, ICAM-1, and E-selectin was independent of cell size, as identical forward scatter gates of high and low density cells were always used in the FACS® analysis. Furthermore, the change in adhesion molecule expression was observed, whether the cells were stained in situ before detachment, or after extraction from matrix (data not shown).


Novel cytokine-independent induction of endothelial adhesion molecules regulated by platelet/endothelial cell adhesion molecule (CD31).

Litwin M, Clark K, Noack L, Furze J, Berndt M, Albelda S, Vadas M, Gamble J - J. Cell Biol. (1997)

Expression of  E-selectin, VCAM-1, and  ICAM-1 are confluency dependent. ECs plated at subconfluent density (stripes,  0.25 × 105 cells per cm2) and  confluent density (solid, 105  cells per cm2) were stained  20 h later for a, E-selectin, b,  VCAM-1, c, ICAM-1, or d,  PECAM-1. Expression is  shown as the MFI of 23, 11,  7, and 7 EC lines, respectively. Error bars represent  the SEM. Asterisks denote  values significantly different  from cells plated at cobblestone density (P < 0.03) by  paired t test.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2139821&req=5

Figure 4: Expression of E-selectin, VCAM-1, and ICAM-1 are confluency dependent. ECs plated at subconfluent density (stripes, 0.25 × 105 cells per cm2) and confluent density (solid, 105 cells per cm2) were stained 20 h later for a, E-selectin, b, VCAM-1, c, ICAM-1, or d, PECAM-1. Expression is shown as the MFI of 23, 11, 7, and 7 EC lines, respectively. Error bars represent the SEM. Asterisks denote values significantly different from cells plated at cobblestone density (P < 0.03) by paired t test.
Mentions: VCAM-1 and ICAM-1 (as well as E-selectin) demonstrated EC density–dependent expression, but the expression of PECAM-1 (or CD31), a non-inducible adhesion protein was not altered (Fig. 4). Increased expression of VCAM-1, ICAM-1, and E-selectin was independent of cell size, as identical forward scatter gates of high and low density cells were always used in the FACS® analysis. Furthermore, the change in adhesion molecule expression was observed, whether the cells were stained in situ before detachment, or after extraction from matrix (data not shown).

Bottom Line: In contrast, cells plated at low density displayed a 17-fold greater expression of E-selectin than did high density ECs with 57 +/- 4% (n = 5) positive for E-selectin expression 15 h after plating, and significant expression still evident 72 h after plating.In contrast, untransfected L cells or L cells transfected with an adhesion-defective domain 2 deletion PECAM-1 mutant failed to regulate E-selectin expression.In an in vitro model of wounding the wound front displayed an increase in the number of E-selectin-expressing cells, and also an increase in the intensity of expression of E-selectin positive cells compared to the nonwounded monolayer.

View Article: PubMed Central - PubMed

Affiliation: Division of Human Immunology, Hanson Centre for Cancer Research, Adelaide, South Australia.

ABSTRACT
Tumor necrosis factor-alpha, interleukin-1, and endotoxin stimulate the expression of vascular endothelial cell (EC) adhesion molecules. Here we describe a novel pathway of adhesion molecule induction that is independent of exogenous factors, but which is dependent on integrin signaling and cell-cell interactions. Cells plated onto gelatin, fibronectin, collagen or fibrinogen, or anti-integrin antibodies, expressed increased amounts of E-selectin, vascular cell adhesion molecule-1, and intercellular adhesion molecule-1. In contrast, ECs failed to express E-selectin when plated on poly-L-lysine or when plated on fibrinogen in the presence of attachment-inhibiting, cyclic Arg-Gly-Asp peptides. The duration and magnitude of adhesion molecule expression was dependent on EC density. Induction of E-selectin on ECs plated at confluent density was transient and returned to basal levels by 15 h after plating when only 7 +/- 2% (n = 5) of cells were positive. In contrast, cells plated at low density displayed a 17-fold greater expression of E-selectin than did high density ECs with 57 +/- 4% (n = 5) positive for E-selectin expression 15 h after plating, and significant expression still evident 72 h after plating. The confluency-dependent inhibition of expression of E-selectin was at least partly mediated through the cell junctional protein, platelet/endothelial cell adhesion molecule-1 (PECAM-1). Antibodies against PECAM-1, but not against VE-cadherin, increased E-selectin expression on confluent ECs. Co- culture of subconfluent ECs with PECAM-1- coated beads or with L cells transfected with full-length PECAM-1 or with a cytoplasmic truncation PECAM-1 mutant, inhibited E-selectin expression. In contrast, untransfected L cells or L cells transfected with an adhesion-defective domain 2 deletion PECAM-1 mutant failed to regulate E-selectin expression. In an in vitro model of wounding the wound front displayed an increase in the number of E-selectin-expressing cells, and also an increase in the intensity of expression of E-selectin positive cells compared to the nonwounded monolayer. Thus we propose that the EC junction, and in particular, the junctional molecule PECAM-1, is a powerful regulator of endothelial adhesiveness.

Show MeSH
Related in: MedlinePlus