Endotoxin, tumor necrosis factor, and dexamethasone effects on human endothelial cell fibronectin dynamics: synthesis, matrix assembly, and receptor expression

Abstract

The three inflammatory modulators endotoxin, tumor necrosis factor (TNF) α, and dexamethasone (DEX) were studied for their effects on fibronectin (FN) dynamics in human umbilical vein endothelial cells. Cell culture supernatants were analyzed for new soluble pool FN synthesis. Endotoxin (LPS) (10 μg/mL) decreased the newly synthesized soluble pool of FN (p < 0.05). An increase in soluble FN was demonstrated with 1 and 10 ng/mL TNF α (p < 0.05). DEX decreased newly synthesized endothelial cell (EC) FN in the soluble pool at 4, 40, and 400 μg/mL (p < 0.05). Extracellular matrix FN content was examined using immunofluorescence. The thick FN mesh seen in control cells contrasted with a decreased FN matrix after treatment with each of the three study agents. Immunoprecipitation of the FN receptor α5β1integrin from [35S]methionine-labelled cell extracts demonstrated down regulation of receptor expression by both TNF α and DEX as compared with control samples. These data indicate that LPS, TNF α, and DEX may weaken EC–substratum adhesion by differential effects on FN synthesis and secretion, FN incorporation into the extracellular matrix, and down regulation of FN receptor expression.Key words: endothelium, fibronectin, extracellular matrix, integrin, cytokine.