FIBRONECTIN ATTENUATES INCREASED ENDOTHELIAL MONOLAYER PERMEABILITY AFTER RGD PEPTIDE, ANTI-ALPHA(5)BETA(1), OR TNF-ALPHA EXPOSURE

Endothelial permeability can be altered by tumor necrosis factor-alpha (TNF-alpha), a cytokine released in association with inflammation-ind uced tissue injury. In the subendothelial matrix, fibronectin (Fn) inf luences endothelial cell adhesion by the interaction of integrins with RGD and non-RGD attachment sites in Fn. We compared the effect of TNF -alpha, RGD-containing peptides (GRGDSP), or antibody to alpha(5) beta (1)-integrins on the protein permeability of bovine lung endothelial m onolayers as assessed by transendothelial I-125-labeled albumin cleara nce. We also examined the influence of purified human plasma fibronect in (hFn) on this permeability response. TNF-alpha, RGD peptides, and a ntibodies to alpha(5) beta(1)-integrins elicited a dose- and time-depe ndent increase in protein permeability as well as a reorganization and /or disruption of the endogenous Fn matrix. A control RGE peptide (GRG ESP) as well as immunoglobulin G purified from nonimmune rabbit serum did not increase endothelial protein permeability or disrupt the endog enous fibrillar Fn pattern in the matrix. Likewise, a LDV peptide deri ved from the alternatively spliced type III connecting segment (IIICS) within bovine Fn (bFn) was unable to increase permeability of the bov ine endothelial monolayer. Co-incubation of purified soluble hFn (300 or 600 mu g/ml) with either TNF-alpha, the RGD peptide, or the antibod y to alpha(5) beta(1)-integrins prevented the increase in endothelial permeability. This protective effect was also observed when the purifi ed hFn (600 mu g/ml) was added after the TNF-alpha-induced increase in endothelial permeability had taken place. Immunofluorescent analysis confirmed the incorporation of the hFn into the subendothelial matrix and its co-localization with the endogenous bFn. The similar alteratio n of the subendothelial matrix after exposure to RGD peptides, anti-al pha(5) beta(1) antibodies, or TNF-alpha, coupled with the ability for hFn to attenuate the permeability increase typically elicited by all t hree agents, suggests that disruption of cell-matrix interactions may be the mechanism by which TNF-alpha alters endothelial permeability.