RESUMO
OBJECTIVE: To study the effect of hyaluronan on cell adhesion and recruitment both in vitro and in vivo, since hyaluronan both inhibits restenosis and is anti-inflammatory. When administered to animals undergoing angioplasty the recruitment of cells into the restenotic plaque is inhibited, as well as into inflammatory lesions. The recent discovery that ICAM-1 binds hyaluronan and exhibits the B(X(7))B HA binding motif, led us also to investigate whether cell adhesion could be modulated by hyaluronan. MATERIALS AND METHODS: Human neutrophils were adhered to human umbilical vein (HUVEC) or Ea.hy.926 HUVEC cells stimulated with phorbol myristate acetate (PMA) or tumour necrosis factor (TNFalpha). Neutrophil binding in vivo utilized FMLP-stimulated hamster cheek pouch post-capillary venules. RESULTS: Hyaluronan inhibited human neutrophil adhesion to both PMA and TNFalpha-stimulated HUVEC. Ea.hy.926 human immortal HUVECs expressed ICAM-1 in response to TNFalpha and PMA. E-selectin was also upregulated by 6 h with TNFalpha but not significantly with PMA. TNFalpha induced CD44 expression within 4 h, but PMA not significantly up to 6 h. However, specific binding of [125I]hyaluronan to Ea.hy.926 cells was increased by PMA-stimulation at 4 h. Neutrophil adhesion to PMA-stimulated Ea.hy.926 HUVECs was inhibited in a concentration dependent fashion by both anti-ICAM-1 and hyaluronan (1 ng/ml-10 microg/ml) at 4 h. At 1 mg/ml adhesion was stimulated by hyaluronan. Hyaluronan had no effect on neutrophil adhesion to resting Ea.hy.926 cells. Hyaluronan (25 mg/kg, i.v.) inhibited cell adhesion to FMLP-stimulated post capillary venules of the hamster cheek pouch, whilst leaving cell rolling unaffected. CONCLUSIONS: These results show that hyaluronan, at concentrations below those where intra-molecular associations occur, binds selectively to stimulated endothelial cells and inhibits neutrophil adhesion in vitro and in vivo via a mechanism which may involve molecules other than CD44, such as ICAM-1.
