The inhibition of neutrophil-endothelial cell adhesion by hyaluronan independent of CD44.
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.


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.
Alam
C A S
CA
Experimental Pathology Group, Biochemical Pharmacology, William Harvey Research Institute, Saint Bartholomew's & Royal London School of Medicine & Dentistry, Queen Mary & Westfield College, Charterhouse Square, London EC1M 6BQ, UK.
Seed
M P
MP
Freemantle
C
C
Brown
J
J
Perretti
M
M
Carrier
M
M
Divwedi
A
A
West
D C
DC
Gustafson
S
S
Colville-Nash
P R
PR
Willoughby
D A
DA
D000818:Animals; D002196:Capillaries / Q000187:drug effects / Q000502:physiology; D002448:Cell Adhesion / Q000187:drug effects; D002460:Cell Line; D002610:Cheek / Q000098:blood supply; D006224:Cricetinae; D004305:Dose-Response Relationship, Drug; D019040:E-Selectin / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D018960:Hyaluronan Receptors / Q000378:metabolism*; D006820:Hyaluronic Acid / Q000494:pharmacology*; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism; D007457:Iodine Radioisotopes; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D019040:E-Selectin; D018960:Hyaluronan Receptors; D007457:Iodine Radioisotopes; D014409:Tumor Necrosis Factor-alpha; D018799:Intercellular Adhesion Molecule-1; D009240:N-Formylmethionine Leucyl-Phenylalanine; D006820:Hyaluronic Acid; D013755:Tetradecanoylphorbol Acetate