Evidence for two distinct pathways in TNFalpha-induced membrane and soluble forms of ICAM-1 in human osteoblast-like cells isolated from osteoarthritic patients.
OBJECTIVE
The present study aimed to investigate the modulation of membrane-bound intercellular adhesion molecule-1 (mICAM-1) and soluble ICAM-1 (sICAM-1) expression by tumor necrosis factor-alpha (TNFalpha) in human osteoarthritic (OA) osteoblasts.


METHODS
Cultured human primary osteoblasts were stimulated with increasing concentrations of human recombinant TNFalpha. Expression of mICAM-1 and sICAM-1 was evaluated by immunocytochemistry, enzyme-linked immunosorbent assay and semi-quantitative reverse transcriptase-polymerase chain reaction. In addition, we investigated the molecular mechanisms underlying ICAM-1 induction by TNFalpha, focusing on the activation of the mitogen-activated protein kinases (MAPKs) and nuclear factor-kappaB (NF-kappaB) pathways.


RESULTS
Our data showed that TNFalpha dose-dependently increased mICAM-1 and sICAM-1 expression at the protein and mRNA levels in OA osteoblasts. The inhibitor of de novo mRNA synthesis, actinomycin D, suppressed TNFalpha-induced mICAM-1 and sICAM-1 expression. Upon examination of the signaling components, we found that TNFalpha was a potent activator of p38, p44/42, p54/46 MAPK, and IkappaBalpha (IkappaBalpha). The chemical inhibitors of p38, p44/42 MAPK, and NF-kappaB blocked TNFalpha-induced mICAM-1 expression but not that of sICAM-1. Transfection experiments revealed that p38 MAPK or IkappaB kinase alpha (IKKalpha) overexpression enhanced TNFalpha-induced mICAM-1 production. Furthermore, osteoblasts treatment with a chemical inhibitor of metalloproteinase-9 (MMP-9) activity, a proteolytic enzyme involved in ICAM-1 cleavage, evoked a significant 25% decrease of TNFalpha-induced sICAM-1 release.


CONCLUSIONS
Taken together, these findings illustrate the central role played by TNFalpha in the regulation of ICAM-1. We suggest that TNFalpha differentially regulates sICAM-1 and mICAM-1 expression and that sICAM-1 release involves, in part, the proteolytic cleavage of mICAM-1 by MMP-9. The capacity of the MMP-9 inhibitor to prevent sICAM-1 production may be useful for the development of novel therapeutic approaches relevant to OA.
Shi
Q
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Orthopaedics Research Laboratory, Department of Orthopaedics, Sacre-Coeur Hospital, Montréal, Québec, Canada.
Benderdour
M
M
Lavigne
P
P
Ranger
P
P
Fernandes
J C
JC
D003609:Dactinomycin / Q000494:pharmacology; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism*; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D010003:Osteoarthritis / Q000378:metabolism*; D010006:Osteoblasts / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D018799:Intercellular Adhesion Molecule-1; D003609:Dactinomycin; D020929:Mitogen-Activated Protein Kinase Kinases; D020780:Matrix Metalloproteinase 9