Cyclooxygenase (COX) is the key enzyme in the production of prostaglandins, which are essential for the response of bone to mechanical loading. We determined which COX-isoform, COX-1 or COX-2, determines loading-induced prostaglandin production in primary bone cells in vitro. Mouse and human bone cells reacted to I It of pulsating fluid flow (PFF, 0.6 +/- 0.3 Pa at 5 Hz) with an increased prostaglandin E-2 production, which continued 24h after cessation of PFF. Inhibition of COX-2 activity with NS-398 abolished the stimulating effect of PFF both at I It and at 24 h post-incubation, while inhibition of COX-I by SC-560 affected neither the early nor the late response to flow. PFF rapidly stimulated COX-2 mRNA expression at I It but did not affect COX-1 mRNA expression. COX-2 mRNA expression was still significantly enhanced 24 It after cessation of PFF. We conclude that COX-2 is the mechanosensitive form of COX that determines the response of bone tissue to mechanical loading. (C) 2003 Elsevier Science (USA). All rights reserved.