Effects of a putative prostaglandlin E² antagonist, AH6809, on chondrogenesis in serum‐free cultures of chick limb mesenchyme

Abstract

In the present study, we have examined the effects of a putative antagonist of prostaglandin E2 (PGE2), AH6809, on chondrogenesis in serum-free cultures of mesenchyme from distal tips of stage 25 chick limb buds in order to test the hypothesis that endogenous PGE2, through receptor-linked adenylate cyclase (AC), initiates differentiation of cartilage in limb mesenchyme. Daily addition of 10(-4) M concentrations of AH6809 produced marked inhibition of chondrogenesis over a 5-day period of cell culture as evaluated by Alcian green binding to cartilage matrix components. Inhibition of chondrogenesis by this compound was further shown to be reversible and treatment of cells with the antagonist limited to periods when chondrocytes had differentiated and were actively secreting cartilage-specific matrix components had little effect. Preincubation of control cells in 10(-4) M concentrations of AH6809 inhibited PGE2-induced activation of AC by greater than 80% without significant (P greater than .05) inhibition of basal activity by the antagonist. Responses to parathyroid hormone, which increased AC activity by 7-fold, and forskolin which increased AC activity by 23-fold in control cells, were also uninhibited by preincubation in AH6809. The results demonstrate that blockade of PGE2-AC linked receptors in prechondrogenic limb mesenchyme inhibits chondrogenesis supporting the hypothesis that endogenous PGE2 concentrations in undifferentiated limb mesenchyme play an initiating role in the differentiation of cartilage.

Neuroprotective role of bradykinin because of the attenuation of pro‐inflammatory cytokine release from activated microglia

“…The amount of TNF‐α release was reduced from 2910 ± 12.6 to 263 ± 34 and to 2019 ± 73 pg/mL ( n = 3) by EP‐2 agonist (ONO‐AE1‐259‐01, 1 μmol/L) and EP‐4 agonist (ONO‐AE1‐329, 10 μmol/L), respectively. The involvement of prostaglandins and their receptors were confirmed by the facts that both COX‐2 inhibitor (NS398, 10 μmol/L) and PGE 2 antagonist (AH6809, 10 μmol/L) (Capehart and Biddulph 1991) antagonized the effect of BK on LPS‐induced TNF‐α release (Fig. 2c).…”

Identification of γA‐like protocadherin expressed during chick development

“…High density micromass cultures derived from stage 22 to 23 [Hamburger and Hamilton, 1951] whole chick limb buds were prepared as described previously [Capehart and Biddulph, 1991]. After 18 h, cultures were washed with phosphate buffered saline (PBS) and incubated 1 h on ice with 150 μg/ml biotinylated peanut agglutinin (bPNA, Vector Labs, Burlingame, CA).…”

Skeletal developmental effects of selective and nonselective cyclooxygenase-2 inhibitors administered through organogenesis and fetogenesis in Wistar CRL:(WI)WUBR rats

“…Such changes accelerate the pathophysiological cascade of cartilage destruction (Pilbeam et al, 1993;Morisset et al, 1998). It is postulated that prostacycline and PGF 2␣ may exert positive effects on cartilage, increasing the level of glucocorticoid receptors in chondrocytes (DiBattista et al, 1991), influencing chondrocytes differentiation (O'Keefe et al, 1992) and proliferation (Capehart and Biddulph, 1991), and mediating the effects of vitamin D (Schwartz, 1992).…”