Suppressed thromboxane production in endotoxin-desensitized THP-1 cells is not a result of decreased prostaglandin H synthase activity.

Pre-exposure of THP-1 cells to low concentrations of endotoxin (lipopolysaccharide, LPS) down-regulates thromboxane (Tx) A(2), an arachidonic acid (AA) metabolite, production in response to a subsequent LPS stimulation. To further delineate the mechanisms of LPS-induced down-regulation of TxA(2), we examined expression of prostaglandin H synthase (PGHS)-2 mRNA, changes in PGHS activity, and content of PGHS-1 and -2. Pre-exposure to LPS (1 mu g/mL for 18 h to desensitize cells) inhibits production of TxB(2), the stable metabolite of TxA(2), in response to secondary stimulation of LPS (10 mu g/mL), when compared with LPS-stimulated naive cells (p < .05, n = 5). LPS (10 mu g/mL) induced expression of PGHS-2 mRNA at 1 and 2 h in naive cells, but this expression was decreased in the LPS-desensitized cells. However, exogenous AA (16 mu M) or phorbol myristic acid (PMA), 3 mu M) stimulated greater TxB(2) production in the LPS-desensitized cells than in the naive cells (p < .05). Protein content of PGHS-1 and -2 were examined by Western blot analysis, using antibodies specific for PGHS-1 and PGHS-2. Densitometric analysis demonstrated a significant increase in PGHS-2 induction in LPS-stimutated naive cells (405 +/- 174%) over its respective basal group (p < .05, n = 5). PGHS-1 was constitutively present, but there was no significant difference in quantity between naive and LPS-desensitized basal or LPS-stimulated groups. Thus, despite the reduction in expression of PGHS-2 mRNA, these composite data demonstrate that down-regulation of PGHS activity (assessed with exogenous AA or PMA) cannot be responsible for the inhibition of AA metabolism observed in LPS desensitization.