Expression of prostaglandin H synthase-2 in endotoxic shock induced in rats.

We investigated the expression of prostaglandin H synthase-2 in rats subjected to endotoxic shock. The prostaglandin H synthase activities were assessed by measuring the plasma prostaglandins (PGE2 and 6-keto-PGF1 alpha) after arachidonic acid administration (3 mg/kg, i.v.). The plasma prostaglandin concentrations increased immediately after administration of arachidonic acid, reached a peak at 30-60 seconds, and then rapidly decreased. Lipopolysaccharide (1 mg/kg, i.v.) also increased the plasma prostaglandin concentrations, reached a peak 1 hour after administration, and then gradually decreased to normal levels. The production of plasma prostaglandin, induced by administration of arachidonic acid, was markedly enhanced in the lipopolysaccharide-treated rats. A low dose of acetylsalicylic acid (3 mg/kg, i.v.) blocked the prostaglandin production in the nontreated rats but not in the lipopolysaccharide-treated rats. In the latter group of rats, a high dose of acetylsalicylic acid (30 mg/kg, i.v.), given 10 to 30 minutes before administration of arachidonic acid, completely blocked the prostaglandin production, but recovery of this production was seen with acetylsalicylic acid (30 mg/kg) treatment at 1 to 2 hours before administration of arachidonic acid. These data suggest that pretreatment with lipopolysaccharide enhances the prostaglandin production by forming newly synthesized prostaglandin H synthase. Immunoblots of the levels of enzyme protein from rat aorta endothelial cells were analyzed. The enzyme protein cross-reacting with antibody against prostaglandin H synthase-2 was increased by lipopolysaccharide treatment in endothelial cells, and was constitutively expressed in the stomach, kidney and liver, but not in the lung and the intestine. The induction of prostaglandin H synthase-2 by lipopolysaccharide treatment was observed only in endothelial cells. The enhancement of the prostaglandin production in lipopolysaccharide-treated rats was blocked by pretreatment with dexamethasone, prior to administration of lipopolysaccharide, this suppression is apparently the result of a decrease of the prostaglandin H synthase-2 protein in endothelial cells, as determined by Western blotting. The enhanced production of prostaglandin, induced by lipopolysaccharide, seems to be due to the in vivo expression of prostaglandin H synthase-2.

NS-398, a novel non-steroidal anti-inflammatory drug with potent analgesic and antipyretic effects, which causes minimal stomach lesions.

1. NS-398 (N-[2-cyclohexyloxy-4-nitrophenyl] methanesulfonamide) is a new non-steroidal anti-inflammatory drug (NSAID) with analgesic and antipyretic effects. 2. The anti-inflammatory potency of NS-398 in rat carrageenin-induced edema was as potent as that of indomethacin and 8 times more potent than diclofenac. In rat adjuvant arthritis, NS-398 showed a therapeutic effect comparable to that seen with loxoprofen but less than that seen with indomethacin and diclofenac. 3. The analgesic potency of NS-398 in rat adjuvant arthritic pain was much the same as that of indomethacin, and was about 3-5 times higher than that of diclofenac and loxoprofen. In the Randall-Selitto method in rats, NS-398 was 2-7 times as potent as loxoprofen, diclofenac and indomethacin. In acetic acid-induced writhing in mice, NS-398 was equipotent to indomethacin and diclofenac. 4. In LPS-induced fever in rats, NS-398 was 1.5-4.5 times as potent as loxoprofen and indomethacin, but less potent than diclofenac. 5. NS-398 produced little gastric ulceration in doses of up to 1000 mg/kg, while reference drugs produced distinct stomach lesions in doses of 10-30 mg/kg. 6. NS-398 inhibited prostaglandin (PG) endoperoxide synthase from sheep seminal vesicle microsomes less potent than that of ibuprofen.

A new test for evaluating nonsteroidal anti-inflammatory drugs in vitro: inhibition of prostaglandin E2 production in minced intestinal tissue.

A new method for evaluating the inhibitory effect of nonsteroidal anti-inflammatory drugs on prostaglandin E2 production is presented. Minced rat intestinal tissue was incubated with a nonsteroidal anti-inflammatory drug and homogenized. After centrifugation, prostaglandin E2 in the supernatant was measured by radioimmunoassay. The production of prostaglandin E2 was decreased by nonsteroidal anti-inflammatory drugs, in a concentration-dependent manner, the order of potency being diclofenac greater than loxoprofen greater than indomethacin greater than ibuprofen greater than TA-847 (imidazole derivative). Loxoprofen, a prodrug, inhibited the prostaglandin E2 production. In a classical enzyme assay with sheep seminal vesicle microsomal prostaglandin endoperoxide synthase, the order of inhibitory potency was TA-847 greater than diclofenac greater than indomethacin greater than ibuprofen. The inhibitory effect of loxoprofen was very weak. On the other hand, the potency order of these nonsteroidal anti-inflammatory drugs, with regard to their anti-inflammatory effect as determined in the rat carrageenin-induced paw edema model, was loxoprofen greater than indomethacin greater than diclofenac greater than ibuprofen greater than TA-847. The results of the new method with intestinal tissue showed thus a good correlation with the anti-inflammatory activity in vivo.

Immunopharmacological studies of new 3-benzoyl-4-mercaptobutyric acids. Immunomodulating effects.

A number of D-penicillamine (PA) derivatives (3-benzoyl-4-mercaptobutyric acids) having acetylthio groups on an alpha or beta position of a carboxylic acid, were synthesized and examined for their immunological effects compared with PA. New PA derivatives suppressed adjuvant-induced arthritis (AA) in SD rats and enhanced AA in Lewis rats like PA. Suppressive effects of 2-acetylthiomethyl-3-(4-methyl-benzoyl)propionic acid (compound II-3) on AA in SD rats was most potent among PA derivatives and PA. II-3 enhanced type II collagen-induced arthritis in rats more effectively than PA, and it slightly prolonged the survival time of NZBXNZW hybrid (BWF1) mice. Hemolytic plaque forming cells in the spleen cells of BDF1 and aged Balb/c mice were potentiated but those of BWF1 were suppressed by both compounds. In in vitro experiments, both compounds enhanced lymphocyte transformation. On the contrary, II-3 had no effect on the acute inflammatory response, delayed type hypersensitivity and IgE antibody response. The abnormal release of lysosomal enzymes from the peritoneal macrophages of aged MRL/l mice were suppressed by both compounds. These results suggest that II-3 is an immunomodulator like PA but more effective than PA. II-3 may be clinically effective for rheumatoid arthritis.