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Hintergrund Die In-vitro-Effekte selektiver COX-2-Inhibitoren (L-745,337, NS-398 und DFU) und des COX-unspezifischen Diclofenacs auf die Freisetzung von PGE2 und 6-keto-PGF1α aus Bursitis-subacromialis-Gewebe (BSG) von 35 Patienten mit Subakromialsyndrom wurden verglichen.

Background: The aim of this study was to investigate whether the pulmonary response to inflammatory stimulation, resulting in increased vascular resistance and permeability, could be attenuated by short-term infusion of triglycerides containing ω-3 fatty acids. With the concept of altering the composition of membrane phospholipids in such a manner that stimulation resulted in the release of less vasoconstrictive and permeability-enhancing metabolites of eicosapentaenoic acid instead of those of arachidonic acid (AA), the parenteral application of a lipid emulsion prepared from fish oil (Omegavenös®) was tested in comparison with a soy oil preparation (Lipovenös®). Methods: Isolated lungs from anesthetized rabbits were ventilated and recircuatingly perfused (200 ml/min) with 200 ml cell-free buffer solution to which either 2 ml saline (controls, n = 6), 2 ml Lipovenös 10% (n = 6) or 2 ml Omegavenös 10% (n = 6) were added. To study the possible metabolic alterations in states of an enhanced AA turnover, lungs of each group were stimulated with smaller doses of A23187 (10–8M) during the 180 -min lipid perfusion period, followed by a 10 times higher calcium ionophore A 23187 (10–7M) challenge after washing out the lipids by exchange of perfusion fluid. Pulmonary artery pressure (PAP) and the lung weight gain indicating edema formation were monitored, and eicosanoids were analyzed in samples of the perfusate. Results: Upon A23187 injection lung weight gain and PAP increase were significantly reduced (50%) in Omegavenös-perfused lungs in comparison with controls and Lipovenös treatment. The vascular reactions were accompanied by a shifting from LTC4 to LTC5 during and after Omegavenös perfusion. Conclusion: The data demonstrate that ω-3 fatty acids seem to be incorporated into the phospholipid pool of the pulmonary tissue, even after short-term infusion (3 h) resulting in an attenuated pressure reaction and edema formation due to an altered spectrum of metabolites in the case of inflammatory stimulation.

1. 1. Specific radioimmunoassays for the prostaglandins E 2 , F 2α and A 2 and the metabolites 13,14-dihydro-15-keto-prostaglandin E 2 , 15-keto-prostaglandin F 2α and 13,14-dihydro-15-keto-prostaglandin F 2α were used to study the metabolism of prostaglandins by gastroscopically obtained small biopsy specimens of human gastric fundus mucosa. 2. 2. Three prostaglandin-metabolizing enzymes were found in the 100000 × g supernatant of human gastric fundus mucosa, 15-hydroxy-prostaglandin-dehydrogenase, Δ 13 -reductase and Δ 9 -reductase. The specific activity was highest for 15-hydroxy-prostaglandin-dehydrogenase and lowest for Δ 9 -reductase. 3. 3. Formation of prostaglandin A 2 (or B 2 ) was not observed under the same conditions. 4. 4. None of the three enzyme activities detected in the 100000 × g supernatant was found in the 10000 × g and 100000 × g pellets of human gastric fundus mucosa. 5. 5. The results indicate that high speed supernatant derived from human gastric mucosa can rapidly metabolize prostaglandin E 2 and prostaglandin F 2α , to the 15-keto and 13,14-dihydro-15-keto-derivatives. Furthermore, prostaglandin E 2 can be converted to prostaglandin F 2α , the biological activity of which, on gastric functions, differs from that of prostaglandin E 2 .

Sensitive and relatively specific radioimmunoassays for the prostaglandin (PG) derivatives 15(S)-15-methyl-PGE2 and 15(S)-15-methyl-PGF2α are described. The radioimmunoassays were used for an in vitro study on the metabolism of the PG derivatives. While both methylated PGs are resistant to the action of 15-hydroxy-PG-dehydrogenase, 15(S)-15-methyl-PGE2 is converted to the corresponding PGF2α derivative by the action of the enzyme PGE-9-keto-reductase.

Rat jejunal tissue in vitro synthesizes large amounts of prostaglandin (PG) D 2 and smaller amounts of 6-keto-PGF 1α and PGE 2 , whereas human small intestinal mucosa synthesizes much smaller amounts of the three PG determined with about equal amounts of PGE 2 and PGD 2 . Intraperitoneal administration of bacterial endotoxin to rats induces fluid accumulation in the small intestine and increases significantly the release of PGD 2 , PGE 2 and 6-keto-PGF 1α into the small intestinal lumen in vivo. Endotoxin-induced stimulation of PG release is particularly pronounced for PGD 2 . Fluid accumulation and PG output are inhibited by indomethacin. It seems possible that the different total amounts of PG synthesized by small intestinal tissue of man and rat as well as the different pattern of PG released might contribute to species-specific responses of the gastrointestinal tract to various pathophysiological stimuli.

The recent advances in hapten immunoassays, especially radioimmunoassays for drugs, are reviewed. The progress in immunoassay methodology as well as the development of new radioimmunoassays for low molecular weight compounds are described.

1. 1. Specific radioimmunoassays for the prostaglandins E2, A2 and F2α were used to study the synthesis of prostaglandins by gastroscopically obtained small biopsy specimens of human gastric corpus mucosa. 2. 2. Both prostaglandin E2 and prostaglandin F2α were found to be synthesized from arachidonic acid by the microsomal fraction of human gastric mucosa. The synthesis of prostaglandin E2 exceeded that of prostaglandin F2α by a factor of about 10. 3. 3. Synthesis of prostaglandin A2 or prostaglandin B2 was not observed under the same incubation conditions. 4. 4. Indometacin effectively inhibited synthesis of both prostaglandin E2 (ID50 4.2 μg/ml) and prostaglandin F2α (ID50 1.8 μg/ml) by human gastric mucosa, while paracetamol even in a concentration of 310 gm/ml did not influence prostaglandin synthesis. The anti-ulcer agent carbenoxolone, which has been shown to inhibit prostaglandin inactivation, at the same concentration only slightly inhibited (about 20%) prostaglandin synthesis. 5. 5. The results support the hypothesis that the gastro-intestinal effects or side effects of several drugs are mediated by an influence on the enzymes of prostaglandin sythesis or inactivation.

A specific radioimmunoassay for prostaglandin D2 was developed. Using the radioimmunoassay, prostaglandin D2 synthesis by human thrombocytes was measured. While the cyclooxygenase inhibitor indomethacin inhibits formation of prostaglandin D2, increased formation of prostaglandin D2 was observed in the presence of the thromboxane synthetase inhibitor imidazole.

Radioimmunoassays for prostaglandin D2 (PGD2) and thromboxane B2 (TXB2) were developed. Antisera were obtained from rabbits immunized with the corresponding hapten-bovine serum albumine conjugates. The conjugates were synthesized using N,N′ -carbonyldiimidazole as coupling reagent (Axen, Prostaglandins 5, 45, 1974). The radioactive tracer for the TXB2 assay was synthesized by incubation of 3H-arachidonic acid as substrate with washed human thrombocytes as enzyme source. The label for the PGD2 assay was also synthesized from 3H-arachidonic acid using tne microsomal fraction of sheep seminal vesicles as enzyme source. Both radioimmunoassays are highly specific and recognize the ring structure and the hydroxyl group at position C15 as immunodominant parts of the hapten molecules. The detection limits are 40 pg for TXB2 and 200 pg for PGD2. The radioimmunoassays were used to determine the release of TXB2 and PGD2 from washed human thrombocytes after addition of thrombin. While only small amounts of immunoreactive PGD2 were released in control incubates, the TX synthetase inhibitor imidazole (1.47 mM) increased PGD2 as well as PGF2α and PGE2 release and simultaneously reduced TXB2 release significantly. Indometacin (2.8 μM) abolished both TXB2 and PG release in response to thrombin. The PGD2 found in the incubates could be formed either enzymatically or non-enzymatically by isomerization from its endoperoxide precursor PGH2. The thrombin-induced formation of PGD2 by thrombocytes could be important in view of the potent antiaggregatory activity of PGD2.

Human gastric juice contains higher concentrations of PG metabolites than of unmetabolized PG indicating that local metabolism might play a role in limiting the biological activity of PG in gastric mucosa and has to be considered when investigating endogenous gastric PG. A major fraction of the 15-keto-13,14-dihydro-PGE 2 (KH 2 PGE 2 ) formed in gastric mucosa and released into the gastric lumen seems to be rapidly dehydrated to a compound co-chromatographing with KH 2 PGA 2 , while the amounts of the bicyclic degradation product 11-deoxy-13,14-dihydro-15-keto-11,16-cyclo-PGE 2 (11-deoxy-KH 2 -cyclo-PGE 2 ), as measured by radioimmunoassay, in freshly extracted gastric juice are negligible. Stimulation of secretion with pentagastrin does not influence significantly the concentrations of PG and PG metabolites in human gastric juice, but total output tends to increase parallel to the increase in secretion volume. Levels of immunoreactive 6-keto-PGF 1α in human gastric juice are much lower than those of PGE 2 . Since human gastric mucosa synthesizes considerable amounts of PGI 2 and 6-keto-PGF 1α in vitro, the low levels of 6-keto-PGF 1α in gastric juice might indicate that PGI 2 formed by gastric mucosa in vivo is, like PGE 2 and PGF 2α , rapidly metabolized and/or removed preferentially via the blood stream.