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Synthesis of Prostaglandin E2, Thromboxane B2 and Prostaglandin Catabolism in Gastritis and Gastric Ulcer

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Synthesis of Prostaglandin E2, Thromboxane B2 and Prostaglandin Catabolism in Gastritis and Gastric Ulcer Gut: first published as 10.1136/gut.27.12.1484 on 1 December 1986. Downloaded from Gut, 1986, 27, 1484-1492 Synthesis of prostaglandin E2, thromboxane B2 and prostaglandin catabolism in gastritis and gastric ulcer C J HAWKEY (WITH THE TECHNICAL ASSISTANCE OF N K BHASKAR AND B FILIPOWICZ) From the Department of Therapeutics, University Hospital, Nottingham SUMMARY Because endogenous prostaglandins may protect the gastric mucosa a study was conducted to determine factors influencing (a) the synthesis of immunoreactive prostaglandin (iPG) E2 and thromboxane (iTx) B2 as measured by radioimmunoassay and (b) prostaglandin catabolism measured radiometrically, in human gastric mucosa. Gastric mucosa was obtained at endoscopy. Synthesis of iPE2 and iTxB2 was inhibited in vitro by indomethacin; iTxB2 synthesis was also selectively inhibited by the thromboxane synthesis inhibitor dazmegrel. Prostaglandin catabolism was inhibited by carbenoxolone. Multivariate analysis showed that synthesis of iPGE2 from endogenous precursor during homogenisation was decreased in patients on non-steroidal anti-inflammatory drugs. Mucosal inflammation was associated with significantly increased synthesis of iPGE2 and decreased prostaglandin catabolism. There were no differences between the mucosa of patients with or without gastric ulcers, nor between the ulcer rim and mucosa 5 cm away. Age, sex, smoking history, and ingestion of antisecretory drugs appeared to exert no influence. In this study gastritis was the major influence on prostaglandin synthesis. It seems unlikely that prostaglandin deficiency is a strong predisposing factor for gastric ulceration. http://gut.bmj.com/ Reports suggesting gastric mucosal protection by ment of thromboxane synthesis as thromboxane endogenous prostaglandin synthesis'-3 raise the synthesis is associated with gastric ulceration in rats possibility that this process might be defective in and inhibitors of thromboxane synthesis appear to patients who develop gastric ulcers. Previous evi- be protective.7 Alternatively reduced levels of dence on this point has been conflicting. Some protective prostaglandins might result from in- studies have found reduced prostaglandin synthesis creased prostaglandin catabolism.'2I3 on September 27, 2021 by guest. Protected copyright. in gastric ulcer patients,4 even in the presence of In this study synthesis of immunoreactive prosta- gastritis: this would be surprising because other glandin (iPG)E, and of thromboxane (iTx)B,, alnd inflammatory lesions lead to enhanced prostaglan- prostaglandin catabolism in humatn gastric mucosa din synthesis. Another study reported increased have been investigated with reference both to prostaglandin synthesis in patients with gastric histological appearances and to the presence or ulcers6 and attributed this to the associated gastritis: absence of gastric ulcerattion. The relationship of however, even where this occurs it remains possible age, sex or smoking or ingestion of non-steroidal that prostaglandin synthesis could be deficient in anti-inflammatory drugs (NSAIDS) or antisecretory relation to the prevailing levels of mucosal in- drugs to these parameters has also been investigated flammation. in view of the known influence of these factors on Factors other than reduced prostaglandin synth- the incidence or natural history of gastric ulceration. esis might render the mucosa liable to ulcerate. These could include an absolute or relative enhance- Methods Addrcss for corrcspondcicncc: I)r ( J llawikcy. i)cpartnicrit of ThhcralpcLtics. PATI ENTS Univcrsity tIospitl.ll Nottiighitm NG7 2L i-. The following radiolabelled compounds were used: Reccivc(d for puhlication 22 April 1986 (H3)PGE,, 160 Ci/mmol (Amersham International); 1484 Gut: first published as 10.1136/gut.27.12.1484 on 1 December 1986. Downloaded from HiTll(t711l g(lstric Iro1sftalloi(ds 1485 (H3) TxB1, 139 3 Ci/mmol (New England Nuclear) slight increase in inflarmmatory cell infiltrate) or and (9 H3)PGF1. 13 7 Ci/mmol (Amersham Interna- inflamed (moderate or matrked increatse in in- tional). The following compounds were gifts: in- flammatory cell infiltraite) using staindard histological domethaicin (Merck Sharp & Dohme). dazmegrel criteria of inflammation. No attempt was maide to (Pfizer UK). carbenoxolone (Biorex Laboratories). differentiate between inflammnation caused malinly TxB, antiserum wtas kindlv donated by Dr Lawrence by neutrophils and inflarmmation caused mainly by Levine. PGE, antiserum wias bought from the lymphocytes. As mucosal atrophy and Imlucosail Institut Pasteur. Other chemicals were obtained inflammation were frequently associated no attempt from standard lAboratory suppliers. was made to assess them independently. Patients undergoing upper gastrointestinal endo- scopy with an Olympus T endoscope were studied. HOMOGENATES ANI) INCUBATIONS Where there was a possibility that mucosa might be The rest of the biopsy specimens were homogenised taken for research patients were asked to give immediately on ice in Tris HCI t)()5M. pH 7-4 (20 written informed consent before the endoscopy. The mg wet weight/ml finatl proportions) for 15 seconds study was approved by the South Nottingham using an Ultraturax homogenizer. Ethical Committee. In studies used to develop and validate the SYNTHESIS FROM ENDOGENOUS STORES incubation conditions and radioimmunoassalys and (ENDOGENOUS iPGE2ANI) iTxB,) to assess reproducibility with repeated study. both Ten milligram aliquots in 0 5 ml (final proportions) normail subjects aind paitients with a vairiety of of the homogenaite were extracted immediately into gastroduodenal conditions were studied. During the chloroform,'1 for the determination of prostanoids mnain study 27 patients with gastric ulcers and 43 synthesised from endogenous stores (henceforth control patients were studied. As gastritis was one of designated Endogenous iPGE, or iTxB ). the factors which the study wats designed to investi- gate. endoscopic or histological evidence of in- INHIBITOR EXPERIMENTS flarmmation was not an exclusion criterion. In all Some samples were pre-incubated in Tris HCI other respects. the control patients had normal containing either indomethacin 50 [tg/ml (4x 10 4M) endoscopic findings. The characteristics of the for 15 minutes before being homogenised in fresh patients studied are detailed in Table 1. Tris HCI also containing indomethacin 50 Rg/ml (4x10-4 M). These samples were extracted in the http://gut.bmj.com/ HIS (IOGY1 usual way and results compared with those obtained Four to five mucosail biopsy specimens were taIken from biopsies from the same patient but not exposed from the lesser curve between 45 and 50 cm from the to indomethacin. incisor teeth and used immediately. A small piece from two of the specimens, or a separate biopsy SYNTHESIS AFTER INCUBATION WITH ARACHIDONIC specimen from the sarme irea wats exatmined histolo- ACID (TOTAL iPGE2 SYNTHESIS) gically (without knowledge of the biochemical re- Other aliquots (10 mg, 0 5 ml, final proportions) sults) and graded as uninflamed (normal or only were incubated with arachidonic acid 2 utg for 30 on September 27, 2021 by guest. Protected copyright. minutes at 37°C in a rocking water bath before extraction into chloroform.'5 Because this proce- Table I (Characteristics of tile 1i)(lit.e.s Istdied dure results in synthesis of prostanoids from both endogenous and exogenous arachidonic acid it is Control GU p)atielIS designated Total iPGE2 synthesis. Total 43 27 Mcn 29 13 INHIBITOR EXPERIMENTS Stilokcrs 16 17 In some experiments aliquots of homogenates were Antisecrctory dtirgs 9 17 incubated with or without indomethacin 1 [ig/ml NSAIDS 5 4 (2*8x10-6M) or dazmegrel 0-3 [tg/ml (10-6M). The Agc <3(0 ysears 7 3(-50 II- effect of the inhibitor was calculated after allowance 50-65 9 11 for synthesis during homogenisation. 65-75 11 14 75+ 4 RADIOIMMUNOASSAYS After extraction into chloroform, samples were Pa'aticnts on rcgular NSAID's. last dosc in cach case approximiatcly 12 hours bcforc cindoscopsy Individual druLgS aIS follows:- resuspended in phosphate buffered saline (pH 7-4 indomcthacin: flivc, ispirin: tsAo. naproxen: onc. flurhuprotcn: 0-15 M), and stored at -50°C for radioimmunoas- onc. say, usually within one month (maximum three Gut: first published as 10.1136/gut.27.12.1484 on 1 December 1986. Downloaded from 1486 Hawkey months). iTxBI3 was measured by radioimmunoassay dilutions gave similar results (value at higher dilu- using anti-serum kindly donated by Dr Lawrence tion 99+3%/,, SEM, of value at lower dilution, n=6). Levine. For thromboxane assays standard TxB,, Authentic PGE, added to samples before extraction TxBI anti-serum and H13] TxB2 tracer (Amersham) was assayed as 103±4°/O (n=48) of that added. were dissolved and samples were diluted in Tris Prostaglandin E, re-assayed in samples after one to saline pH 7-4, 0-15 M with 1% gelatin. iPGE2 was three months of storage was 105±9'S (n=101) of measured by radioimmunoassay as previously initial value. The PGE, antiserum showed selective described. 16 17 reactivity with material cochromatographing with authentic PGE,. CATABOLISM Aliquots of homogenate (10 mg 0 5 ml, final propor- (ii) iTxB2 tions) were also incubated with (90-H3)PGF2, Under the assay conditions used, the cross reactivity (1 mg, 0-1 iCi) and NAD 1 mm for five minutes at of the TxB2 antiserum with other prostanoids (based 37°C. They were extracted into chloroform at pH on concentrations required to produce 50% dis- 3 5 and re-suspended in 75 Rl; chloroform:ethanol
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