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Effects of pH on the inhibition of fatty acid amidohydrolase by ibuprofen.
Umeå universitet, Medicinska fakulteten, Institutionen för farmakologi och klinisk neurovetenskap, Farmakologi.
Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
Umeå universitet, Medicinska fakulteten, Institutionen för farmakologi och klinisk neurovetenskap, Farmakologi.
Visa övriga samt affilieringar
2001 (Engelska)Ingår i: British Journal of Pharmacology, ISSN 0007-1188, E-ISSN 1476-5381, Vol. 133, nr 4, s. 513-520Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

The pharmacological properties of fatty acid amidohydrolase (FAAH) at different assay pH values were investigated using [(3)H]-anandamide ([(3)H]-AEA) as substrate in rat brain homogenates and in COS-1 [corrected] cells transfected with wild type and mutant FAAH. Rat brain hydrolysis of [(3)H]-AEA showed pH dependency with an optimum around pH 8-9. Between pH 6.3 and 8.2, the difference in activity was due to differences in the V(max), rather than the K(M) values. For inhibition of rat brain [(3)H]-AEA metabolism by a series of known FAAH inhibitors, the potencies of the enantiomers of ibuprofen and phenylmethylsulphonyl fluoride (PMSF) were higher at pH 5.28 than at pH 8.37, whereas the reverse was true for oleyl trifluoromethylketone (OTMK) and arachidonoylserotonin. At both pH values, (-)ibuprofen was a mixed-type inhibitor of FAAH. The K(i)((slope)) and K(i)((intercept)) values for (-)ibuprofen at pH 5.28 were 11 and 143 microM, respectively. At pH 8.37, the corresponding values were 185 and 3950 microM, respectively. The pH dependency for the inhibition by OTMK and (-)ibuprofen was also seen in COS-1 [corrected] cells transiently transfected with either wild type, S152A or C249A FAAH. No differences in potencies between the wild type and mutant enzymes were seen. It is concluded that the pharmacological properties of FAAH are highly pH-dependent. The higher potency of ibuprofen at lower pH values raises the possibility that in certain types of inflamed tissue, the concentration of this compound following oral administration may be sufficient to inhibit FAAH.

Ort, förlag, år, upplaga, sidor
2001. Vol. 133, nr 4, s. 513-520
Nyckelord [en]
Anandamide, fatty acid amidohydrolase, ibuprofen, pH dependency
Identifikatorer
URN: urn:nbn:se:umu:diva-4508DOI: 10.1038/sj.bjp.0704113PubMedID: 11399668OAI: oai:DiVA.org:umu-4508DiVA, id: diva2:143640
Tillgänglig från: 2005-04-19 Skapad: 2005-04-19 Senast uppdaterad: 2019-11-19Bibliografiskt granskad
Ingår i avhandling
1. Fatty acid amide hydrolase - A target for anti-inflammatory therapies?
Öppna denna publikation i ny flik eller fönster >>Fatty acid amide hydrolase - A target for anti-inflammatory therapies?
2005 (Engelska)Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
Alternativ titel[sv]
Fettsyraamidohydrolas - Ett mål för antiinflammatoriska läkemedel?
Abstract [en]

Anti-inflammatory drugs are a widely used class of therapeutic agents, but the use of non-steroidal anti-inflammatory drugs (NSAID) is hampered by their gastrointestinal side-effects. Recent reports that cyclooxygenase-2 inhibitors may cause cardiovascular events underline the importance of identifying new therapeutic strategies for the treatment of inflammation. One such target could be agents modifying the endogenous cannabinoid (endocannabinoid) system, since there is evidence that this system plays a role in our natural defence against inflammation. The levels of the endocannabinoid anandamide (arachidonoyl ethanolamide, AEA) are low under normal conditions, and stand under strict regulatory control of synthesising and degrading enzymes. Fatty acid amide hydrolase (FAAH) is the main enzyme degrading AEA, hydrolysing it to ethanolamine and arachidonic acid. The focus of this thesis lies in exploring the pharmacology of FAAH to evaluate its possibilities as a target for new anti-inflammatory drugs.

In Papers I and II, the effects of the ambient pH on the properties of FAAH were investigated, since tissue pH is known to decrease under inflammatory conditions. In homogenates, it was found that the activity of FAAH decreased as the assay pH was decreased, consistent with the known pH profile of the enzyme. More importantly, the sensitivity of the enzyme to inhibition by FAAH inhibitors changed. In particular, the sensitivity of the enzyme to inhibition by the NSAID ibuprofen increased seventeen-fold as the assay pH decreased from 8.37 to 5.28. A similar pattern was found using intact C6 glioma cells when the extracellular, but not the intracellular pH was reduced. Thus, at an extracellular pH value of 6.2, (R)-ibuprofen, (S)-flurbiprofen and (R,S)-flurbiprofen inhibited the metabolism of AEA with IC50 values of 26, 14 and 15 µM, respectively. These values are in theory reachable upon normal dosing of the compounds. In Paper III, the effect of the selective FAAH inhibitor URB597 and the NSAID indomethacin were investigated in vivo upon the oedema response to carrageenan administration in the paw of anaesthetised mice. Both compounds reduced the oedema in a manner completely blocked by the CB2 receptor antagonist SR144528. In Paper IV, the effect of inflammation upon endocannabinoid synthesis was investigated in mice. Lipopolysaccharide-induced pulmonary inflammation was found not to affect the release of AEA to any obvious extent, and did not change the activities of the AEA synthesising enzymes N-acyl transferase or N-acyl phosphatidylethanolamine phospholipase D, or of FAAH in lung tissue.

The results of this thesis would suggest that FAAH inhibitors can produce anti-inflammatory effects, and that the endocannabinoid system contributes to the actions of the NSAID indomethacin in the carrageenan model of inflammation, but that an increased endocannabinoid synthesis (a prerequisite for FAAH inhibition as a therapeutic strategy) is not an obligatory response to an inflammatory stimulus.

Ort, förlag, år, upplaga, sidor
Umeå: Farmakologi och klinisk neurovetenskap, 2005. s. 97
Serie
Umeå University medical dissertations, ISSN 0346-6612 ; 944
Nyckelord
Pharmacology, FAAH, anandamide, endocannabinoid, pH, inflammation, NSAID, Farmakologi
Nationell ämneskategori
Farmakologi och toxikologi
Forskningsämne
medicinsk farmakologi
Identifikatorer
urn:nbn:se:umu:diva-504 (URN)91-7305-823-8 (ISBN)
Disputation
2005-05-13, E04, 6E, Norrlands universitetssjukhus, Umeå, 13:00 (Engelska)
Opponent
Handledare
Tillgänglig från: 2005-04-19 Skapad: 2005-04-19 Senast uppdaterad: 2018-01-13Bibliografiskt granskad

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