An ex vivo skin model to probe modulation of local cutaneous arachidonic acid inflammation pathway
Main Article Content
Keywords
skin, inflammation, COX-2, PGE2, ex vivo
Abstract
There is a need for inexpensive and reliable means to determine the modulation of cutaneous inflammation. The method outlined in this article draws together a number of scientific techniques and makes use of generally unwanted biological tissues as a means of determining skin inflammation ex vivo, and focuses on probing aspects of the arachidonic acid inflammation pathway. Freshly excised skin contains elevated levels of short-lived inducible cyclooxygenase-2 (COX-2) and, under viable conditions, COX-2 and its eicosanoid products will continue to be produced until tissue necrosis, providing a window of time in which relative levels can be probed to determine exacerbation due to an upregulating factor or downregulation due the presence of an agent exerting anti-inflammatory activity. Ex vivo porcine skin, mounted in Franz diffusion cells, is dosed topically with the xenobiotic challenge and then techniques such as Western blotting and immunohistochemistry can then be used to probe relative COX-2 levels on a semi-quantitative or qualitative level. Enzyme-linked immunosorbent assay or LCMS can be used to determine relative prostaglandin E-2 (PGE-2) levels. Thus far, the technique has been used to examine the effects of topically applied anti-inflammatories (betamethasone, ibuprofen, ketoprofen and methotrexate), natural products (fish oil, Devil’s claw extract and pomegranate rind extract) and drug delivery vehicle (polyNIPAM nanogels). Topically applied xenobiotics that modulate factors such as COX-2 and PGE-2 must penetrate the intact skin, and this provides direct evidence of overcoming the "barrier function" of the stratum corneum in order to target the viable epidermis in sufficient levels to be able to elicit such effects. This system has particular potential as a pre-clinical screening tool for those working on the development of topical delivery systems, and has the additional advantage of being in line with 3 Rs philosophy.
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References
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4. Monteleone G, Pallone F, MacDonald TT, Chimenti S, Costanzo A (2011) Psoriasis: from pathogenesis to novel therapeutic approaches. Clin Sci 120: 1-11. DOI: 10.1042/CS20100163
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6. Ahuja M, Dhake AS, Sharma SK, Majumdar DK (2008) Topical ocular delivery of NSAIDs. AAPS J. 10: 229-241. PMID: 18437583
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8. Marks F, Furstenberger G, Muller-Decker K (1998) Arachidonic acid metabolism as a reporter of skin irritancy and target of cancer chemoprevention. Toxicol Lett 97: 111–118. PMID: 9820655
9. Marnett LJ, Rowlinson SW, Goodwin DC, Kalgutkar AS, Lanzo CA (1999) Arachidonic acid oxygenation by COX-1 and COX-2. Mechanisms of catalysis and inhibition. J Bio Chem 274: 22903–22906. DOI: 10.1074/jbc.274.33.22903
10. Chandrasekharan NV, Simmons DL (2004) The cyclooxygenases. Genome Biology 5: 241. PMID: 15345041
11. Chen Y, Chen P, Hanaoka M, Droma Y, Kubo K. (2008) Enhanced levels of prostaglandin E2 and matrix metalloproteinase-2 correlate with the severity of airflow limitation in stable COPD. Respirology 13:1014-1021. PMID: 18699805
12. Minghetti L (2004) Cyclooxygenase-2 (COX-2) in inflammatory and degenerative brain diseases. J Neuropathol Exp Neurol 63: 901-910. PMID: 15453089
13. Nassar A, Radhakrishnan A, Cabrero IA, Cotsonis G, Cohen C (2007) COX-2 expression in invasive breast cancer: correlation with prognostic parameters and outcome. Appl Immunohistochem Mol Morphol 15: 255-259. DOI: 10.1097/01.pai.0000213130.63417.b3
14. Menter DG, Schilsky RL, DuBois RN (2010) Cyclooxygenase-2 and cancer treatment: understanding the risk should be worth the reward. Clin Cancer Res 16: 1384–1390. PMID: 20179228
15. Lawrence J (2016) Anti-inflammatory drugs linked with higher risk of heart failure. The Pharmaceutical Journal. DOI: 10.1211/PJ.2016.20201783
16. Cannon CP, Cannon PJ (2012) Physiology COX-2 inhibitors and cardiovascular risk. Science 336: 1386–1387. DOI: 10.1126/science. PMID: 22700906
17. Barkin RL (2015) Topical nonsteroidal anti-inflammatory drugs: the importance of drug, delivery, and therapeutic outcome. Am J Ther 22: 388-407. PMID: 22367354
18. Suto B, Berko S, Kozma G, Kukovecz A, Budai-Szucs M, Eros G, Kemeny L, Sztojkov-Ivanov A, Gaspar R, Csanyi E (2016) Development of ibuprofen-loaded nanostructured lipid carrier-based gels: characterization and investigation of in vitro and in vivo penetration through the skin. Int J Nanomedicine 11: 1201–1212. PMID: 27099487 Sütő B, Berkó S, Kozma G, Kukovecz Á, Budai-Szűcs M, et al. (2016) Development of ibuprofen-loaded nanostructured lipid carrier-based gels: characterization and investigation of in vitro and in vivo penetration through the skin.. Int J Nanomedicine 11: 1201-1212. doi: 10.2147/IJN.S99198. [View Article] [PubMed]
19. Potts R, Guy RH (1992) Predicting skin permeability. Pharm Res 9: 663–669. PMID: 1608900
20. Lademann J, Jacobi U, Surber C, Weigmann HJ, Fluhr TW (2008) The tape stripping procedure-evaluation of some critical parameters. Eur J Pharm Biopharm 72: 317–323. PMID: 18775778
21. Flecknell P (2002) Replacement, reduction and refinement. ALTEX 19: 73-78. PMID: 12098013
22. Bralley EE, Greenspan P, Hargrove JL, Wicker L, Hartle DK (2008) Topical anti-inflammatory activity of Polygonum cuspidatum extract in the TPA model of mouse ear inflammation. J Inflamm 5: 1. PMID: 18261214
23. Kim EJ, Park H, Kim J, Park JH (2010) 3,3'-diindolylmethane suppresses 12-O-tetradecanoylphorbol-13-acetate-induced inflammation and tumor promotion in mouse skin via the downregulation of inflammatory mediators. Mol Carcinog 49: 672-683. PMID: 20564344
24. Cichocki M, Blumczyńska J, Baer-Dubowska W (2010) Naturally occurring phenolic acids inhibit 12-O-tetradecanoylphorbol-13-acetate induced NF-κB, iNOS and COX-2 activation in mouse epidermis. Toxicol 268: 118-124. PMID: 20026373
25. Burke JR, Davern LB, Stanley PL, Gregor KR, Banville J, Remillard R, Russell JW, Brassil PJ, Witmer MR, Johnson G, Tredup JA, Tramposch KM (2001) BMS-229724 is a tight-binding inhibitor of cytosolic phospholipase A2 that acts at the lipid/water interface and possesses anti-inflammatory activity in skin inflammation models. J Pharmacol Exp Ther 298: 376-385. PMID: 11408565
26. De Vry CG, Valdez M, Lazarov M, Muhr E, Buelow R, Fong T, Iyer S (2005) Topical application of a novel immunomodulatory peptide, RDP58, reduces skin inflammation in the phorbol ester-induced dermatitis model. J Invest Dermatol 125: 473–481. PMID: 16117788
27. Morris CJ (2003) Carrageenan-induced paw edema in the rat and mouse. Methods Mol Biol 225: 115-121. PMID: 12769480
28. Sharma JN, Samud AM, Asmawi MZ (2004) Comparison between plethysmometer and micrometer methods to measure acute paw oedema for screening anti-inflammatory activity in mice. Inflammopharmacology 12: 89-94. PMID: 15035782
29. Albanesi C, Scarponi C, Giustizieri ML, Girolomoni G. (2005) Keratinocytes in inflammatory skin diseases. Curr Drug Targets Inflamm Allergy 4: 329-334. PMID: 16101542
30. Lau WM, Ng KW, White AW, Heard CM (2011) Therapeutic and cytotoxic effects of the novel anti-psoriasis co-drug, naproxyl-dithranol on HaCaT cells. Mol Pharm 8: 2398-2407. PMID: 21882816
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Oct 26, 2020
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Heard CM. An ex vivo skin model to probe modulation of local cutaneous arachidonic acid inflammation pathway. J Biol Methods [Internet]. 2020 Oct. 26 [cited 2024 Apr. 24];7(4):e138. Available from: https://jbmethods.org/index.php/jbm/article/view/319
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Author Biography
Charles M. Heard, Cardiff University
School of Pharmacy and Pharmaceutical Sciences. Reader in Pharmaceutical Chemistry
