An ex vivo skin model to probe modulation of local cutaneous arachidonic acid inflammation pathway.
An ex vivo skin model to probe modulation of local cutaneous arachidonic acid inflammation pathway.
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DOI：
10.14440/jbm.2020.319
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发表时间：
2020
期刊：
影响因子：
--
通讯作者：
Heard CM
中科院分区：
文献类型：
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作者：
Heard CM
需要一种廉价和可靠的方法来确定皮肤炎症的调节。本文概述的方法汇集了许多科学技术，并利用通常不需要的生物组织作为体外确定皮肤炎症的手段，并侧重于探测花生四烯酸炎症途径的各个方面。新切除的皮肤含有高水平的短寿命诱导型环氧合酶-2 (COX-2)，在有活力的条件下，COX-2及其类二十烷酸产物将继续产生，直到组织坏死，这提供了一个时间窗口，在这个时间窗口中，可以探测相对水平，以确定由于发挥抗炎活性的药物的存在而导致的加剧或下调。将离体猪皮肤置于Franz扩散细胞中，局部给药，然后使用Western blotting和免疫组织化学等技术，在半定量或定性水平上探测COX-2的相对水平。酶联免疫吸附法或LCMS可用于测定相对前列腺素E-2 （PGE-2）水平。到目前为止，这项技术已被用于检查局部应用的消炎药（倍他米松、布洛芬、酮洛芬和甲氨喋呤）、天然产品（鱼油、魔鬼爪提取物和石榴皮提取物）和药物递送载体（聚尼帕姆纳米凝胶）的效果。局部应用调节COX-2和PGE-2等因子的外源药物必须穿透完整的皮肤，这提供了克服角质层“屏障功能”的直接证据，以便在足够的水平上靶向可存活的表皮，从而能够引起这种效果。对于那些致力于局部给药系统开发的人来说，该系统具有作为临床前筛选工具的特殊潜力，并且具有符合3rs理念的额外优势。
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.