Mao Ganming, Douglas Dalon, Prajapati Milankumar, Janardhanam Raghavendra Rao Trishaal, Zheng Haiyan, Zhao Caifeng, Billack Blase
Department of Pharmaceutical Sciences, St. John's University, Queens, NY 11439, USA.
Department of Pathology and Laboratory Medicine, Brown University, Providence, RI 02912, USA.
Curr Res Toxicol. 2024 Jun 23;7:100184. doi: 10.1016/j.crtox.2024.100184. eCollection 2024.
Croton oil is liquid at room temperature, with a pale-yellow color and spicy odor. It is commonly used in combination with phenol as a chemical peeling agent in dermatology, which reveals its caustic exfoliating effects. Topical use of croton oil at a high dose produces skin irritation, inflammation, swelling, pain, and even tumors. Therefore, croton oil has been widely used for inflammation, pain, and tumor related research, with different animal models having been established. However, mechanistic studies through which croton oil induces skin swelling, injury and activates tissue repair/regeneration are limited. The present study used croton oil to induce mouse ear edema and examined tissue responses 4 h after exposure. To this end, croton oil was applied to the ventral side of mouse ears, followed by tissue collection. Samples were analyzed by hematoxylin and eosin (H&E) staining, toluidine blue staining, and immunohistochemistry staining for myeloperoxidase (MPO) and matrix metalloproteinase-9 (MMP-9). Western blotting and ELISA were also carried out for MMP-9 together with unbiased proteomic analysis using mass-spectrometry. Results from our study demonstrated that as soon as 4 h of exposure to 2.5 % croton oil, the expression levels of MPO and MMP-9 in the dermis significantly increased compared to acetone-treated (vehicle) control ears, as did other inflammatory reactions such as swelling and neutrophil aggregation and infiltration. Subsequently, proteomic analysis confirmed that croton oil treatment resulted in significant upregulation of proteins such as myeloperoxidase (MPO), matrix metalloproteinase-9 (MMP-9), and matrix metalloproteinase-8 (MMP-8) in the ear skin. Interestingly, mouse ears treated with acetone vehicle showed differential expression of 2,478 proteins relative to naïve tissues; among those differentially expressed in acetone-treated samples were members of the phosphatidylinositol-glycan biosynthesis class N, T and U proteins (PIGN, PIGT, and PIGU). Overall, this work confirms the presence of neutrophil-derived MPO and MMP-9 and extends the body of knowledge to show that MMP-8 is also present during croton oil-mediated skin inflammation in the mouse ear; moreover, we find that acetone vehicle is not inert and has effects on the skin that should be considered moving forward.
巴豆油在室温下呈液体状，颜色淡黄，气味辛辣。在皮肤科，它通常与苯酚联合用作化学剥脱剂，这显示出其腐蚀性剥脱作用。高剂量局部使用巴豆油会引起皮肤刺激、炎症、肿胀、疼痛，甚至肿瘤。因此，巴豆油已被广泛用于炎症、疼痛和肿瘤相关研究，并建立了不同的动物模型。然而，关于巴豆油诱导皮肤肿胀、损伤并激活组织修复/再生的机制研究有限。本研究使用巴豆油诱导小鼠耳部水肿，并在暴露4小时后检查组织反应。为此，将巴豆油涂抹于小鼠耳部腹侧，随后收集组织。样本通过苏木精和伊红（H&E）染色、甲苯胺蓝染色以及髓过氧化物酶（MPO）和基质金属蛋白酶-9（MMP-9）的免疫组织化学染色进行分析。还对MMP-9进行了蛋白质印迹法和酶联免疫吸附测定（ELISA），并使用质谱进行了非偏向蛋白质组学分析。我们的研究结果表明，暴露于2.5%巴豆油仅4小时，与丙酮处理（赋形剂）的对照耳部相比，真皮中MPO和MMP-9的表达水平显著增加，其他炎症反应如肿胀以及中性粒细胞聚集和浸润也是如此。随后，蛋白质组学分析证实，巴豆油处理导致耳部皮肤中髓过氧化物酶（MPO）、基质金属蛋白酶-9（MMP-9）和基质金属蛋白酶-8（MMP-8）等蛋白质显著上调。有趣的是，用丙酮赋形剂处理的小鼠耳部相对于未处理组织显示出2478种蛋白质的差异表达；在丙酮处理样本中差异表达的蛋白质包括磷脂酰肌醇聚糖生物合成N、T和U类蛋白（PIGN、PIGT和PIGU）。总体而言，这项工作证实了中性粒细胞来源的MPO和MMP-9的存在，并扩展了知识体系，表明在巴豆油介导的小鼠耳部皮肤炎症过程中也存在MMP-8；此外，我们发现丙酮赋形剂并非惰性物质，其对皮肤有影响，在后续研究中应予以考虑。
