三黄凝胶对耳郭皮肤缺损兔创面愈合、碱性成纤维细胞生长因子和白细胞介素17水平的影响

Effects of Sanhuang gel on wound healing,basic fibroblast growth factor and interleukin 17 levels of auricular skin defect in rabbits

背景:由于耳结构的特殊性,耳郭损伤处皮肤通常活动度及组织再生能力差,愈合困难,因此如何加速创面愈合并获得理想的修复效果一直是外科领域探索的问题及研究热点。目的:探究三黄凝胶对耳郭皮肤缺损兔创面愈合、碱性成纤维细胞生长因子和白细胞介素17水平的影响。方法:取60只雄性新西兰兔,采用随机数字法分6组,每组10只,A组不做任何处理,在B-F组兔耳郭处制作一个直径为1 cm的圆形全层皮肤缺损创面,B组创面注射生理盐水,C组创面涂抹积雪苷霜软膏,D组创面涂抹三黄凝胶,E组创面注射PI3K/AKT激动剂740 Y-P,F组创面涂抹三黄凝胶并注射740 Y-P,2次/d,连续治疗30 d。观察并记录家兔耳郭创面愈合情况,苏木精-伊红染色观察耳郭组织病理形态,RT-PCR法检测家耳郭组织碱性成纤维细胞生长因子和白细胞介素17的mRNA相对表达,免疫印迹法检测家耳郭组织PI3K/AKT通路的蛋白表达。结果与结论:①经过治疗后,皮肤创面愈合速度加快,治疗30 d后,三黄凝胶与PI3K/AKT激动剂单独应用的创面愈合率高于积雪苷霜软膏,二者联合应用的创面愈合率最高;②苏木精-伊红染色显示,B组表皮明显增厚且出现血管扩张,可见大量炎性细胞浸润及纤维组织增生;与B组相比,C、D、E、F组形态明显改善,可见新生毛细血管及大量成纤维细胞和内皮细胞增殖,其中以F组改善最明显;③RT-PCR检测显示,耳郭皮肤损伤后,碱性成纤维细胞生长因子mRNA相对表达降低、白细胞介素17 mRNA相对表达升高,经过不同治疗后两因子mRNA相对表达有不同程度地改善,三黄凝胶与PI3K/AKT激动剂单独应用的改善作用均强于积雪苷霜软膏,二者联合应用的改善作用最强;④免疫印迹检测显示,耳郭皮肤损伤后,PI3K、AKT、p-PI3K、p-AKT蛋白表达降低,经过不同治疗后4种蛋白表达均有不同程度地升高,三黄凝胶与PI3K/AKT激动剂单独应用的升高作用均强于积雪苷霜软膏,二者联合应用的升高作用最强;⑤结果表明,三黄凝胶可促进耳郭皮肤缺损创面的愈合,其作用机制可能与激活PI3K/AKT通路有关。

BACKGROUND:Due to the particularity of the ear structure,the skin at the auricle injury usually has poor motion range and tissue regeneration ability,and it is difficult to heal.Therefore,how to accelerate the wound healing and obtain an ideal repair effect has always been a problem and research hotspot in the field of surgery.OBJECTIVE:To explore the effects of Sanhuang gel on the wound healing,basic fibroblast growth factor and interleukin 17 levels of auricle skin defect in rabbits.METHODS:Sixty male New Zealand rabbits were randomly divided into six groups(n=10 per group).In group A,no treatment was given.In groups B-F,a circular skin defect area with the diameter of 1 cm was made in the auricle.In group B,the wound was injected with physiological saline.In group C,the wound was coated with asiaticoside cream ointment.In group D,the wound was coated with Sanhuang gel.In group E,the wound was injected with PI3K/AKT agonist 740 Y-P.In group F,the wound was coated with Sanhuang gel and injected with 740 Y-P.The treatment was performed twice a day,for 30 consecutive days.The wound healing of rabbit ear was observed and recorded,and the pathological morphology of ear tissue was detected by hematoxylin-eosin staining.The mRNA relative expression levels of basic fibroblast growth factor and interleukin 17 in rabbit ear tissue were detected by real-time PCR.The protein expression of PI3K/AKT pathway in rabbit ear tissue was detected by western blot assay.RESULTS AND CONCLUSION:(1)After treatment,the skin wound healing speed was accelerated.After 30 days of treatment,the wound healing rate of Sanhuang gel and PI3K/AKT agonist alone was higher than that of asiaticoside cream ointment,and the combined application of the two had the highest wound healing rate.(2)Hematoxylin-eosin staining results showed that the epidermis of group B was significantly thickened and blood vessels were dilated;and a large number of infiltrated inflammatory cells and fibrous tissue hyperplasia were seen.Compared with group B,the morphology of groups C,D,E,and F was significantly improved,and new capillaries and numerous proliferated fibroblasts and endothelial cells were seen.Among them,group F had the most obvious improvement.(3)RT-PCR detection showed that the relative expression of basic fibroblast growth factor mRNA decreased and the relative expression of interleukin 17 mRNA increased after auricular skin injury.After different treatments,the relative mRNA expression of the two factors was improved to varying degrees.The improvement effect of Sanhuang gel and PI3K/AKT agonist alone was stronger than that of asiaticoside cream ointment,and the improvement effect of their combination was strongest.(4)Western blot assay showed that the expression of PI3K,AKT,p-PI3K,and p-AKT proteins decreased after auricular skin injury.The expression levels of the four proteins increased to varying degrees after different treatments.The elevating effect of Sanhuang gel and PI3K/AKT agonist alone was stronger than that of asiaticoside cream ointment,and the elevating effect of their combination was strongest.(5)The results show that Sanhuang gel can promote the wound healing of auricular skin defects,and its mechanism may be associated with the activation of PI3K/AKT pathway.