MEBT/MEBO对慢性难愈合创面组织中CK15表达水平的影响

Influence of MEBT/MEBO on the Expression of CK15 in Chronic Refractory Wound Tissues

目的通过检测大鼠慢性难愈合创面组织中细胞角蛋白(cytokeratin,CK)15的动态表达水平,探讨皮肤再生医疗技术(moist exposed burn therapy/moist exposed burn ointment,MEBT/MEBO)促进慢性难愈合创面愈合的部分作用机制。方法选取SPF级健康成年雄性Wistar大鼠90只适应性喂养7 d后,按照随机数表法随机分为空白组、对照组、模型组、MEBO组与rb-bFGF组,每组18只,其中空白组大鼠只做背部备皮处理,对照组大鼠于背部建立急性创面模型,模型组、MEBO组和rb-bFGF组大鼠于背部建立慢性难愈合创面模型;模型建立后,空白组大鼠备皮处皮肤及对照组、模型组大鼠创面予以0.9%氯化钠注射液纱布湿敷,MEBO组大鼠创面予以湿润烧伤膏(moist exposed burn ointment,MEBO)药纱换药治疗,rb-bFGF组大鼠创面予以重组牛碱性成纤维细胞生长因子(recombinant bovine basic fibroblast growth factor,rb-bFGF)药纱换药治疗。分别于治疗第3、7、14天,采用Western blotting技术及Real-time PCR技术检测各组大鼠皮肤或创面组织中CK15蛋白及CK15mRNA的表达水平。结果空白组大鼠皮肤组织中CK15蛋白及CK15 mRNA表达水平无明显变化(F=0.054、1.333,P=0.948、0.293),其他各组大鼠创面组织中CK15蛋白及CK15 mRNA表达水平均呈先降低后升高的趋势(CK15蛋白:F=192.766、58.065、126.888、118.244,P均=0.000;CK15 mRNA:F=580.869、272.569、569.254、736.054,P均=0.000)。治疗第3、7天,对照组、MEBO组、rb-bFGF组大鼠创面组织中CK15蛋白及CK15 mRNA表达水平均显著低于模型组(P均<0.05),而治疗第14天对照组、MEBO组、rb-bFGF组大鼠创面组织中CK15蛋白及CK15 mRNA表达水平均显著高于模型组(P均<0.05);治疗第3、7、14天,MEBO组与rb-bFGF组大鼠创面组织中CK15蛋白及CK15 mRNA表达水平均无明显差异(P均>0.05)。结论通过调控创面组织中CK15的表达水平促进表皮干细胞的增殖可能是MEBT/MEBO加快慢性难愈合创面愈合的部分作用机制,且其治疗效果与rb-bFGF相当,作用机制与rb-bFGF相似。

Objective To explore the mechanism of action of Moist Exposed Bum Therapy/Moist Exposed Burn Ointment(MEBT/MEBO)in promoting the healing of chronic refractory wound through detecting the dynamic expression of cytokeratin(CK)15 in chronic refractory wound tissues of rats.Methods Ninety healthy SPF Wistar male rats were selected as subjects to be fed for 7 days to adapt surrounding environment and then divided,according to the random number table,into blank group,control group,model group,MEBO group and rb-bFGF group,with 18 rats in each group.For rats in blank group,only back skin preparation was performed.For rats in control group,acute wound models were established.For rats in model group,MEBO group and rb-bFGF group,chronic refractory wound models were established on their backs.After the models were established,gauze with 0.9%sodium chloride injection was applied to the skin preparation sites of blank group and wounds of control group and model group,MEBO dressing change was performed on the wounds of MEBO group and rb-bFGF dressing change was performed on wounds of the rb-bFGF group.On day 3,day 7 and day 14 of treatment,the expression levels of CK15 and CK15 mRNA in the skin or wound tissues of each group were detected with Western blotting and Real-time PCR.Results No significant change was observed in the expression levels of CK15 and CK15 mRNA in skin tissues of blank group(F=0.054,1.333,P=0.948,0.293).The expression levels of CK15 and CK15 mRNA in the wound tissues of other groups showed a trend of increase first followed by decrease(CK15:F=192.766,58.065,126.888,118.244,all P=0.000;CK15 mRNA:F=580.869,272.569,569.254,736.054,all P=0.000).On day 3 and day 7 of treatment,the expression levels of CK15 and CK15 mRNA in the wound tissues of control group,MEBO group,and rb-bFGF group were significantly lower than those of the model group(all P<0.05)while on day 14 of treatment,the expression levels of CK15 and CK15 mRNA in wound tissues of control group,MEBO group and rb-bFGF group were significantly higher than those of model group(all P<0.05).On day 3,day 7,and day 14 of treatment,no significant difference was observed between MEBO group and rb-bFGF group(all P>0.05)in the expression levels of CK15 and CK15 mRNA in the wound tissues.Conclusion MEBT/MEBO can promote the proliferation of ESCs by regulating the expression level of CK15 in tissue cells.This may constitute its mechanism of action in accelerating the healing of chronic refractory wounds and its therapeutic effect and mechanism of action are similar to rb-bFGF.