鱼鳞膏调节IL-6/STAT3信号通路对难愈性皮肤溃疡模型大鼠创面愈合的影响

Impact of Fish Scale Ointment on wound healing in rats with refractory skin ulcers by regulating the IL-6/STAT3 signaling pathway

目的初步探讨鱼鳞膏调节白细胞介素-6(IL-6)信号转导与转录激活因子3(STAT3)信号通路对难愈性皮肤溃疡模型大鼠创面愈合的影响。方法高脂高糖喂养联合腹腔注射链脲佐菌素(STZ)建立糖尿病大鼠模型,手术剪去背部皮肤构建难愈性皮肤溃疡大鼠模型;造模成功后按照随机数字表法分为模型组、鱼鳞膏组、激活剂组,每组12只,另外取12只大鼠作对照组(模型对照),各组贴敷和腹腔注射相应药物,每天1次,连续21 d;给药结束后测量大鼠创面愈合情况;HE染色观察创面损伤及新生毛细血管情况;酶联免疫吸附法(ELISA)检测大鼠胎盘创面组织中白细胞介素-6(IL-6)、白细胞介素-1β(IL-1β)、肿瘤坏死因子α(TNF-α)水平;免疫组化染色测定碱性成纤维细胞生长因子(bFGF)、血小板-内皮细胞黏附分子-1(CD31)表达;实时荧光定量PCR(qRT-PCR)法检测创面组织中IL-6与STAT3 mRNA表达;蛋白免疫印迹法(Western blotting)检测大鼠创面组织中血管内皮生长因子(VEGF)、转化生长因子-β1(TGF-β1)及IL-6/STAT3通路相关蛋白的表达。结果与对照组比较,模型组大鼠创面愈合率、新生血管数及bFGF、CD31、VEGF、TGF-β1蛋白表达水平显著降低,IL-6、IL-1β、TNF-α水平、IL-6、STAT3 mRNA及蛋白表达水平显著升高(P<0.05);与模型组比较,鱼鳞膏组大鼠创面愈合率、新生血管数及bFGF、CD31、VEGF、TGF-β1蛋白表达水平显著升高,IL-6、IL-1β、TNF-α水平、IL-6、STAT3 mRNA及蛋白表达水平显著降低(P<0.05);与鱼鳞膏组比较,激活剂组大鼠上述指标变化均显著逆转(P<0.05)。结论鱼鳞膏具有抑制炎症反应,促进难愈性皮肤溃疡大鼠血管形成及创面愈合的作用,其作用机制可能与抑制IL-6/STAT3信号通路有关。

Objective To explore the impact of Fish Scale Ointment on wound healing in rats with refractory skin ulcer by regulating interleukin-6(IL-6)/signal transducer and activator of transcription 3(STAT3)signaling pathway.Methods The model of diabetes rats was established by feeding with high fat and high sugar and intraperitoneal injection of streptozotocin(STZ),and the model of refractory skin ulcer was established by removing the back skin through surgical scissors;After successful modeling,the rats were randomly divided into model group,Fish Scale Ointment group and activator group according to the random number table method,with 12 rats in each group.Another 12 rats were taken as the control group(model control),and the corresponding drugs were applied and injected intraperitoneally in each group,once a day for 21 days;The wound healing status of rats was measured after administration;HE staining was applied to observe wound damage and neovascularization;Enzyme linked immunosorbent assay(ELISA)was applied to detect the levels of interleukin-6,-1β(IL-6,IL-1β)and tumor necrosis factor-α(TNF-α)in placental wound tissue;Immunohistochemical staining was applied to determine the expression of basic fibroblast growth factor(bFGF)and platelet endothelial cell adhesion molecule 1(CD31);Real-time fluorescence quantitative PCR(qRT-PCR)method was applied to detect the expression of IL-6 and STAT3 mRNA in wound tissue;And Western blotting was applied to detect the expression of vascular endothelial growth factor(VEGF),transforming growth factor-β1(TGF-β1)and IL-6/STAT3 pathway related proteins in rat wound tissues.Results Compared with the control group,the wound healing rate,number of new blood vessels,and the expression levels of bFGF,CD31,VEGF,and TGF-β1 proteins in the model group were obviously reduced,the levels of IL-6,IL-1β,TNF-α,and the expression levels of IL-6,STAT3 mRNAs and proteins were obviously increased(P<0.05).Compared with the model group,the wound healing rate,number of new blood vessels,and the expression levels of bFGF,CD31,VEGF,and TGF-β1 proteins in the Fish Scale Ointment group were obviously increased,the levels of IL-6,IL-1β,TNF-α,and the expression levels of IL-6,STAT3 mRNAs and proteins were obviously reduced(P<0.05);Compared with the Fish Scale Ointment group,the changes in the above indicators in the activator group were obviously reversed(P<0.05).Conclusion Fish Scale Ointment has the effects of inhibiting inflammatory response,promoting angiogenesis and wound healing in rats with refractory skin ulcers,and its mechanism of action may be related to the inhibition of IL-6/STAT3 signaling pathway.