象皮生肌膏对糖尿病足溃疡大鼠创面组织内上皮-间质转化过程的影响

Effects of Xiangpi Shengji ointment on EMT in wound tissues of diabetic ulcer rats

目的:观察象皮生肌膏对糖尿病足溃疡大鼠创面组织内转化生长因子-β1(TGF-β1)、胰岛素样生长因子-1(IGF-1)、纤维结合蛋白(FN)表达的影响,以及在愈合过程中上皮-间质转化(EMT)过程。方法:选择SPF级雄性SD大鼠40只,建立溃疡大鼠模型,其中10只作为空白组,其余30只大鼠建立糖尿病大鼠模型,造模成功后分为模型组、对照组、实验组。比较各组大鼠第7、14天创面愈合情况;实时荧光定量聚合酶链式反应法(RT-qPCR)检测各组第7、14天TGF-β1、IGF-1、FN的mRNA表达情况;Western blotting检测各组第7、14天EMT相关蛋白E-钙黏蛋白(E-cadherin)、纤连蛋白(Fibronectin)蛋白表达情况;酶联免疫反应法测定组织中血管内皮生长因子(VEGF)含量。结果:干预第7天,空白组、实验组的创面愈合率均高于模型组;与模型组相比,空白组、实验组E-cadherin表达降低,Fibronectin表达升高,而TGF-β1、IGF-1、FN的mRNA表达均高;与对照组相比,实验组E-cadherin蛋白表达降低,Fibronectin蛋白表达升高;与模型组相比,空白组、对照组、实验组的VEGF含量均高,差异有统计学意义(均P<0.05)。干预第14天,模型组的创面愈合率低于其他组;与模型组相比,空白组、实验组E-cadherin蛋白表达较低,Fibronectin蛋白表达较高,空白组、对照组、实验组FN mRNA表达均高,空白组、实验组TGF-β1的mRNA表达均高。与模型组相比,空白组、对照组、实验组的VEGF含量均高,差异有统计学意义(均P<0.05)。干预第14天,模型组、对照组、实验组的IGF-1的mRNA表达比较,差异无统计学意义(均P>0.05)。结论:象皮生肌膏能促进EMT过程,加快愈合速度,其途径可能是通过促进TGF-β1表达,介导IGF-1,调控FN实现。

Objective:To observe expression of TGF-β1,IGF-1,FN in wound tissue of diabetic ulcer rats with Xiangpi Shengji ointment,and to explore effect of healing process on epithelial-mesenchymal transition(EMT).Methods:40 male SD rats of SPF grade and establish ulcer rat models.Ten of rats were used as the blank group,and the other 30 rats used to establish diabetic rat models.The 30 rats were successfully divided into model group,control group and experimental group.Compared the wound healing of rats in each group on the 7th and 14th day.Real-time quantitative polymerase chain reaction(RT-qPCR)was used to detect the mRNA expressions of TGF-β1,IGF-1 and FN on the 7th and 14th day in each group.Western blotting was used to detect the protein expression of EMT-related proteins,E-cadherin and Fibronectin on the 7th and 14th day in each group.ELISA was used to measure the content of vascular endothelial growth factor(VEGF)in tissues.Results:On the 7th day of intervention,the wound healing rate of the blank group and the experimental group were significantly higher than those in the model group.On the 7th day of intervention,compared with the model group the expression of E-cadherin in the blank group and the experimental group decreased,expression of Fibronectin increased,and the mRNA expressions of TGF-β1,IGF-1 and FN increased.On the 7th day of intervention,compared with the control group the expression of E-cadherin protein decreased and the expression of Fibronectin protein increased in the experimental group.On the 7th day of intervention,compared with the model group VEGF content of the blank group,the control group and the experimental group were all higher,difference statistically significant(all P<0.05).On the 14th day of intervention,the wound healing rate of the model group was lower than that of the other three groups.On the 14th day of intervention,compared with model group expression of E-cadherin protein in the blank group and the experimental group decreased,but expression of Fibronectin protein increased,expression of FN mRNA in the other three groups increased,expression of TGF-β1 mRNA in the blank group and the experimental group increased.On the 14th day of intervention,compared with the model group the content of VEGF in the blank group,the control group and the experimental group higher,difference statistically significant(all P<0.05).There was no significant difference in IGF-1 mRNA expression among the other three groups except the blank group(all P>0.05).Conclusion:Xiangpi Shengji ointment can promote the EMT process and accelerate the healing speed.It may be achieved by promoting the expression of TGF-β1 and mediating the regulation of IGF-1 on FN.