MEBO对糖尿病大鼠创面自噬信号因子Beclin1、LC3、p62的调控作用

目的分析湿润烧伤膏(MEBO)对创面组织自噬信号因子Beclin1、LC3、p62的调控作用及其促进创面愈合的机制。方法选取SPF级Wistar雄性大鼠250只,随机分为空白组、对照组、模型组、康复新组、MEBO组各50只。对照组建立常规创面模型,模型组、康复新组、MEBO组建立糖尿病创面模型,对照组、模型组大鼠创面予生理盐水外敷治疗,康复新组、MEBO组分别予康复新和MEBO外敷治疗。分别于干预第1、5、11、18、25天观察创面愈合情况、苏木素-伊红(HE)染色病理及自噬体的变化;免疫荧光法及Western印迹检测创面组织Beclin1、LC3、p62的变化;qRT-PCR分析Beclin1、LC3、p62 mRNA的表达。结果治疗过程中,对照组愈合最快,模型组愈合最慢,早期各组创面愈合率无显著差异(P>0.05),晚期各组创面愈合率:对照组>MEBO组>康复新组>模型组(P<0.05)。对照组、康复新组、MEBO组创面组织病理形态改善优于模型组。对照组、模型组、康复新组、MEBO组创面组织中Beclin1、LC3平均光密度(OD)值上升后下降,p62 OD值下降后上升;细胞内自噬体数目升高后下降;创面组织Beclin1、p-Beclin1表达量及LC3Ⅱ/LC3Ⅰ比值上升后下降,p62表达下降后上升;Beclin1、LC3 mRNA表达上升后下降,p62表达下降后上升差异均有统计学意义(均P<0.05)。结论MEBT/MEBO可有效促进糖尿病创面愈合,细胞自噬激活可能是其作用机制之一。

Regulation of autophagy signaling factors Beclin1, LC3 and p62 in diabetic rats by MEBO

Objective To observe the effect of Moist Exposed Burn Ointment(MEBO)on the expression of Beclin1,LC3 and p62 of autophagy signals in wound tissues,and to explore the mechanism of MEBO promoting wound healing.Methods Two hundred and fifty male SPF-grade Wistar rats were randomly divided into blank group,control group,model group,Kangfuxin group and MEBO group,with 50 rats in each group.Conventional wound models were established in the control group,while diabetic wound models were respectively established in the model group,Kangfuxin group and MEBO group.The wounds of rats in the control group and model group were treated with normal saline,while the wounds of Kangfuxin group and MEBO group were treated with Kangfuxin and MEBO,respectively.The wound healing,and the changes of HE staining pathology and autophagosomes were observed on day 1,day 5,day 11,day 18 and day 25 after treatment.Meanwhile,the changes of Beclin1,LC3 and p62 in wound tissues were detected by immunofluorescence assay and Western blotting.The expressions of Beclin1,LC3 and p62 mRNA were analyzed by qRT-PCR.Results During the treatment:the healing rate was the fastest in the control group and the slowest in the model group,and there was no significant difference in the wound healing rate of each group in the early stage(P>0.05).Wound healing rate of each group in the late stage:control group>MEBO group>Kangfuxin group>model group(P<0.05).The improvement of wound histopathological morphology in the control group,Kangfuxin group and MEBO group was better than that in model group.The OD values of Beclin1 and LC3 in wound tissues of the control group,model group,Kangfuxin group and MEBO group increased and then decreased,while that of p62 increased after decreasing(P<0.05).The number of intracellular autophagosomes in the control group,model group,Kangfuxin group and MEBO group increased and then decreased(P<0.05).The expressions of Beclin1,p-Beclin1 and the LC3Ⅱ/LC3Ⅰratio in wound tissues of the control group,model group,Kangfuxin group and MEBO group decreased after increasing,and the expression of p62 increased after decreasing(P<0.05).Beclin1 and LC3 mRNA expression increased and then decreased in the control group,model group,Kangfuxin group and MEBO group,and p62 mRNA expression decreased and then increased(P<0.05).Conclusions MEBO could effectively promote wound healing in diabetes mellitus,and autophagy activation may be one of its mechanisms.