黄芩苷对大鼠压疮模型的干预作用及机制

Intervention Effect and Mechanism of Baicalin on Rat Pressure Ulcer Model

目的研究黄芩苷预防性和治疗性用药对大鼠压疮创面愈合的影响及对Th17、Treg细胞亚群的调节作用。方法通过磁力压迫局部组织缺血损伤法构建大鼠压疮模型,实验分为模型对照组(A组)、黄芩苷预防性用药组(B组)、黄芩苷治疗性用药(C组)和阳性对照组(D组)。采用HE染色观察创面皮肤和肌肉病理学变化,TUNEL法检测创面细胞凋亡情况,qPCR法检测创面EGF、VEGF、IL-17、IL-6、TGF-β和IL-10 mRNA水平,Western blot法检测脾脏RORγt及Foxp3蛋白表达,FCM法检测脾脏Th17、Treg细胞亚群比例变化,免疫组化法检测创面IL-17表达量。结果B、C、D组创面红肿、水泡、渗出、溃疡和坏死五种情况及细胞凋亡量较比A组有不同程度的改善(P<0.05);与A组比较,B、C和D组创面组织EGF、VEGF、TGF-β和IL-10 mRNA水平呈上升趋势(P<0.05),IL-17和IL-6水平呈下降趋势(P<0.05),且C组各因子变化幅度高于B组(P<0.05);B、C、D组RORγt蛋白表达较比A组有不同程度降低(P<0.05),Foxp3蛋白表达有小幅增加(P>0.05);B、C和D组脾脏Th17细胞比例较比A组呈下降趋势(P<0.01),且C组降低幅度高于B组(P<0.05),B、C和D组Treg细胞比例较比A组有小幅上升(P>0.05)。结论大鼠压疮模型Th17/Treg比例失衡诱导炎症反应发生,黄芩苷通过下调Th17细胞比例促进Th17/Treg恢复免疫平衡,进而降低炎症反应,促进创面愈合。

Objective To investigate the preventive and therapeutic effect of baicalin on wound healing and the regulation of Th17 and Treg cells subsets in pressure ulcer rats.Methods The rat models of pressure ulcer were established by magnetic compression of ischemic tissue injury,and then divided into model control group(Group A),preventive baicalin group(Group B),therapeutic baicalin group(Group C)and positive control group(Group D).HE staining was used to observe the pathological changes of the skin and muscle of the wound,and TUNEL assay was used to detect the apoptosis of the wound cells.The levels of EGF,VEGF,IL-17,IL-6,TGF-βand IL-10 mRNA of the wound were detected by qPCR.The expression of RORγt and Foxp3 protein in the spleen was detected by Western blot.FCM was used to detect the changes of the proportions of Th17 and Treg cells subsets in the spleen.The expression of IL-17 in the wound was detected by immunohistochemistry.Results Compared with Group A,the wound redness,blister,exudation,ulcer,necrosis and the amount of apoptosis in Group B,C and D were relieved to different degrees(P<0.05).The levels of EGF,VEGF,TGF-βand IL-10 mRNA in wound tissues of Group B,C and D increased(P<0.05),while the levels of IL-17 and IL-6 decreased,in comparison with Group A(P<0.05).The amplitude of variations of each factor in Group C was higher than that in Group B(P<0.05).Compared with Group A,the protein expression of RORγt in Group B,C and D decreased to different extents(P<0.05),while the protein expression of Foxp3 increased slightly(P>0.05).The proportions of Th17 cells in the spleen of Groups B,C and D were lower than that of Group A(P<0.01),and the proportion in Group C decreased more than that in Group B(P<0.05).There was a slight increase of Treg cells in Group B,C and D compared with Group A(P>0.05).Conclusion The imbalance of Th17/Treg cells subsets in rat models of pressure ulcer induces an inflammatory reaction.Baicalin inhibits the inflammatory response and promotes wound healing by promoting the restoration of immune balance of Th17/Treg through down-regulating the proportions of Th17 cells.