人脐带间充质干细胞在糖尿病大鼠皮肤创面中的应用

Application of human umbilical cord-mesenchymal stem cells in skin wound of diabetic rats

目的:探讨人脐带间充质干细胞(hUC-MSCs)治疗糖尿病大鼠创面的安全性和有效性及相应机制。方法:24只雄性SD大鼠采用完全随机分组方法分为hUC-MSCs组和磷酸缓冲盐溶液(PBS)组,每组12只,在其背部形成一全层皮肤缺损创面模型,分别于创缘注射hUC-MSCs悬液和PBS液。于3、7、14 d观察创面愈合,苏木精-伊红(HE)染色观察病理学改变,免疫组织化学及蛋白免疫印迹(Western blot)检测血管内皮细胞生长因子(VEGF)蛋白的表达。组间比较采用t检验。结果:hUC-MSCs组第3天时创面愈合率为(20.12±4.50)%,PBS组创面愈合率为(13.85±3.89)%。第7天hUC-MSCs组创面持续缩小,创面愈合率为(61.26±9.25)%,PBS组创面愈合率为(40.23±8.30)%。第14天hUC-MSCs组创面基本愈合,创面愈合率为(93.28±4.27)%,PBS组创面愈合率为(81.56±3.89)%。hUC-MSCs组创面愈合率在各时间点(3、7、14 d)均较PBS组显著增高(P<0.05)。HE染色显示hUC-MSCs组创面愈合质量(皮肤厚度、弹力纤维密度等)优于PBS组。免疫组织化学血管染色显示hUC-MSCs组创面新生组织新生血管数量及血管密度均较PBS组高。免疫组织化学显示hUC-MSCs组VEGF在3 d时表达最多,7、14 d逐渐减少,但均较PBS组增高(P<0.05)。Western blot结果显示hUC-MSCs组3、7、14 d创面VEGF蛋白表达显著高于PBS组(P<0.05)。结论:hUC-MSCs治疗糖尿病大鼠创面安全有效,可通过增加新生血管数量,促进皮肤创面的修复。

Objective To study the safety and effectiveness of human umbilical cord-mesenchymal stem cells(hUC-MSCs)in treating diabetic rat wounds and the corresponding mechanisms.Methods Twenty four male SD rats were randomly divided into hUC-MSCs group and phosphate buffered saline(PBS)group,with 12 rats in each group.A full-thickness skin defect wound model was performed on their back,and hUC-MSCs suspension and PBS solution were injected at the wound margin respectively.The wound healing was observed at 3,7 and 14 days,the pathological changes were observed by hematoxylin and eosin(HE)staining,and vascular endothlial growth factor(VEGF)protein was detected by immunohistochemistry and Western blot.SPSS 20.0 statistical software was used for analysis.The comparison between groups was performed by t-test.The difference was statistically significant if P<0.05.Results The wound healing rate was(20.12±4.50)%in hUC-MSCs group and(13.85±3.89)%in PBS group on the third day.On the 7th day,the wound of hUC-MSCs group continued to shrink,and the wound healing rate was(61.26±9.25)%,while that of PBS group was(40.23±8.30)%.On the 14th day,the wound healing rate of hUC-MSCs group was(93.28±4.27)%,and that of PBS group was(81.56±3.89)%.The wound healing rate in hUC-MSCs group was significantly higher than that in PBS group at all time points(3,7,14 d)(P<0.05).HE staining showed that the wound healing quality(skin thickness,elastic fiber density,etc)of hUC-MSCs group was better than that of PBS group.Immunohistochemistry showed that the expression of VEGF in hUC-MSCs group was the highest at 3 days,decreased gradually at 7 and 14 days,but was higher than that in PBS group(P<0.05).Western blotting results showed that the expression of VEGF protein in the wounds of hUC-MSCs group was significantly higher than that of PBS group on the 3rd,7th and 14th day(P<0.05).Conclusion hUC-MSCs are safe and effective in treating diabetic rat wounds,which can promote the repair of skin wounds by increasing the number of new blood vessels.