微孔化羊ADM复合hUCMSC促进创面修复的机制研究

目的探讨微孔化羊脱细胞真皮基质(ADM)复合人脐带间充质干细胞(hUCMSC)促进全层损伤皮肤创面愈合的治疗机制。方法将hUCMSC种植于微孔化羊ADM上共培养,形成复合生物敷料。选用72只裸鼠制作成皮肤全层损伤模型,按随机数字表法分为3组,每组24只,hUCMSC种植于微孔化羊ADM上治疗全层损伤创面组(hUCMSC+微孔化羊ADM组);羊ADM治疗全层损伤组(羊ADM组);碘伏纱布治疗全层损伤创面组(碘伏纱布组)。术后14、21、28 d检测各组裸鼠创面愈合率,运用qRT-PCR技术检测创面组织中Bax和Bcl-2的mRNA的表达,免疫组织化学染色检测创面愈合相关蛋白细胞胶原蛋白Ⅰ和血管内皮生长因子(VEGF)的表达。数据比较采用单因素方差分析和t检验。结果术后14 d,hUCMSC+微孔化羊ADM组的愈合率为(65.34±14.72)%,明显高于碘伏纱布组(37.54±10.21)%],也高于羊ADM组(49.08±11.16)%],差异均有统计学意义(t=19.52、14.72,P<0.05)。随着损伤创面的逐步愈合,术后28 d,hUCMSC+微孔化羊ADM组的愈合率为(98.63±15.41)%,明显高于羊ADM组(81.74±16.27)%]和碘伏纱布组(63.47±14.80)%],差异均有统计意义(t=-16.42、20.35,P<0.05)。Bax在hUCMSC+微孔化羊ADM组的创面组织中的表达显著降低,尤其在术后21 d的表达量为0.25±0.06,明显低于碘伏纱布组(0.53±0.16)和羊ADM组(0.41±0.12),差异均有统计学意义(t=3.52、-2.83,P<0.05)。Bcl-2在hUCMSC+微孔化羊ADM组的表达显著高于其他2组,尤其是在术后21 d的表达量为0.63±0.19,明显高于碘伏纱布组(0.34±0.09)和羊ADM组(0.46±0.13),差异均有统计学意义(t=5.31、-6.07,P<0.05)。免疫组织化学技术检测发现hUCMSC+微孔化羊ADM组胶原蛋白Ⅰ和VEGF的表达深棕色细胞虽然比羊ADM组和碘伏纱布组稍微有所增多,但是效果并不显著。结论hUCMSC+微孔化羊ADM复合敷料,通过携带hUCMSC更能促进皮肤全层损伤的愈合并减少凋亡细胞的产生。

Study on the mechanism of microporous sheep ADM combined with hUCMSC in promoting wound healing

Objective To explore the therapeutic mechanism of microporous sheep acellular dermal matrix(ADM)combined with human umbilical cord mesenchymal stem cell(hUCMSC)to promote the healing of full-thickness wounds.Methods hUCMSC was co-cultured on microporous sheep ADM to form composite biological dressings.Seventy-two nude mice were selected to make a full-thickness skin injury model and randomly divide them into 3 groups,each group of 24.hUCMSC+microporous sheep ADM group,sheep ADM group and iodophor gauze group.The wound healing rate of each group was detected at 14,21 and 28 days after operation,qRT-PCR technique was used to detect the expression of Bax and Bcl-2 in wound tissue,and immunohistochemical technique was used to detect the expression of wound healing related proteins.Data were analyzed with one-way analysis of variance and t test.Results On the 14 days after operation,the healing rate of the hUCMSC+microporous sheep ADM group was(65.34±14.72)%,which was significantly higher than that of the Iodophor gauze group(37.54±10.21)%,and also higher than that of the sheep ADM group(49.08±11.16)%],the differences were statistically significant(t=19.52,14.72;P<0.05).With the gradual healing of the wound,the healing rate of the hUCMSC+microporous sheep ADM group was(98.63±15.41)%at 28 days after operation,which was significantly higher than that of the sheep ADM group(81.74±16.27)%]and the Lodophor gauze group(63.47±14.80)%],the differences were statistically significant(t=-16.42,20.35;P<0.05).The expression of Bax in the wound tissue of the hUCMSC+microporous sheep ADM group was significantly reduced,especially on the 21 days after operation,the expression level was 0.25±0.06,which was significantly lower than the Lodophor gauze group(0.53±0.16)and sheep ADM group(0.41±0.12),the differences were statistically significant(t=3.52,-2.83;P<0.05).The expression of Bcl-2 in the hUCMSC+microporous sheep ADM group was significantly higher than that of the other two groups,especially at the 21 days after operation postoperative day,the expression level was 0.63±0.19,which was significantly higher than the Lodophor gauze group(0.34±0.09)and sheep ADM group(0.46±0.13),the differences were statistically significant(t=5.31,-6.07;P<0.05).Immunohistochemical detection showed that the expression of CollagenⅠand VEGF in hUCMSC+microporous sheep ADM group was slightly more than that of sheep ADM group and iodophor gauze group,but the effect was not significant.Conclusion hUCMSC+microporous sheep ADM composite dressing can promote the healing of full-layer skin injury and reduce the production of apoptotic cells by carrying hUCMSC.