小鼠烧伤坏死真皮基质的制备方法及生物学评价  

Preparation and biological assessment of burned-necrotic dermal matrix in mouse

作　　者：于冠英[1,2,3] 李川[2,3] 朱绪国[2,3] 王晓川[2,3] 李耀南[2,3] 姜笃银[2,3] 

机构地区：[1]山东大学附属济南市中心医院,山东济南250013 [2]山东大学组织工程研究所 [3]山东大学第二医院整形烧伤科,山东济南250033

出　　处：《感染．炎症．修复》2013年第3期146-149,F0003,共5页Infection Inflammation Repair

基　　金：国家自然科学基金资助项目(81071560);山东省重点攻关项目(2009GG10002078)

摘　　要：目的:探讨小鼠烧伤坏死真皮基质(BNDM)制备方法,并对其生物学性能进行评价。方法:水浴烫伤法制备昆明小鼠Ⅲ度烫伤模型,3 d后取烫伤部位皮肤,以0.25%Trypsin-0.5%TritonX-100溶液洗脱处理,制成BNDM。以正常小鼠脱细胞真皮基质(ADM)作对照,HE染色及DAPI染色后观察洗脱效果及胶原排列情况。将ADM和BNDM分别埋植入小鼠皮下,于埋植后1、2、3、4周观察真皮基质的外观及其与周围组织的关系,以及ADM和BNDM中胶原排列、细胞浸润及血管生成情况。结果:BNDM质地较ADM柔软,有一定的柔韧性和延展性。HE染色观察BNDM的胶原纤维结构模糊,呈均质玻璃样改变,嗜伊红性增强,经DAPI染色证实基质内无染色质成分存留。皮下埋植试验显示BNDM和ADM均无明显排斥反应和毒副作用。显微镜下观察埋植术后第1周即可见到ADM及BNDM中有成纤维细胞长入并有炎性细胞浸润。埋植后第3周,细胞浸润更多并可见毛细血管长入,较其他时间点更明显,而ADM较BNDM更明显(P<0.05);至埋植后第4周,炎症反应减轻。结论:烧伤坏死皮肤经Trypsin-TritonX-100溶液处理后,可以保持胶原纤维的连续性,无明显的不良反应,成纤维细胞和炎性细胞可以很好地浸润生长,并有毛细血管长入,能满足异体或自体皮肤移植的基本要求。Objective To investigate the preparation and assess the biological property of mouse burned-necrotic dermal matrix （BNDM）. Methods. Full-thickness thermal injury model was replicated in Kunming mice by dipping the back of the mice into heated water of 95 ℃ for 8 seconds, and the burned skin was harvested 3 days later. Acellular dermal matrix （ADM） of normal skin of mouse was prepared as control. The BNDM was treated with 0.25~/oo trypsin and 0.5~ TritonX-100 solution. The arrangement of collagen in BNDM and ADM was observed with hematoxylin-eosin （HE） and 4＇,6-diamidino-2-phenylindole （DAPI） staining. ADM and BNDM was implanted under the dorsal skin of mice . At the end of 1, 2,3,4 weeks after implantation, the arrangement of collagen, cellular infiltration and the angiogenesis were assessed after HE staining. Results.. It was found that the BNDM was softer than ADM, with certain degree of flexibility and extensibility. The loosely arranged collagen fibers were found to be eosin-staining enhanced with obscure structure without cells under the microscope. No rejection or un toward effects were observed after either AMD or BNDM implantation. Infiltration of fibroblasts and inflammatory cells were observed 1 week after implantation in both ADM and BNDM, and the cell number peaked 3 weeks later and decreased 4 weeks after implantation. More capillary growth and cellular infiltration were observed in ADM than in BNDM 3 weeks after implantation （P^0.05）. Conclusions：The burned-necrotic skin could be processed by trypsin-TritonX-100 solution to satisfy the need of allogeneic or even autogeneic skin graft with preservation of col- lagenous structure, and infiltration of fibroblasts and inflammatory cells, and angiogenesis without untoward effect.

关 键 词：烧伤坏死真皮基质 深度烧伤 生物相容性 

分 类 号：R622.1[医药卫生—整形外科]