羊脱细胞真皮基质的体内外毒性实验  

Toxicity studies in vivo and in vitro for sheep skin acellular dermal matrix

作　　者：张军[1] 黄晓元[2] 王凌峰[3] 

机构地区：[1]包头医学院第一附属医院烧伤整形科,内蒙古包头014010 [2]中南大学湘雅医院烧伤重建外科,长沙410008 [3]内蒙古包钢医院烧伤科,内蒙古包头014010

出　　处：《中南大学学报（医学版）》2016年第10期1069-1074,共6页Journal of Central South University ：Medical Science

基　　金：内蒙古自然科学基金(2008040MS1115)~~

摘　　要：目的：利用溶血反应、热原反应和细胞毒性反应实验了解羊脱细胞真皮基质（acellular dermal matrix,ADM）在动物体内外的毒性反应强度.方法：溶血反应：分别制作交联型和非交联型羊ADM的浸提液,为交联组和非交联组;并设置阳性对照组10mL灭菌注射用水,阴性对照组10 mL 0.9％氯化钠溶液.将新鲜抗凝稀释兔血加入实验组和对照组试管中,离心后取上清液测量吸光度值,计算溶血度.热原实验：按照条件入选16只新西兰兔,分为初试A,B两组各3只和复试C,D两组各5只.分别将交联与未交联的两种ADM浸提液注射人两组新西兰兔体内,注射后每隔0.5 h测量一次体温,共测量6次,以6次中最高一次减去正常体温即为该兔的体温升高度数.据此判断是否符合生物制品热原实验的评价标准.MTT实验：收集对数期的小鼠成纤维细胞细胞,种植于含有不同浓度羊ADM浸提液的培养基,测量吸光度值进行细胞活力测定.结果：实验A,B组溶血度均小于5％,热原反应测得新西兰兔体温升高之和低于1.8℃.MTT实验显示10％～90％羊ADM浸提液培养基对体外细胞的毒性为1级,100％的羊浸提液对体外细胞的毒性为2级,交联型与非交联型组的浸提液对体外细胞的毒性的无显著性差异（P＞0.05）.均对体外细胞的增殖影响轻微.结论：羊DM植入新西兰兔体内引起的溶血反应和热原反应在评价标准之内,在体外对细胞的增殖和活力影响不明显.Objective： To detect the toxic reaction degree for sheep acellular dermal matrix （ADM） in vivo or vitro by using hemolytic, pyrogen and cell-cytotoxic reaction experiments, respectively. Methods： Leach liquor of cross-linked and non-cross-linked sheep ADMs were set for cross-linked group and non-cross-linked group, respectively3 with a positive control group （10 mL sterile water for injection in test tube） and a negative control group （10 mL 0.9% sodium chloride solution in test tube）. The supernatants were obtained from each group and were measured for the absorbance. The hemolysis degree was calculated; 16 New-Zealand rabbits were selected and then divided into 4 groups, A, B, C and D group. The leach liquor of cross-linked and non-cross-linked sheep ADMs were injected into bodies of the 6 New-Zealand rabbits in the A and B groups, and then the body temperatures were measured in every half hour after injection, 6 times in total. The value of highest temperature among 6 measurements minus the normal temperature was the fever degree for the body temperature. Based on these fever degree, the criterion of biological pyrogen reaction for sheep ADM pyrogen experiment was evaluated; the mice fibroblasts were collected during logarithmic phase and were cultured in the nutrient medium containing sheep ADM leach liquor with different density. The absorbance was measured to evaluate relative growth rate for fibroblast. Results： The hemolysis degree for the group A and B are less than 5%. The summary of fever degree for New-Zealand rabbits were lower than 1.8 ℃. MTT experiment showed that the toxicity of 10%-90% or 100% leach liquor nutrient medium with sheep ADM for the mice fibroblast is at level 1 or level 2. There was no significant difference between leach liquor of cross-linked and non-cross- linked sheep ADMs （P〉0.05）. The effects on relative growth rate for mice fibroblasts were minor. Conclusion： The hemolytic and pyrogen reactions for the sheep ADMs embedded in New- Zealand rabbit were

关 键 词：羊皮 脱细胞真皮基质 溶血度 热原反应 相对增殖率 

分 类 号：R318.08[医药卫生—生物医学工程]