摘要
目的探讨玻璃化冻存组织工程肌腱植入大鼠体内修复跟腱缺损后不同时期,大鼠的免疫排斥反应及对肝、肾、心血管功能的影响。方法抽取1周龄SD大鼠尾腱,采用组织块法培养肌腱细胞。取第2～4代肌腱细胞,以5×106个/mL细胞密度接种至长1.5cm生物衍生肌腱材料,复合培养构建组织工程肌腱。采用21%DMSO作为玻璃化冻存保护液,Eurocollins solution作为4℃条件下预冷液和洗脱液的基础液体,按自制的玻璃化冻存方法冻存。健康SD大鼠72只,雌雄不限,体重210～230g。随机分成A(n=32)、B(n=32)、C(n=8)3组。A、B组实验动物于跟腱中段制备长约0.5cm肌腱缺损模型,分别将玻璃化冻存后及未冻存的组织工程肌腱移植修复缺损肌腱;C组大鼠未手术,为正常对照。术后2、4、6、8周,行大体观察、肝、肾及心血管功能检测;术后2、4、6周行血清免疫学检测。结果A、B组植入部位均未见组织坏死、积液及化脓感染;术后2周,A、B组均出现粘连现象,4～6周逐渐好转,8周未见明显粘连,新生组织连续性完整,肌腱桥接部已愈合,与正常肌腱类似。术后2、4、6周,血清中IgG和IgM含量A、B、C组组间比较差异均无统计学意义(P>0.05)。肝功能检测:A组术后4周,B组术后4、6周,谷草转氨酶均低于C组,差异有统计学意义(P<0.05);其余各时间点A、B组谷草转氨酶与C组比较差异均无统计学意义(P>0.05)。各时间点A、B组谷丙转氨酶、ALP、直接胆红素和总胆红素与C组比较差异均无统计学意义(P>0.05)。肾功能检测:术后2周,A组血清白蛋白和B组肌酐明显下降,与C组比较差异均有统计学意义(P<0.05);术后4、6、8周,A、B组各指标浓度与C组比较差异均无统计学意义(P>0.05)。心血管功能检测:A组各时间点血糖、甘油三酯、胆固醇与C组比较差异均无统计学意义(P>0.05);B组术后8周甘油三酯与C组比较差异有统计学意义(P<0.05)。结论玻璃化冻存的组织工程肌腱植入大鼠体内修复跟腱缺损,未引起明显免疫排斥反应,对肝、肾及心血管功能无明显影响。
Objective To study the immunological rejection occurred in different period after the in vivo implantation of vitreous-cryopreservation tissue engineered tendons for the repair of tendon defect and investigate its in？ uences on the hepatic,renal,and cardiovascular function of rats. Methods Tenocytes obtained from tail tendon of one-week-old SD rats were cultured in vitro. The tenocytes at passage 2-4 （5 ×10^6 cells/mL） were co-cultured with 1.5 cm bio-derived tendon material to reconstruct tissue engineered tendon. The 21% DMSO was used as cryopreservation protection solution and the Eurocollins solution served as basic solution for pre-frozen solution （4 ） and eluent. The cell-scaffold composites were vitreous-cryopreserved by self-designed method. Seventy-two healthy SD rats （male and/or female） weighing 210-230 g were randomly divided into three groups：group A （n=32）,group B （n=32）,and group C （n=8）. The 0.5 cm tendon defect model was established in the middle part of Achilles tendon in groups A and B. The defect in group A and B was repaired by the transplantation of tissue engineered tendon with and without vitreous-cryopreservation,respectively. At 2,4,6,and 8 weeks after transplantation,the general observation and the detection of hepatic function,renal function,and cardiovascular function were conducted. At 2,4,and 6 weeks after transplantation,serum immunology test was conducted. Results There were no tissue necrosis,hydrops,and suppurative infection in groups A and B. The adhesion was evident in groups A and B 2 weeks after transplantation,improved gradually during 4-6 weeks,and disappeared at 8 weeks. The neonatal tissue had full integration and continuity,and the bridging region of the tendon healed and was similar to the normal tendon. For serum IgG and IgM content,there was no signifi cant difference when group A or B was compared with group C,and between group A and group B 2,4,and 6 weeks after transplantation （P 〉 0.05）. Hepatic function：aspartate aminotransferase （AST） content of group A was less than that of group C 4 weeks after transplantation （P 〈 0.05）; AST content of group B was less than that of group C 4 and 6 weeks after transplantation （P 〈 0.05）; but there was no signifi cant difference when group A or B was compared with group C in terms of other indexes 8 weeks after transplantation （P 〉 0.05）. Renal function：serum albumin and creatinine in groups A and B were decreased obviously,and signifi cant difference was evident when compared with group C （P 〈 0.05）. Cardiovascular function：there was no signif icant difference between group A and group C in terms of blood glucose,triglyceride,and cholesterol （P 〉 0.05）; there was a signifi cant difference between group B and group C in terms of triglyceride 8 weeks after transplantation （P 〈 0.05）.Conclusion Repairing tendon defect with the implantation of vitreous-cryopreservation tissue engineered tendons results in no obvious immunological rejection and exerts no obvious in？ uences on hepatic,renal,and cardiovascular function.
出处
《中国修复重建外科杂志》
CAS
CSCD
北大核心
2009年第10期1229-1234,共6页
Chinese Journal of Reparative and Reconstructive Surgery
基金
国家自然科学基金资助项目(30570469)
四川省青年科技基金资助项目(06ZQ026-040)~~
关键词
组织工程肌腱
玻璃化冻存
体内植入
免疫排斥反应
大鼠
Tissue engineered tendon Vitreous-cryopreservation Implantation in vivo Immunological rejection Rat
