去细胞异种神经复合同种异体脂肪干细胞修复猕猴周围神经缺损的免疫反应研究  

STUDY ON IMMUNE RESPONSE AFTER REPAIR OF NERVE DEFECT WITH ACELLULAR NERVE XENOGRAFT LADEN WITH ALLOGENIC ADIPOSE-DERIVED STEM CELLS IN RHESUS MONKEY

作　　者：黄喜军[1] 朱庆棠[1] 江丽[2] 郑灿镔[1] 朱昭炜[1] 路庆森[1] 许银峰[3] 顾立强[1] 刘小林[1] 

机构地区：[1]中山大学附属第一医院显微创伤骨科,广州510080 [2]中山大学附属第一医院骨科研究所,广州510080 [3]中山大学附属第一医院黄埔院区上肢骨科,广州510080

出　　处：《中国修复重建外科杂志》2012年第8期993-1000,共8页Chinese Journal of Reparative and Reconstructive Surgery

基　　金：国家自然科学基金资助项目(30700847;31070869);广东省自然科学基金资助项目(9251008901000017);广东省科技计划资助项目(2010B031100006)~~

摘　　要：目的观察去细胞异种神经复合同种异体脂肪干细胞修复猕猴周围神经缺损后的全身及局部免疫排斥反应,评价该修复材料的安全性。方法健康成年雄性长白猪1只,体重48 kg,取胫神经制备去细胞异种神经;健康成年雄性猕猴1只,体重4.5 kg,分离培养脂肪干细胞;健康成年雌性猕猴10只,体重3～5 kg,制备25 mm长桡神经缺损动物模型,随机分为复合细胞组和无细胞组(n=5),前者采用去细胞异种神经复合第3代脂肪干细胞移植修复,后者采用去细胞异种神经移植修复。于术前及术后14、60、90 d抽取外周静脉血行淋巴细胞分析;术后5个月取材观察移植物组织免疫反应和神经再生情况,并与自体神经移植组作比较。结果复合细胞组与无细胞组术前及术后各时间点外周静脉血淋巴细胞数量、T淋巴细胞百分率及其数量、CD8+T淋巴细胞百分率、CD4+/CD8+T淋巴细胞比值间比较,差异均无统计学意义(P>0.05);术后14 d,复合细胞组CD4+T淋巴细胞百分率低于其余时间点,差异有统计学意义(P<0.05);同一时间点两组间比较,除术后14 d复合细胞组CD4+T淋巴细胞百分率低于无细胞组,差异有统计学意义(P<0.05)外,其余各时间点各指标组间比较差异均无统计学意义(P>0.05)。术后5个月,去细胞异种神经与周围组织有轻度粘连,神经外膜较自体神经厚,未见组织坏死、纤维瘢痕形成,移植物内见再生神经纤维,复合细胞组、无细胞组均有稀疏的CD3+、CD4+、CD8+、CD68+、CD163+T淋巴细胞散在分布,细胞浸润情况与自体神经移植组类似。结论去细胞异种神经移植修复猕猴周围神经缺损后未产生全身及局部免疫排斥反应;复合同种异体脂肪干细胞移植后也未发现免疫排斥反应,且术后早期可能抑制CD4+T淋巴细胞增殖。Objective To observe the systemic and local immune response after repair of nerve defect with acellular nerve xenograft laden with allogenic adipose-derived stem cells （ADSCs） in rhesus monkey so as to evaluate the safety of the proposed material for nerve reconstruction. Methods Bilateral tibial nerves were taken from a healthy adult male landrace （weighing 48 kg） to prepare acellular nerve xenograft by chemical extraction. ADSCs were isolated from a healthy adult male rhesus monkey （weighing 4.5 kg）, and were seeded into the acellular nerve grafts. The radial nerve defect models with 25 mm in length were established in 10 healthy adult female rhesus monkeys （weighing 3-5 kg）, and they were divided into cell-laden group （n=5） and non-cell-laden group （n=5） randomly. Defect was repaired with acellular nerve xenograft laden with allogenic ADSCs in cell-laden group, with acellular nerve xenograft only in non-cell-laden group. The blood samples were taken from peripheral vein preoperatively and at 14, 60, and 90 days after operation for lymphocyte analysis; at 5 months after operation, the grafts were harvested to perform histological examination for local immune response and nerve regeneration. The nerve autograft in rhesus monkey was used as control. Results In cell-laden group and non-cell-laden group, no significant difference was found in the count oflymphocytes and T lymphocytes, the percentage ofT lyrnphocytes, CD84^＋ T lymphocytes, as well as the ratio of CD4^＋ T lymphocytes to CD84^＋ T lymphocytes between pre- and post-operation （P 〉 0.05）; in cell-laden group, the percentageof CD4＋ T lymphocytes at 14 days was significantly lower than that at 60 and 90 days postoperatively （P 〈 0.05）. The percentage of CD44^＋ T lymphocytes in cell-laden group was significantly lower than that in non-cell-laden group at 14 days （P 〈 0.05）, but no significant difference was found in the other indexes at the other time between 2 groups （P 〉 0.05）. At 5 months after ope

关 键 词：去细胞异种神经 脂肪干细胞 异种移植 同种异体移植 免疫反应 猕猴 猪 

分 类 号：R651.3[医药卫生—外科学]