羟基荧光素二醋酸盐琥珀酰亚胺脂标记大鼠肝脏NK细胞体内转输示踪  

Tracing of CFSE labeled donor hepatic NK cells in isogenic rats transfused via portal vein

作　　者：龙天柱[1] 商昌珍[1] 李国林[1] 俞建东[1] 许晓婷[1] 吕丽虹[1] 闵军[1] 林浩铭[1] 万云乐[1] 

机构地区：[1]中山大学孙逸仙纪念医院肝胆胰外科,广州510120

出　　处：《中华实验外科杂志》2010年第11期1671-1673,共3页Chinese Journal of Experimental Surgery

基　　金：国家自然科学基金资助项目（30671987）

摘　　要：目的 利用羟基荧光素二醋酸盐琥珀酰亚胺脂（CFSE）标记大鼠肝脏NK细胞进行体内示踪,观察其在受鼠体内的存活.方法 经SD大鼠门静脉转输1×106个/ml CFSE体外标记新鲜分离的SD供鼠肝脏NK细胞,于输人后1、3、7、15 d分别切取肝脏和脾脏行冰冻切片免疫荧光观察,并收集门静脉血监测淋巴细胞和中性粒细胞的比例变化.结果 CFSE对大鼠肝脏NK细胞的标记率为98.63% 转输NK细胞后第1天受鼠肝脏内CFSE荧光最强,7 d时荧光变微弱,15 d时荧光基本消失 脾脏在第1、3天时可见较弱荧光,7 d时荧光消失.转输NK细胞后相同时间点肝脏内CFSE荧光均明显强于脾脏.转输肝脏NK细胞的受鼠门静脉血中性粒细胞比例第1天时最高,随时间延长逐渐下降,第15天时基本回复正常水平,淋巴细胞比例仅一过性降低.结论 转输的肝脏NK细胞在受体内的数量随时间延长而减少,在受体内的存活时间约为15 d.Objective To investigate the survival of donor hepatic NK cells labeled with carboxyfluorescein diacetate succiminidyl ester （CFSE） after transfusion into isogenic rats via portal vein （PV）.Methods The fresh isolated hepatic NK cells from SD rats were identified by anti-rat NKR-P1 antibody staining. 1 × 106/mL NK cells labeled by CFSE in vitro were transfused into recipient SD rats via portal vein. Recipient liver and spleen tissues were harvested for detection of the fluorescence of NK cells labeled with CFSE on the day 1 to 15 after transfusion. Portal vein blood was collected for observation of the ratio of lymphocytes and neutrophilic granulocytes dynamically. Results The CFSE labeling rate of hepatic NK cells in vitro was 98.63%. The fluorescence of CFSE labeled cells in liver was the strongest on the day 1 after transfusion, while that was weak on the day 7. However, it could hardly been detected on the day 15.The fluorescence in spleen was weak on the day 1 and 3, but it could never been detected on the day 7.The ratio of lymphocytes in the portal vein blood of recipients was dropped after liver NK cells were transfused on the day 1. But the ratio in neutrophilic granulocyte field reached its peak on the day 1, dropped as time went on and returned to the normal level one the day 15. Conclusion Donor hepatic NK cells transfused via PV are decreased as time goes on in the recipients. The survival time of transfused cells in the recipient is about 15 days. The results may give us an insight into new strategy for cancer immunotherapy and transplant immunology.

关 键 词：肝脏NK细胞 示踪 免疫荧光 

分 类 号：R730.51[医药卫生—肿瘤]