1, 1′-双十八烷基-3, 3, 3′, 3′-四甲基吲哚碳菁高氯酸盐标记的大鼠骨髓间充质干细胞与聚己内酯薄膜共培养制作细胞补片  

Experimental study on co-culture of 1,1′-doctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate labeled rat bone marrow mesenchymal stem cells and polycaprolactone film in vitro to make cell patch

作　　者：张子畅 周帆 郑建伟 穆军升[1] 伯平[1] 尤斌[1] Zhang Zichang;Zhou Fan;Zheng jianwei;Mu Junsheng;Bo Ping;You Bin(Department of Cardiac Surgery,Beijing Anzhen Hospital Affiliated to Capital Medical University-Beijing Institute of Cardiopulmonary and Vascular Diseases,Beijing 100029,China;Department of Ultrasound,the Third Medical Center,General Hospital of Chinese People’s Liberation Army of China,Beijing 100039,China;Heart Center of Sunshine Fusion Hospital,Weifang 261205,China)

机构地区：[1]首都医科大学附属北京安贞医院-北京市心肺血管疾病研究所心脏外科,100029 [2]解放军总医院第三医学中心超声科,北京100039 [3]阳光融和医院心脏中心,潍坊261205

出　　处：《中华实验外科杂志》2022年第2期295-298,共4页Chinese Journal of Experimental Surgery

基　　金：国家自然科学基金(81870181);中国科学技术协会2021年度公共服务能力提升项目(2021GGFW003)。

摘　　要：目的将聚己内酯(PCL)薄膜与1, 1′-双十八烷基-3, 3, 3′, 3′-四甲基吲哚碳菁高氯酸盐(DiI)标记的骨髓间充质干细胞(BMSCs)共培养, 制备细胞补片。方法取1只清洁级SD大鼠作为实验对象, 通过分离培养获得大鼠BMSCs, 并进行流式细胞仪鉴定表面标志物。BMSCs培养至3代后, 使用DiI染料对BMSCs标记, 将DiI标记的BMSCs与PCL制成的薄膜共培养, 24 h后在荧光显微镜下观察细胞生长情况, 并固定进行电镜扫描, 在培养的1、4、7、10 d分别用细胞计数试剂盒(CCK-8)检测细胞增殖。多组间比较采用单因素方差分析, 两两比较采用LSD分析。结果 BMSCs流式细胞术鉴定:CD90、CD44H强阳性, CD11b/c、CD45阴性。荧光显微镜下, 可见DiI标记的BMSCs发红光, 呈梭形或多边形。扫描电镜下, PCL薄膜与DiI标记的BMSCs共培养形成的细胞补片, 其表面细胞数量多, 细胞状态正常。CCK-8测定结果显示第1天吸光度(A)值为0.330±0.025;第4天A值为0.620±0.012, 第7天A值为1.033±0.144, 第10天A值为1.223±0.133。各时间点A值之间差异有统计学意义(F=66.441, P<0.05)。结论 PCL薄膜可以作为一种DiI标记示踪的支架材料, 用于干细胞治疗。Objective Polycaprolactone(PCL)film was co-cultured with 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate(DiI)labeled bone mesenchymal stem cells(BMSCs)to prepare cell patch.Methods Clean grade SD rats were selected as experimental objects,BMSCs were isolated and cultured,and the surface markers were identified by flow cytometry.After the BMSCs were cultured for 3 passages,the BMSCs were stained with DiI dye,and the BMSCs DiI and PCL biomaterial film were co-cultured.After 24 h,the cell growth was observed under fluorescence microscope,and cells were fixed for scanning under electron microscope.The cell proliferation was detected by CCK-8 at 1,4,7,10 days of culture.The measurement data conforming to normal distribution were expressed as mean±standard deviation(±s).One way ANOVA was used for comparison among groups,and LSD analysis was used for pairwise comparison.Results BMSCs were strongly positive for CD90,CD44H,but negative for CD11b/c,CD45.Under fluorescence microscope,BMSCs DiI showed red light,fusiform or polygonal.Under the scanning electron microscope,the cell patch formed by co-culture of PCL film and DiI-BMSCs had a large number of cells on the surface and normal cell state.CCK-8 assay showed that the absorbance(A)value on the first day,4th day,7th day and 10th day was 0.330±0.025,0.620±0.012,1.033±0.144 and 1.223±0.133,respectively.There was significant difference in A value among the time points(F=66.441,P<0.05).Conclusion PCL film can be used as an in vivo tracer scaffold material for stem cell therapy.

关 键 词：干细胞治疗 聚己内酯 骨髓间充质干细胞 支架 

分 类 号：R457.7[医药卫生—治疗学]