静电纺丝聚乳酸聚乙醇酸／聚乙二醇纳米材料与小鼠胚胎肝细胞生物相容性的研究  

Biological compatibility of electrospun polylactic - co - glycolic acid - polyethylene glycol nanofibers with mouse embryonic hepatocyte in vitro

作　　者：胡昆鹏[1] 刘波[1] 姚志成[1] 罗慧[2] 熊志勇[3] 范伟明[3] 许瑞云[3] 邓美海[3] 

机构地区：[1]中山大学附属第三医院岭南医院普通外科,广州510530 [2]中山大学附属第三医院岭南医院手术室 [3]中山大学附属第三医院肝胆外科

出　　处：《中华实验外科杂志》2015年第12期3024-3027,共4页Chinese Journal of Experimental Surgery

摘　　要：目的探讨小鼠胚胎肝细胞与静电纺丝聚乳酸聚乙醇酸（PLGA）-聚乙二醇（PEG）共聚物纳米材料的体外生物相容性。方法采用胰酶分步消化法分离培养孕16d昆明小鼠胚胎肝细胞并进行免疫荧光鉴定；使用静电纺丝法制备PLGA和PLGA—PEG纳米材料；取第5代胚胎肝细胞分别接种于PLGA和PLGA—PEG纳米纤维材料上，进行体外培养，细胞计数试剂盒（CCK-8）法测定两种材料的细胞毒性及细胞在材料表面的黏附、增殖；扫描电镜（SEM）观察材料结构、细胞复合材料的形态。结果体外分离培养的小鼠胚胎肝细胞呈单层多边形样生长，免疫荧光检测胚胎肝细胞角蛋白18（CK18）和甲胎蛋白（AFP）表达阳性；CCK-8检测显示PLGA（0．255±0．062、0．259±0．039、0．239±0．029、0．257±0．026）和PLGA—PEG（0．293±0．033、0．283±0．200、0．249±0．021、0．244±0．025）两种纳米材料与空白对照组（0．258±0．017）比较均无明显的细胞毒性（P〉0．05）。小鼠胚胎肝细胞在材料表面生长良好，PLGA—PEG组（0．113-4-0．005、0．234±0．008、0．678±0．006、1．108±0．004、1．182±0．004、1．114±0．011）及PLGA组（0．108±0．005、0．206±0．010、0．502±0．009、0．817±0．005、0．807±0．007、0．819±0．005）吸光度（A）值均随时间延长而增大。两组A值在各时间点差异均有统计学意义（P〈0．05）。各时间点两组材料的细胞黏附率差异有统计学意义，PLGA．PEG组（21．250±2．165、51．250±7．806、75．000±3．750）明显高于PLGA组（15．000±3．750、38．750±7．806、65．000±4．330，P〈0．05）。SEM结果显示，与PLGA组比较，PLGA—PEG组中小鼠胚胎肝细胞更易于黏附生长。结论静电纺丝法制备的PLGA—PEG纳米材料具有良好的细胞生物相容性，安全无毒，适宜胚胎肝细胞生长。Objective To study the biological compatibility of electrospun polylactic - co - glycolic acid -polyethylene glycol （PLGA -PEG） nanofibers with mouse embryonic hepatocyte in vitro. Methods Mouse embryonic hepatocytes were isolated from embryos of E16 days KunMing mouse, cultured and identi- fied. Electrospinning was used to prepare PLGA and PLGA - PEG nanofibers. The 5th passage embryonic hepatocytes were seeded onto PLGA and PLGA- PEG scaffolds respectively. Cell counting kit- 8 （ CCK - 8 ） was used to detect the eytotoxicity, adhesion rate and proliferation of embryonic hepatocytes on the two nanofibers. The cell morphological changes on scaffolds were observed by electron microscope （SEM）. Results Mouse fetal hepatocytes were polygonal and the specific expression of cytokeratin 18 （CK18） and alpha - 1 - fetoprotein （AFP） was detected by immunofluoreseence. CCK - 8 assay showed thePLGA （0.255±0.062, 0.259 ±0.039, 0.239 ±0.029, 0.257 ± 0.026） and PLGA - PEG （0. 293 ±0. 033, 0. 283 ±0. 200, 0. 249 ±0. 021, 0. 244±0. 025） nanofibers showed no significant cyto- toxicity compared with blank control group （ 0. 258 ± 0. 017, P 〉 0. 05 ）. Mouse embryonic hepatocytes grew well on both two nanofibers, and absorbanee （A） value in PLGA - PEG group （0. 113 ± 0.005, 0.234s0.008, 0.678 ±0.006, 1.108 ±0.004, 1.182 ±0.004, 1.114 ±0.011） and PLGA group （0. 108 ± 0. 005, 0. 206 ±0. 010, 0. 502 ± 0. 009, 0. 817 ± 0. 005, 0. 807± 0. 007, 0. 819 ± 0. 005 ） in- creased with prolongation of incubation time, and there were statistically significant differences between two groups at each time point （P 〈 0. 05 ）. Besides, the cell adhesion rate was significantly higher in the PLGA-PEG group （21.250 ± 2. 165, 51.250 ± 7.806, 75~ ± 3.750） than in the PLGA group （ 15. 000 ± 3.75,0, 38. 750 ± 7. 806, 65. 000 ± 4. 330, P 〈 O. 05 ） at each time point （ P 〈 0. 05 ）. SEM showed as compa}ed with PLGA scaffold; PLGA - PEG scaffold was better for mouse emb

关 键 词：静电纺丝 聚乳酸聚乙醇酸 聚乙二醇 胚胎肝细胞 生物相容性 

分 类 号：R318[医药卫生—生物医学工程]