组织工程与重建外科杂志

组织工程与重建外科杂志 ›› 2018, Vol. 14 ›› Issue (5): 241-244.doi: 10.3969/j.issn.1673-0364.2018.05.001

• 论著 •    下一篇

3D生物打印构建复层尿道组织的初步研究

皮庆猛，王晓磊，张志亮，周碧玉，任晓芸，陆毅，黄一雄，傅敏刚，范志宏   

  1. 上海交通大学医学院附属仁济医院整形外科；峄城中医院内科
  • 收稿日期:2018-07-19 发布日期:2020-07-23
  • 作者简介:范志宏（E-mail：fanzhihong@163.com）。

Preliminary Application of Urethral Tissue Construction with Multiple Layers Using 3D Bio -printing

PI Qingmeng,WANG Xiaolei,ZHANG Zhiliang,ZHOU Biyu,REN Xiaoyun,LU Yi,HUANG Yixiong,FU Mingang,FAN Zhihong   

  1. Department of Plastic Surgery, Renji Hospital, Shanghai Jiaotong University School of Medicine; Department of Medicine, Yicheng Medical Hospital
  • Received:2018-07-19 Published:2020-07-23
  • Contact: 仁济医院培养基金(PYIII-17-032)

摘要： 目的探讨利用生物打印体外构建具有复层结构的尿道空腔组织的可行性。方法将7%GelMA和2%Alginate制备复合水凝胶,分别同绿色和红色的microbeads或2×106 cells/mL尿道上皮细胞和尿道平滑肌细胞混匀,通过本实验室改进的同轴多通道打印系统,以10～20μL/min的速度推进内、外层,以80～200μL/min的速度推进CaCl2,边打印边交联;打印后不含细胞的空腔结构,置于荧光显微镜下观察;打印后含有细胞的组织,以尿道上皮细胞和尿道平滑肌细胞混合培养液于六孔板内体外培养,分别于第0～7天live/dead染色检测细胞活力,F-actin染色检测细胞铺展情况;将培养24 h的打印后组织,予以红色颜料灌注。结果复合水凝胶混合microbeads后,荧光显微镜下可以清楚看到内、外双层结构,复层管腔的内、外径分别为(663.66±51.74)μm和(976.84±63.43)μm,内外亚层壁厚分别为(62.06±10.45)μm和(93.78±10.25)μm。尿道细胞在第0天和第7天的细胞活力分别为84.52%和86.26%,免疫荧光染色第7天F-actin染色铺展均匀,提示细胞铺展生长。灌注过程中,打印后空腔组织内红色灌注液可以顺利通过管腔结构,颜料并无外溢,管壁无破损。结论利用复合水凝胶和同轴多通道生物打印系统可以初步构建尿道空腔结构,打印后尿道细胞可在空腔结构内长期存活。

关键词: 3D生物打印, 尿道空腔组织, 水凝胶, 海藻酸盐, 甲基丙烯酸酐化明胶

Abstract: Objective To investigate the feasibility of multiple-layers reconstruction of urethral tissue with 3D bioprinting technology. Methods Mixed blend of 7% GelMA and 2% Alginate with green and read microbeads respectively, or 2×106cells/mL urethral epidermal cells and smooth muscle cells, printed on the system of co-axial multiple channel bioprinting platform. The speed of inner and outer layer printing was 10-20 μL/min and CaCl2extruding was 80-200 μL/min. Then the printed structure was observed under microscopy, and the printed hollow tissues loaded urethral cells were tested with live/dead and F-actin staining from day 0 to day 7. The printed hollow structure was perfused with color medium after cultured 24 hours in vitro. Results With hydrogels mixed with microbeads, two distinct sublayers of printed hollow structure can be observed clearly under microscopy. The diameter of inner and outer layer was (663.66±51.74) μm and (976.84±63.43) μm respectively, and wall length of inner and outer layer was (62.06±10.45) μm and (93.78±10.25) μm respectively. The viability of printed urethral tissue was 84.52% and 86.26% at day 0 and day 7. F-actin expression were observed on day 7 after immunofluorescent staining and showed the spread cells. During the process of perfusion of hollow tissue, colored medium can circulate the channel smoothly without color exuding. Conclusion Urethral hollow structure can be printed with combined hydrogel by the method of co-axial multiple channel bioprinting system. And the urethral cells can survive for long term after printing.

Key words: 3D bioprinting, Urethral hollow tissue, Hydrogel, GelMA, Alginate

中图分类号: