【摘要】 目的通过三维(3D)生物打印兔原代肝细胞-海藻酸钠-明胶共混物,探讨3D生物打印技术构建类肝水凝胶结构体片段的可行性。方法采用Ⅳ型胶原酶消化法提取分离新生兔原代肝细胞;制备海藻酸钠-明胶水溶胶,与兔原代肝细胞共混;采用圆柱状微丝逐层交错堆积技术,进行兔肝细胞-海藻酸钠-明胶共混物的3D生物打印,获得三维结构体。用钙黄绿素-AM和碘化丙啶溶液对该结构体进行活/死细胞双荧光染色,评价打印后及培养后兔肝细胞的活性,计算细胞存活率。结果通过按照设计的参数,打印出含有兔原代肝细胞的网格状水凝胶结构体。该结构体的规格为10 mm×10 mm×1.2 mm,微丝间的孔隙相互通连,孔隙大小约450μm×450μm。打印后的兔肝细胞在三维结构体中仍可增殖,活/死细胞荧光染色结果表明结构体中兔肝细胞的存活率为82%±3%,兔原代肝细胞在结构体中分布均匀。结论本研究实现了兔肝细胞共混物结构体的3D生物打印,为构建类肝组织工程结构体片段提供新思路。更多还原

【Abstract】 Objective To construct three dimensional( 3D) structure lamella containing rabbit primary hepatocytes/alginate/gelatin blend using 3D bioprinting technique,so as to investigate the feasibility of 3D biological printing technique for the construction of structural fragments of liver like hydrogels. Methods Rabbit primary hepatocytes were extracted from liver tissue pieces of newborn rabbit by collagenase type Ⅳ digestion. Alginate and gelatin were mixed to form hydrosol,and rabbit primary hepatocytes were added into the hydrosol. 3D bioprinting system was used to construct 3D printed hepatic structures using rabbit primary hepatocytes/alginate/gelatin blend. The hepatic hydrogel structures were cultured,and the cells’ survival rate was detected by live-dead cell double fluorescence staining. Results The 3D structures with the grid hydrogel composed of multilayer staggered cylindrical microfilament were obtained. The size of the structure was about 10 mm × 10 mm × 1. 2 mm,and pore size was about 450 μm × 450 μm. The printed rabbit liver cells can still proliferate in the three-dimensional structure,the live/dead cell fluorescence staining results showed that the hepatocytes’ survival rate was about 82% ± 3% after printing,and the primary hepatocytes of rabbit were evenly distributed in the structure. Conclusion 3D bioprinting of hepatocytes is feasible,which can provide a new way to construct liver-like tissue.更多还原