【摘要】 目的:观察三维受控组装系统下,胚胎肝细胞在三维立体结构的体外生长状态,探讨胚胎肝细胞在肝组织工程中应用的可行性。方法:用清华大学机械工程系研制的"三维受控组装系统",将第15 d小鼠胚胎肝细胞作为肝组织工程的种子细胞,与以明胶为主的复合材料混合,构建成复杂三维立体结构,观察其体外生长发育状态。对体外培养1周及4周的三维类肝组织标本进行苏木精-伊红(HE)染色,免疫组织化学方法检测甲胎蛋白(AFP)及白蛋白(ALB)的表达,并对体外培养4周的三维类肝组织用PAS显色法检测肝糖原表达。结果:HE染色结果显示体外培养的胚胎肝细胞在三维支架材料中,可形成含有类血管和肝组织样结构;体外培养1周的类肝组织AFP表达呈阳性,体外培养4周的三维类肝组织ALB表达呈阳性,PAS显色亦呈阳性。结论:在三维受控组装系统的构建下,呈立体状生长的胚胎肝细胞,可逐渐形成肝组织样结构,并显示一定的肝脏功能。更多还原

【Abstract】 Objective: To observe using embryonic hepatocytes as the seed cells, the growth state of the 3D structure in vitro which structured by "three dimension(3D) controlled assembly system ", and to investigate the feasibility of embryonic hepatocytes for hepatic tissue engineering. Methods: We used "3D controlled assembly system" which was developed by Department of Mechanical Engineering Tsinghua University. We mixed embryonic 15 day mice embryonic hepatocytes with gelatin-based composite materials to assemble into complex 3D structures. The 3D liver-like tissues were cultured in vitro, and their growing situation were observed by Hematoxylin and Eosin(HE) staining after cultured for 1 week and 4 weeks. The expression of alpha fetoprotein(AFP) and albumin(ALB)were evaluated by immunohistochemistry in 3D liver-like tissue section specimens after cultured for 1 week and 4 weeks, and periodic acid schiff’s(PAS) reaction was used for detecting the expression of the liver glycogen after cultured in vitro for 4 weeks. Results: The results showed that in 3D scaffold material, embryonic hepatocytes cultured in vitro could form some structures containing vascular-like and liver-like tissues. AFP was highly expressed in liver-like tissues cultured for 1 weeks. ALB and liver glycogen were expressed in 3D liver-like tissues cultured in vitro for 4 weeks. Conclusions: In 3D controlled assembly system, the embryonic hepatocytes which grow three-dimensionally are able to form liver-like structures and show some liver functions.更多还原