中国组织工程研究 ›› 2012, Vol. 16 ›› Issue (11): 1963-1968.doi: 10.3969/j.issn.1673-8225.2012.11.015

• 组织构建实验造模 experimental modeling in tissue construction • 上一篇    下一篇

一种适合肝细胞生长的无血清培养基*★

赵逸超1,汪  艳1,卢  海1,简国登2,潘明新1,高  毅1   

  1. 1南方医科大学珠江医院肝胆二科,广东省广州市   510282;2佛山市禅城区中心医院肝胆外科,广东省佛山市   528031
  • 收稿日期:2011-12-20 修回日期:2012-01-16 出版日期:2012-03-11
  • 通讯作者: 高毅,主任医师,教授,南方医科大学珠江医院肝胆二科,广东省广州市 510282 gaoyi6146@163.com
  • 作者简介:赵逸超★,男,1987年生,河北省唐山市人,汉族,南方医科大学在读硕士,主要从事生物人工肝方面的研究。 smuzyc@126.com
  • 基金资助:

    国家高技术研究发展计划(863 计划):No.2006AA02A141,人源细胞混合型生物人工肝的研制与开发。

A new serum-free medium for hepatocytes growth

Zhao Yi-chao1, Wang Yan1, Lu Hai1, Jian Guo-deng2, Pan Ming-xin1, Gao Yi1   

  1. Second Department of Hepatobiliary, Zhujiang Hospital of Southern Medical University, Guangzhou  510282, Guangdong Province, China
  • Received:2011-12-20 Revised:2012-01-16 Online:2012-03-11
  • Contact: author: Gao Yi, Chief physician, Professor, Second Department of Hepatobiliary, Zhujiang Hospital of Southern Medical University, Guangzhou 510282, Guangdong Province, China gaoyi6146@163.com
  • About author:Zhao Yi-chao★, Studying for master’s degree, Second Department of Hepatobiliary, Zhujiang Hospital of Southern Medical University, Guangzhou 510282, Guangdong Province, China smuzyc@126.com
  • Supported by:

    National High Technology Research and Development Program of China (863 Program), No. 2006AA02A141*

摘要:

背景:寻找一种既能适合细胞生长又避免添加血清的培养基迫在眉睫,无血清培养应运而生。
目的:探索一种适用于C3A细胞生长的无血清培养基,为肝细胞用于临床治疗以及生物人工肝走入临床奠定基础。
方法:分别用无血清培养基、完全培养基和基础培养基培养C3A细胞,观察细胞的生长状态,绘制细胞生长曲线和活力曲线,全自动生化分析仪测定上清中谷丙转氨酶漏出量、尿素含量,放射免疫分析法检测白蛋白分泌量,实时荧光定量PCR检测基因水平的变化情况。
结果与结论:3种培养基培养的细胞生长良好,无血清培养基组与完全培养基组细胞密度在培养周期内,除第7天以外差异无显著性意义(P > 0.05)。培养第4~7天,其他两组明显高于基础培养基组(P < 0.05);无血清培养基尿素合成水平>完全培养基>基础培养基(P < 0.05)。Real-time qPCR显示,无血清培养基组相对于完全培养基组仅CK18、CK19和HNF4α基因表达量下降(P < 0.05)。说明实验研制了一种适合于肝细胞(C3A)生长的无血清培养基,与传统血清培养能达到同样的效果。
关键词:培养基;无血清培养;C3A细胞;肝细胞;生物人工肝;PCR检测
doi:10.3969/j.issn.1673-8225.2012.11.015
 

关键词: 培养基, 无血清培养, C3A细胞, 肝细胞, 生物人工肝, PCR检测

Abstract:

BACKGROUND: A medium both for cell growth and avoiding adding serum medium is imminent, so serum-free medium emerge as the times require.
OBJECTIVE: To develop a new serum-free medium for C3A cells to lay the foundation for the clinical application of hepatocytes and bioartificial liver.
METHODS: C3A cells were cultivated with serum-free medium, complete medium and basal medium, respectively. The growth and morphological characteristics of cells were observed. Curves of cell growth and the energy were drawn. The leakage volume of alaninetransaminase, urea level and the level of albumin were determined by automatic biochemistry analyzer and radioimmunoassay. The changes of gene expression were determined by real-time fluorescence quantitative PCR.
RESULTS AND CONCLUSION: The cells in the three kinds of mediums grew well. Apart from the 7th day, the cell density had no significant difference between the serum-free medium and complete medium groups in the culture period (P > 0.05), whereas it was obviously higher in these two groups than in the basal medium group at 4-7 days (P > 0.05). The level of urea in the serum-free medium group was significantly higher than that in the other two groups, and the level of urea in the complete group was higher than that in the basal medium group (P < 0.05). Real-time fluorescence quantitative RCR revealed that gene expressions of CK18, CK19 and hepatocyte nuclear factor 4α were lower in the serum-free group as compared with the complete medium group (P < 0.05). The results showed that the new serum-free medium for hepatocytes is effective just like the conventional complete medium.
 

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