中国组织工程研究

中国组织工程研究 ›› 2011, Vol. 15 ›› Issue (6): 990-994.doi: 10.3969/j.issn.1673-8225.2011.06.010

• 干细胞培养与分化 stem cell culture and differentiation • 上一篇    下一篇

大鼠成骨细胞的原代培养和鉴定

李晓峰，赵劲民，苏  伟，范  锲，罗世兴，马爱国   

  1. 广西医科大学第一附属医院创伤骨科手外科，广西壮族自治区南宁市  530021
  • 收稿日期:2010-07-23 修回日期:2010-11-20 出版日期:2011-02-05 发布日期:2011-02-05
  • 通讯作者: 赵劲民，博士，教授，广西医科大学第一附属医院创伤骨科手外科，广西壮族自治区南宁市 530021 zhaojinmin@126.com
  • 作者简介:李晓峰★，男，1974年生，吉林省德惠县人，汉族，2005年福建医科大学毕业，硕士，主治医师，主要从事创伤外科手外科、组织工程方面的研究。 xiaofengli74@163.com
  • 基金资助:

    广西医疗卫生重点科研项目(桂卫重200636)：组织工程种子细胞的实验研究。

Primary culture and identification of rat osteoblasts

Li Xiao-feng, Zhao Jin-min, Su Wei, Fan Qie, Luo Shi-xing, Ma Ai-guo   

  1. Department of Traumatic Orthopaedics and Hand Surgery, First Affiliated Hospital, Guangxi Medical University, Nanning  530021, Guangxi Zhuang Autonomous Region, China
  • Received:2010-07-23 Revised:2010-11-20 Online:2011-02-05 Published:2011-02-05
  • Contact: Zhao Jin-min, Doctor, Professor, Department of Traumatic Orthopaedics and Hand Surgery, First Affiliated Hospital, Guangxi Medical University, Nanning 530021, Guangxi Zhuang Autonomous Region, China zhaojinmin@126.com
  • About author:Li Xiao-feng★, Master, Attending physician, Department of Traumatic Orthopaedics and Hand Surgery, First Affiliated Hospital, Guangxi Medical University, Nanning 530021, Guangxi Zhuang Autonomous Region, China xiaofengli74@163.com
  • Supported by:

    the Key Scientific Research Program of Medical Health of Guangxi Zhuang Autonomous Region, No. 200636*

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1142

被引次数

50

摘要：

背景：组织工程需要大量的种子细胞，成骨细胞已成为骨组织工程构建的重要种子细胞之一。但成骨细胞取材困难，获得的成骨细胞的纯度不一。
目的：建立大鼠新生乳鼠颅骨来源成骨细胞的分离培养纯化方法，观察颅骨来源成骨细胞生物学特点。
方法：采用二次酶消化法对SD大鼠乳鼠成骨细胞进行原代培养，扩增。通过差速贴壁法进行成骨细胞纯化。通过形态学、细胞碱性磷酸酶检测、茜素红染色、钙结节Von kossa法染色、超微结构以及细胞增殖曲线，确定其增殖与成骨活性。
结果与结论：二次酶消化法培养颅骨来源成骨细胞可获得原代细胞增殖，传代扩增细胞具有典型成骨细胞形态学和生物学活性。碱性磷酸酶、茜素红染色、钙结节Von kossa法染色均呈阳性结果。超微结构显示为高分化功能活跃成骨细胞，细胞增殖曲线显示细胞生长活跃。提示，新生SD大鼠颅骨来源成骨细胞具有良好的增殖与成骨活性，能连续传代增殖，纯度高，细胞生物学特征稳定，适用于做体外实验研究。

关键词: 成骨细胞, 组织工程, 原代培养, 鉴定, 形态学

Abstract:

BACKGROUND: Tissue engineering requires a lot of seed cells. Osteoblasts have become an important seed cells in bone tissue engineering. However, there are the difficulties to derive osteoblasts and the different purity of osteoblasts was obtained.
OBJECTIVE: To establish the methods of isolated culture and purification of neonatal rat calvarial osteoblasts, and to observe the biological characteristics of the osteoblasts from the skull.
METHODS: Osteoblasts of Sprague-Dawley neonatal rats were primarily cultured and proliferated by the second enzyme digestion. Osteoblasts were purified by differential attachment method. Osteoblast proliferation and osteogenic activity were identified by morphology, alkaline phosphatase detection, Alizarin red staining and calcium nodules Von kossa staining, ultrastructure and cell proliferation curve.
RESULTS AND CONCLUSION: Primarily cultured calvarial osteoblasts could proliferate by the secondary source of enzyme digestion. The amplification cells showed representative morphological and biological characteristics of osteoblasts. Alkaline phosphatase, Alizarin red staining and calcium nodules Von kossa staining presented positive results. Ultrastructural features appeared as high-differentiated active osteoblasts. Cell proliferation curves showed active cells. Results indicated that calvarial osteoblast of Sprague-Dawley rat has good proliferation and osteogenic activity and can be continuously passaged, with high purity, cell biological characteristics of stability. Calvarial osteoblast is suitable for experiment in vitro.

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