中国组织工程研究

中国组织工程研究 ›› 2010, Vol. 14 ›› Issue (23): 4290-4294.doi: 10.3969/j.issn.1673-8225.2010.23.025

• 干细胞基础实验 basic experiments of stem cells • 上一篇    下一篇

三种方法原代培养人牙周膜细胞

蒋俊强，王忠朝，黎春晖，蔡  炜   

  1. 泸州医学院附属口腔医院口腔内科，四川省泸州市  646000
  • 出版日期:2010-06-04 发布日期:2010-06-04
  • 通讯作者: 蔡 炜，讲师，泸州医学院附属口腔医院口腔内科，四川省泸州市 646000 caiwei311@tom.com
  • 作者简介:蒋俊强，男，1976年生，四川省邻水县人，汉族，2006年四川大学华西口腔医学院毕业，硕士，讲师，主要从事牙周病病因及其防治的研究。 jqjiang@126.com
  • 基金资助:

    四川省卫生厅科研基金资助项目(川卫办发[2007]431号)。

Primary culture of human periodontal ligament cells using three methods

Jiang Jun-qiang, Wang Zhong-chao, Li Chun-hui, Cai Wei   

  1. Department of Oral Medicine, Stomatological Hospital Affiliated to Luzhou Medical College, Luzhou  646000, Sichuan Province, China
  • Online:2010-06-04 Published:2010-06-04
  • Contact: Cai Wei, Lecturer, Department of Oral Medicine, Stomatological Hospital Affiliated to Luzhou Medical College, Luzhou 646000, Sichuan Province, China caiwei311@tom.com
  • About author:Jiang Jun-qiang, Master, Lecturer, Department of Oral Medicine, Stomatological Hospital Affiliated to Luzhou Medical College, Luzhou 646000, Sichuan Province, China jqjiang@126.com
  • Supported by:

    the Scientific Research Foundation of Health Department of Sichuan Province, No. [2007]431*

PDF

575

被引次数

74

摘要：

背景：牙周膜细胞培养是牙周细胞生物学研究的基础。牙周膜细胞原代培养因其成功率较低，一直是制约牙周细胞生物学研究的瓶颈之一。如何寻找一种简单有效的培养方法，提高原代培养成功率一直是近年来牙周细胞生物学的研究热点之一。

目的：比较组织块法、酶消化法、酶消化结合组织块法对人牙周膜细胞的培养效果，探索一种更有效的人牙周膜细胞的体外培养方法。

方法：取58例正常恒牙牙周膜组织，随机分为3组，分别采用组织块法、酶消化法、酶消化结合组织块法进行原代培养，再进行传代培养、倒置显微镜下观察细胞生长情况以及细胞形态；免疫组织化学鉴定波形蛋白与角蛋白；取第4代细胞，绘制生长曲线。

结果与结论：酶消化结合组织块法培养成功率(35%)明显高于其他两种方法(11%和21%)。3种方法获得的细胞均符合正常人牙周膜细胞形态学特征和生物学特征，生长良好。第4代细胞生长曲线均为近似的“S”形，有明显的滞留期、对数生长期和平台期。提示实验建立的酶消化结合组织块法较为简单并具有较高的成功率，是一种培养牙周膜细胞的可行办法。

关键词: 人牙周膜细胞, 细胞培养, 组织块法, 酶消化法, 酶消化结合组织块法

Abstract:

BACKGROUND: The culture of human periodontal ligament cell (hPDLC) is the basis of biological research of periodontal cells. The success rate of primary culture of periodontal membrane cells is low, which restricts the biological research of periodontal cells. How to seek a simple effective culture method or how to elevate success rate of primary culture is always one of hot focus studies of biology of periodontal cells.
OBJECTIVE: To explore an effective culture technique of hPDLC by comparing the effects of tissue explant, enzymatic digestion, the combination of tissue explant and enzymatic digestion.
METHODS: The tissues of periodontal ligament isolated from 58 normal human permanent teeth were collected and divided into three groups randomly, and then primarily cultured using tissue explant, enzymatic digestion, the combination of tissue explant and enzymatic digestion, followed by subculture. Cell growth and morphology were observed under an inverted microscope. Vimentin and keratin were identified by immunohistochemistry. The fourth passage of cells were collected to draw growth curve.
RESULTS AND CONCLUSION: The successful rate of the combination of tissue explant and enzymatic digestion (35%) was significantly higher than others (11% and 21%). The morphological and biological characteristics of the hPDLC in present study were similar to those typical hPDLC, which grew well. The growth curve of the fourth passage of cells was similar to “S” shape, with significant detention period, logarithmic phase and plateau phase. Results have suggested that the combination of tissue explant and enzymatic digestion is relatively simple and has a high successful rate, so it can be regarded as an effective method for hPDLC culture.

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