解剖学报 ›› 2015, Vol. ›› Issue (3): 348-353.doi: 10.16098/j.issn.0529-1356.2015.03.010

• 肿瘤生物学 • 上一篇    下一篇

神经生长因子在食管鳞癌细胞中的表达及分泌

李秋霞 裴学莲 刘洋 王玉婷 黄燕燕 周宗瑶 慕晓玲*   

  1. 石河子大学医学院 新疆地方与民族高发病教育部重点实验室,新疆 石河子 832000
  • 收稿日期:2014-10-29 修回日期:2014-12-24 出版日期:2015-06-06 发布日期:2015-06-06
  • 通讯作者: 慕晓玲 E-mail:xiaolingmur@163.com
  • 基金资助:

    新疆兵团科技支疆计划项目;国家自然科学基金项目

Expression and secretion of nerve growth factor in esophageal squamous cell cancer cell line Eca109

LI Qiu-xia PEI Xue-lian LIU Yang WANG Yu-ting HUANG Yan-yan ZHOU Zong-yao MU Xiao-ling*   

  1. Key Laboratory of Xinjiang Endemic and Ethnic Disease School of Medicine,Shihezi University,Xinjiang Shihezi 832000,China
  • Received:2014-10-29 Revised:2014-12-24 Online:2015-06-06 Published:2015-06-06
  • Contact: MU Xiao-ling E-mail:xiaolingmur@163.com
  • Supported by:

    ;National Natural Science Foundation of China

摘要:

目的 探讨神经生长因子(NGF)与食管鳞癌细胞分化的相关性。方法 采用有限稀释法分选出圆形和梭形单细胞克隆,采用无血清悬浮培养获得细胞球细胞, 贴壁的Eca109细胞分别在含血清和不含血清培养基中培养48h;分别采用反转录-聚合酶链式反应(RT-PCR)技术和免疫印迹法,检测NGF在食管鳞癌细胞中mRNA水平和蛋白水平的表达;免疫荧光技术检测NGF在食管鳞癌细胞中的表达定位;免疫组织化学法检测食管鳞癌组织中NGF的表达定位;用酶联免疫吸附法(ELISA)检测NGF在Eca109细胞培养基中的分泌情况。结果 在Eca109细胞中能检测到NGF的mRNA水平和蛋白水平的表达,其中NGF在细胞球细胞中的mRNA水平和蛋白水平的表达量最高。食管癌组织中检测到NGF表达于细胞质。Eca109细胞在无血清培养条件下能够分泌NGF,且明显高于含血清培养基中的含量。结论 食管癌细胞系Eca109表达并分泌NGF,并且食管癌组织中表达NGF,NGF可能在维持食管鳞状细胞癌的干细胞特性发挥了重要作用。

Abstract:

Objective To investigate the relationship between the nerve growth factor (NGF) and the cell differentiation of esophageal squamous cell cancer. Methods Round and spindle single-cell clones were sorted by limited dilution, and spheres were obtained by cultured in serum-free medium. Adherent Eca109 cells were cultured in serum-containing and serum-free culture medium 48hours, respectively.The expression of NGF mRNA and protein levels in esophageal squamous cell carcinoma (ESCC) were assessed by reverse transcriptase expression of the polymerase chain reaction (RT-PCR) technology and Western blotting analysis, respectively.Immunohistochemical staining determined the nerve growth factor expression in esophageal squamous cell carcinoma specimen. The secretion of NGF in Eca109 was determined by enzyme-linked immunosorbent assay (ELISA). Results The expression of NGF mRNA and protein levels was detected in ESCC,and the spheres cells expressed the highest amount of NGF in mRNA and protein expression level. NGF was expressed in the cytoplasm of ESCC cells in esophageal squamous cell carcinoma specimen. ESCC cells could secreted NGF in serum-free culture conditions, and the NGF secreted was significantly higher than that in serum-containing medium. Conclusion ESCC cells of Eca109 express and secrete NGF, and esophageal squamous cell carcinoma specimen could express nerve growth factor. NGF may play an important role in maintaining ESCC stem cell characteristics.