脾源性酪氨酸激酶基因启动子甲基化与髓母细胞瘤细胞侵袭转移的关系

doi:10.13459/j.cnki.cjap.2016.02.010

中国应用生理学杂志 ›› 2016, Vol. 32 ›› Issue (2): 132-136.doi: 10.13459/j.cnki.cjap.2016.02.010

脾源性酪氨酸激酶基因启动子甲基化与髓母细胞瘤细胞侵袭转移的关系

吉海龙¹, 史鹏飞¹, 周洁², 毛天明³, 罗永康³, 陈茜⁴, 周开宇¹

1. 温州医科大学附属第一临床学院, 浙江温州 325000;
2. 浙江省台州学院医学院, 台州 318000;
3. 浙江省台州市立医院神经外科, 台州 318000;
4. 浙江省台州市立医院病理科, 台州 318000

收稿日期:2015-11-02 修回日期:2015-12-09 出版日期:2016-03-28 发布日期:2018-06-12
通讯作者: 周开宇,Tel:13957680507;E-mail:kerry2000year@163.com E-mail:kerry2000year@163.com
基金资助:
台州市科技局科技计划资助项目（14SF05）；浙江省中医药管理局科技计划资助项目（2013ZA133，2015ZB133）

The relationship between hypermethylation of Syk gene promoter and medulloblastoma cell invasion and metastasis

JI Hai-long¹, SHI Peng-fei¹, ZHOU Jie², MAO Tian-ming³, LUO Yong-kang³, CHEN Xi⁴, ZHOU Kai-yu¹

1. The First Affiliated Hospital of Wenzhou Medical University, Wenzhou 325000;
2. Taizhou University Medical School, Taizhou 318000;
3. Department of Neurosurgery, Zhejiang Taizhou Municipal Hospital, Taizhou 318000;
4. Department of Pathology, Zhejiang Taizhou Municipal Hospital, Taizhou 318000, China

Received:2015-11-02 Revised:2015-12-09 Online:2016-03-28 Published:2018-06-12
Supported by:
台州市科技局科技计划资助项目（14SF05）；浙江省中医药管理局科技计划资助项目（2013ZA133，2015ZB133）

摘要/Abstract

摘要： 目的：探讨甲基化转移酶抑制剂5-氮杂-2-脱氧胞苷（5-aza-CdR）抑制脾源性酪氨酸激酶（Syk）基因启动子的甲基化后对髓母细胞瘤Daoy细胞侵袭转移能力的影响。方法：用甲基化转移酶抑制剂5-aza-CdR处理体外培养的髓母细胞瘤Daoy细胞，通过甲基化特异性PCR（MSP）、Real time-PCR、Western blot及Transwell实验方法分别检测不同浓度5-aza-CdR处理后髓母细胞瘤Daoy细胞中脾源性酪氨酸激酶（Syk）基因启动子区甲基化、mRNA表达、蛋白表达及细胞穿膜数的变化。结果：髓母细胞瘤Daoy细胞中Syk基因启动子存在过甲基化，与对照组比较，经不同浓度5-aza-CdR处理后，其Syk基因启动子区甲基化受到不同程度抑制，Syk mRNA的表达量最高上调（3.40±0.24）倍（P<0.01）；Syk蛋白的表达量最高上调（3.23±0.19）倍（P<0.01）；细胞侵袭及转移能力降低（P<0.05），差异有统计学意义。结论：髓母细胞瘤Daoy细胞中Syk基因启动子甲基化导致其表达下调，可能是髓母细胞瘤发生转移的机制之一；而甲基化转移酶抑制剂5-aza-CdR可抑制其启动子区的甲基化，使Syk的表达水平上调，抑制肿瘤细胞侵袭及转移能力。

关键词: 髓母细胞瘤, 脾源性酪氨酸激酶(Syk)基因, 甲基化, 5-氮杂-2-脱氧胞苷, 侵袭转移

Abstract: Objective: To investigate the effect of demethylation of Syk gene promoter by the methylation transferase inhibitor 5-aza-CdR on the invasion and metastasis of medulloblastoma cell line Daoy. Methods: Medulloblastoma cell line Daoy was treated with 5-aza-CdR in vitro. Methylation-specific PCR, real time-PCR and Western blot were used to detect Syk gene promoter methylation status, Syk mRNA and protein expression respectively. Transwell was employed to study the invasion and metastasis of medulloblastoma cell line Daoyby counting the cells that had invaded through Matrigel and migrated to the undersurface of the membrane before and after treatment of 5-aza-CdR. Results: In comparison to control group, Syk gene promoter of 5-aza-CdR-treated groups was demethylated and expression of Syk mRNA and protein was significantly up-regulated by 3.40±0.24 folds (P<0.01) and 3.23±0.19 folds (P<0.01) respectively. The invasiveness and metastasis of medulloblastoma cell line Daoy was decreased(P<0.05). Conclusion: Hypermethylation of Syk gene promoter is responsible for the down-regulation of Syk gene expression in medulloblastoma cell line Daoy, which may be one of the mechanisms that enhanced cell invasion and metastasis. While 5-aza-CdR can reverse the hypermethylation of Syk gene promoter and restore Syk gene expression and thus suppresses invasiveness and metastasis of tumor cells.

Key words: medulloblastoma, Syk gene, methylation, 5-aza-CdR, invasion and metastasis

吉海龙, 史鹏飞, 周洁, 毛天明, 罗永康, 陈茜, 周开宇. 脾源性酪氨酸激酶基因启动子甲基化与髓母细胞瘤细胞侵袭转移的关系[J]. 中国应用生理学杂志, 2016, 32(2): 132-136.

JI Hai-long, SHI Peng-fei, ZHOU Jie, MAO Tian-ming, LUO Yong-kang, CHEN Xi, ZHOU Kai-yu. The relationship between hypermethylation of Syk gene promoter and medulloblastoma cell invasion and metastasis[J]. CJAP, 2016, 32(2): 132-136.

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脾源性酪氨酸激酶基因启动子甲基化与髓母细胞瘤细胞侵袭转移的关系

The relationship between hypermethylation of Syk gene promoter and medulloblastoma cell invasion and metastasis

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