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[1]张琪,唐慧娴,韩一芳,等.HBV X基因突变促进肝癌发生的生物学机制[J].医学研究与战创伤救治(原医学研究生学报),2019,21(4):337-342.[doi:10.3969/j.issn.1672-271X.2019.04.001]
　ZHANG Qi,TANG Hui?xian,HAN Yi?fang,et al.Biological mechanisms of HBV X gene mutations in promoting hepatocellular carcinoma[J].JOURNAL OF MEDICALRESEARCH —COMBAT TRAUMA CARE,2019,21(4):337-342.[doi:10.3969/j.issn.1672-271X.2019.04.001]
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HBV X基因突变促进肝癌发生的生物学机制()
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《医学研究与战创伤救治》（原医学研究生学报）[ISSN:1672-271X/CN:32-1713/R]

卷:
第21卷
期数:
2019年4期
页码:
337-342
栏目:
基础研究
出版日期:
2019-07-20

文章信息/Info

Title:
Biological mechanisms of HBV X gene mutations in promoting hepatocellular carcinoma
文章编号:
1672-271X（2019）04-0337-06
作者:
张琪;  唐慧娴;  韩一芳;  叶福强;  王太武;  吕恒;  张锦海
作者单位：210002 南京，东部战区疾病预防控制中心（张 琪、韩一芳、叶福强、王太武、吕 恒、张锦海）; 211198 南京，中国药科大学药学院（唐慧娴）
Author(s):
ZHANG Qi1; 2;  TANG Hui?xian1; 2;  HAN Yi?fang1; 2;  YE Fu?qiang1; 2;  WANG Tai?wu1; 2;  L? Heng1; 2;  ZHANG Jin?hai1; 2
(1.Center for Disease Control and Prevention of Eastern Theater Command，Nanjing 210002，Jiangsu，China; 2.School of Pharmacy，China Pharmaceutical University，Nanjing 211198，Jiangsu，China)
关键词:
HBV X基因;  病毒突变;  细胞增殖;  细胞迁移;  细胞侵袭
Keywords:
hepatitis B virus X gene;  viral mutation;  cell proliferation;  cell migration;  cell invasion
分类号:
R373.2
DOI:
10.3969/j.issn.1672-271X.2019.04.001
文献标志码:
A
摘要:
目的 探讨乙型肝炎病毒(HBV) X基因(HBx)羧基末端四个突变位点C1653T、T1753C及A1762T/G1764A致癌的生物学机制。 方法 利用体外基因合成和定点突变构建野生型和复合突变型HBx重组质粒，转染至人肝癌细胞HepG2筛选稳定表达细胞株。通过绘制细胞生长曲线（CCK8法）、平板克隆形成实验、划痕实验、Transwell侵袭实验比较转染空载体(Vector)、野生型HBx(WT)和突变型HBx(Combo)细胞的增殖、迁移和侵袭能力。 结果 细胞生长曲线结果显示WT和Combo细胞的生长速率明显快于Vector（P<0.01），但WT和Combo之间差异无统计学意义（P>0.05）。平板克隆形成实验结果提示Combo细胞的克隆形成率显著高于WT（31.90% vs 16.00%，P<0.01），而WT略高于Vector(12.46%，P<0.05）。划痕实验结果显示Combo细胞48 h迁移距离显著高于WT细胞（P<0.01），而WT细胞显著高于Vector细胞（P<0.01）。Transwell侵袭实验结果显示Combo细胞穿过小室的细胞数最多[（227.80±17.85）个]，其次为WT细胞[（181.75±10.06）个]，Vector细胞最少[（85.72±3.19）个]，三者两两比较差异均有统计学意义（P<0.05）。 结论 野生型和突变型HBx均可显著增强HepG2细胞的增殖、迁移和侵袭能力，且突变型HBx促进细胞恶性生物学行为的能力显著强于野生型HBx，这为明确HBV致癌机制提供了一定的依据。
Abstract:
Objective To explore the biological functions of four mutations (C1653T，T1753C，and A1762T/G1764A) locating in the C?terminal region of hepatitis B virus (HBV) X gene (HBx) in hepatocarcinogenesis. Methods Recombinant constructs containing wild?type and compound mutations of HBx were constructed by gene synthesis and site?directed mutagenesis. Stable transfection was carried out in human hepatoma cell line HepG2. CCK?8 assay was used to draw the growth curves of the transfected cells. The proliferation，migration，and invasion abilities of cells transfected with empty vector (Vector)，wild?type HBx (WT)，and compound mutations of HBx (Combo) were evaluated by colony formation assay，wound healing assay，and transwell method. Results Cell growth curves revealed that the growth rates of WT and Combo cells were significantly higher than that of Vector cells (P<0.01)，but there was no difference between WT and Combo cells（P>0.05）. The colony formation rate of Combo cells (31.90%) was significantly higher than that of WT cells (16.00%) (P<0.01)，while it was slightly higher for WT cells when compared to Vector cells (12.46%) (P<0.05). Wound healing assay showed that the migration distance of Combo cells in 48h was significantly higher than WT cells (P<0.01)，while it was significantly higher in WT cells than that in Vector cells (P<0.01). Similarly，transwell assay turned out that the most number of invasive cells was observed in Combo cells (227.80±17.85)，followed by WT cells (181.75±10.06)，and then Vector cells (85.72±3.19)，and the differences between the three groups were all significant (P<0.05). Conclusion Wild?type and mutated HBx can both significantly enhance the proliferation，migration，and invasion ability of HepG2 cells，and the mutational HBx can further strengthen malignant biological behavior than wile?type HBx，which provided evidence for the mechanism of HBV?induced hepatocarcinogenesis.

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备注/Memo

备注/Memo:
收稿日期：2019-01-21
更新日期/Last Update: 2019-07-11