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目的 研究乙型肝炎病毒ｘ（ Ｈ Ｂｘ） 蛋白对细胞周期影响，探讨 Ｈ Ｂｘ 蛋白致细胞周期紊乱的分子机理。方法 构建 Ｈ Ｂｘ 基因真核表达载体ｐ Ｃ Ｅ Ｐ４ Ｘ，电转染入 Ｈｅｐ Ｇ２ 细胞（ Ｘ 细胞） ，以ｐ Ｃ Ｅ Ｐ４空载体为对照（ Ｘ０ 细胞） ，潮霉素选择培养，克隆扩增转染细胞，经无血清培养同步化后，流式细胞仪检测细胞周期变化；细胞组化和 Ｗｅｓｔｅｒｎ ｂｌｏｔ 检测增殖细胞核抗原（ Ｐ Ｃ Ｎ Ａ） 和ｐ２１ Ｃ Ｉ Ｐ１／ Ｗ Ａ Ｆ１ 表达变化。结果 流式细胞仪检测示对照 Ｘ０ 细胞７０ ％ 进入 Ｇ０ Ｇ１ 期，而转 Ｘ 细胞仅５６ ％ 进入 Ｇ０ Ｇ１ 期，与含血清培养的亲本细胞 Ｈｅｐ Ｇ２ 几乎相同。ｐ２１ Ｃ Ｉ Ｐ１／ Ｗ Ａ Ｆ１ Ｄ 表达在 Ｘ０ 细胞阳性率为（８６ ±８） ％ ， Ｘ 细胞为（１６ ±６） ％ 。 Ｐ Ｃ Ｎ Ａ 表达在 Ｘ０ 细胞阳性率为（９ ±５） ％ ， Ｘ 细胞为（８６ ±３） ％ 。结论 Ｈ Ｂｘ 有推进细胞周期作用，并使细胞生长对血清依赖减少。 Ｈ Ｂｘ 可抑制细胞周期抑制蛋白ｐ２１ Ｃ Ｉ Ｐ１／ Ｗ Ａ Ｆ１ 表达，同时增强细胞 Ｄ Ｎ Ａ 合成。
Objective To study the effect of hepatitis B virus X (H Bx) on the cell cycle and to explore the molecular mechanism of Hbx induced cell cycle disorder. Methods The eukaryotic expression vector pC E P4 X of H Bx gene was constructed and transfected into Hep G2 cells (X cells). The pC E P4 empty vector was used as control (X0 cells), hygromycin selection culture, clone expansion The transfected cells were transfected with the recombinant plasmids. After synchronized by serum-free culture, the cell cycle was detected by flow cytometry. The expressions of P C N A and p21 C I P1 / W A F1 were detected by Western blot. Results Flow cytometry showed that 70% of the control X0 cells entered the G0G1 stage, while only 56% of the Xtransfering cells entered the G0G1 stage, almost identical to the Hep G2 cells in the serum-containing parental cells. The positive rate of p21 C I P1 / W A F1 D expression was (86 ± 8)% in X0 cells and (16 ± 6)% in X cells. The positive expression rate of P C N A was (9 ± 5)% in X0 cells and (86 ± 3)% in X cells. Conclusion H Bx can promote the cell cycle and decrease the dependence of serum on the cell growth. H Bx inhibits the expression of the cell cycle inhibitory protein p21 C I P1 / W A F1 while enhancing cellular D N A synthesis.