@@ 1 病例资料rn患者,女,47岁,因"双下肢疼痛、冰冷1月余,疼痛加重1 d"入院.既往史:慢性食管炎、慢性浅表性胃炎并糜烂性出血病史5年,贫血病史5年,卵巢囊肿病史2年.查体发现左足部苍白冰凉,足背动脉搏动较弱,右下肢皮色、皮温尚可,足背动脉搏动较有力.血常规检查提示中度贫血,血小板波动在(324～645)×109/L.肿瘤标记物、免疫全套、蛋白S(C)、雌、孕激素检查均正常.彩超检查发现左侧股浅动脉、腘动脉、右侧腘动脉血栓形成,心脏彩超无异常.初步诊断为双下肢动脉血栓形成,考虑到患者下肢缺血1个月,手术取栓效果不明确且易破坏侧枝循环加重缺血,故行抗凝溶栓治疗.%Objective To investigate the mechanism of HBx gene on cell cycle of human hepatocellular carcinoma cell line HepG2 through miR-192 regulation. Methods Flow cytometry was used to analyze the cell cycle of these three cell lines:HepG2/HBx cells( HepG2 cells stably transfected with HBx) , HepG2/pcDNA3. 1 cells ( HepG2 cells stably transfected with pcDNA3. 1) and HepG2 cells. MiR-192 expression of these cells was detected by Taqman fluorescence quantitative PCR. Following transfection with miR-192, cell cycle distribution of HepG2 cells was tested by flow cytometry, and SYBR Green quantitative PCR and Western blot were used to examine the expression of p53 and CDKNIA mRNA and protein. Results The proportion of HepG2/HBx cells was decreased significantly in G0/G1, phase [(52. 78±11. 08) ％ vs(67. 37±11. 87) ％ , (65. 08±5. 15) ％ ] , but increased remarkably in S phase and G2/M phase [S phase: ( 25. 22± 1. 84) ％ vs ( 19. 78± 1. 26) ％ , (18. 84±1. 68) ％ ; G2 /M phase : (22. 00±2. 07) ％ vs(12. 85± 1. 29) ％ . (16. 08± 1. 44) ％]. The expression of miR-192 was down-regulated in HepG2/HBx cells [(49. 1±5. 9) ％ vs ( 98. 0± 8. 9) ％ , ( 100. 0± 9. 1) ％]. Transfection of miR-192 into HepG2 cells caused cell cycle arrest at G0 /G1, and G2 /M phases. Meanwhile the mRNA( p53 :1. 68±0. 12 vs 0. 90±0. 06 ; CDKNIA : 2. 36±0. 12 vs 1. 05±0. 06)and protein( p53 : 3. 07-fold; CDKNIA : 2. 82fold) expression levels of p53 and CDKN1A genes were dramatically increased. Conclusion miR-192 could induce cell cycle arrest by augmentation of p53 and CDKN1A levels in HepG2 cells and HBx could down-regulate the expression of miR-192 to facilitate cell cycle progression.
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