HBx基因对HepG2.2.15细胞MICA-A5.1表达及侵袭、迁移的影响

李沛; 刘宇

doi:10.3969/j.issn.1001-5256.2020.04.020

HBx基因对HepG2.2.15细胞MICA-A5.1表达及侵袭、迁移的影响

DOI: 10.3969/j.issn.1001-5256.2020.04.020

李沛,
刘宇

1. 南华大学附属第一医院检验科
2. 南华大学附属第一医院急诊科

基金项目:

湖南省自然科学基金(14JJ2092)；

详细信息

中图分类号: R735.7
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出版历程
- 收稿日期: 2019-10-24
- 出版日期: 2020-04-20

Effect of hepatitis B x gene on the expression of major histocompatibility complex class I chain-related gene A-A5. 1,invasion,and migration of HepG2. 2. 15 cells

Li Pei,
Liu Yu

Department of Clinical Laboratory,The First Affiliated Hospital of Nanhua University

Research funding:

摘要

摘要: 目的探究HBx基因对HepG2. 2. 15细胞侵袭、迁移及主要组织相容性复合体（MHC）Ⅰ类链相关基因A （MICA）-A5. 1表达的影响。方法体外培养HepG2. 2. 15细胞系（插入HBV全基因组并持续表达的HepG2细胞）,随机分为对照组、HBx过表达质粒组、HBx空载质粒组、HBx siRNA组、HBx siRNA阴性对照组,分别转染质粒及siRNA后,以CKK-8法分别检测24 h、48 h后细胞增殖情况,筛选合适药物作用时间;以Transwell侵袭实验和细胞划痕实验检测各组细胞侵袭迁移能力;以免疫印记法检测HBx、MICA-A5. 1蛋白和上皮间质转化标志蛋白E-cadherin、Vimentin的表达;以酶联免疫吸附法检测各组细胞培养基中s MICA水平。计量资料多组间比较采用单因素方差分析,进一步两两比较采用SNK q检验。结果与对照组比较,24 h后,HBx过表达质粒组细胞活力升高,HBx siRNA组细胞活力降低,差异均有统计学意义（q值分别为8. 268、4. 365,P值分别为<0. 001、0. 036）; 48 h后,HBx过表达质粒组细胞活力...
- 肝炎病毒,乙型 /
- Hep G2细胞 /
- 基因,MHCⅠ类
Abstract: Objective To investigate the effect of hepatitis B x( HBx) gene on the expression of major histocompatibility complex class I chain-related gene A( MICA)-A5. 1,invasion,and migration of HepG2. 2. 15 cells. Methods HepG2. 2. 15 cells( HepG2 cells with the insertion and continuous expression of whole HBV genome) were cultured in vitro and were randomly divided into control group,HBx overexpression plasmid group,HBx empty plasmid group,HBx siRNA group,and HBx siRNA negative control group. After the transfection of plasmid or siRNA,CCK-8 assay was used to measure cell proliferation after 24 and 48 hours to screen out the appropriate duration of drug action; the Transwell invasion test and wound-healing assay were used to observe the changes in cell invasion and migration abilities;immunoblotting was used to measure the expression of HBx,MICA-A5. 1,and the marker proteins for epithelial-mesenchymal transition E-cadherin and vimentin; ELISA was used to measure the level of soluble MICA( s MICA) in cell culture medium. A one-way analysis of variance was used for comparison of continuous data between multiple groups,and the SNK q test was used for further comparison between two groups. Results Compared with the control group after 24 hours,the HBx overexpression plasmid group had a significant increase in cell viability and the HBx siRNA group had a significant reduction in cell viability( q = 8. 268 and 4. 365,P < 0. 001 and 0. 036); compared with the control group after 48 hours,the HBx overexpression plasmid group had a significant increase in cell viability and the HBx siRNA group had a significant reduction in cell viability( q = 12. 680 and 7. 523,both P < 0. 001). Compared with the control group,the HBx overexpression plasmid group had significant increases in the protein expression of HBx,MICA,and vimentin in cells,the level of s MICA in cell culture medium,the number of migrating cells,and the number of cells out of the Transwell chamber,as well as a significant reduction in the expression of E-cadherin( q = 9. 427,6. 697,10. 500,5. 042,22. 740,15. 720,and 5. 258,all P < 0. 05); the HBx siRNA group had significant reductions in the protein expression of HBx,MICA,and vimentin in cells,the level of s MICA in cell culture medium,the number of migrating cells,and the number of cells out of the Transwell chamber,as well as a significant increase in the expression of E-cadherin( q = 8. 133,8. 828,7. 616,7. 673,5. 391,7. 694,and 6. 226,all P < 0. 05). Conclusion The HBx gene regulates theexpression of MICA-A5. 1 in HepG2. 2. 15 cells and the invasion and migration of HepG2. 2. 15 cells,and upregulation of the HBx gene can promote the expression of MICA-A5. 1 and enhance the invasion and migration abilities of HepG2. 2. 15 cells.
- hepatitis B virus /
- Hep G2 cells /
- genes,MHC classⅠ

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参考文献(20)

[1] SHIRATA C,HASEGAWA K,KOKUDO T,et al. Liver resection for hepatocellular carcinoma in patients with renal dysfunction[J]. World J Surg,2018,42(12):4054-4062.

[2] ABOU-ALFA GK,MEYER T,CHENG AL,et al. Cabozantinib in patients with advanced and progressing hepatocellular carcinoma[J]. N Engl J Med,2018,379(1):54-63.

[3] GARDINI AC,FOSCHI FG,CONTI F,et al. Immune inflammation indicators and ALBI score to predict occurrence and recurrence of hepatocellular carcinoma in HCV-related cirrhosis treated with direct-acting antivirals[J]. Dig Liver Dis,2018,50(1):50-51.

[4] WANG M,XI D,NING Q. Virus-induced hepatocellular carcinoma with special emphasis on HBV[J]. Hepatol Int,2017,11(2):171-180.

[5] GAO Q,WANG K,CHEN K,et al. HBx protein-mediated ATOH1 downregulation suppresses ARID2 expression and promotes hepatocellular carcinoma.[J]. Cancer Sci,2017,108(7):1328-1337.

[6] SHI T,HUA Q,MA Z,et al. Downregulation of miR-200a-3p induced by hepatitis B virus X(HBx)protein promotes cell proliferation and invasion in HBV-infection-associated hepatocarcinoma[J]. Pathol Res Pract,2017,213(12):1464-1469.

[7] HUANG CF,HUANG CY,YEH ML,et al. Genetics variants and serum levels of MHC class I chain-related a in predicting hepatocellular carcinoma development in chronic hepatitis C patients post antiviral treatment[J]. Ebiomedicine,2017,15:81-89.

[8] HUANG CF,WANG SC,YEH ML,et al. Association of serial serum major histocompatibility complex class I chain-related A measurements with hepatocellular carcinoma in chronic hepatitis C patients after viral eradication[J]. J Gastroenterol Hepatol,2019,34(1):249-255.

[9] SUN D,WANG X,ZHANG H,et al. MMP9 mediates MICA shedding in human osteosarcomas[J]. Cell Biol Int,2011,35(6):569-574.

[10] HEIMBACH JK,KULIK LM,FINN RS,et al. AASLD guidelines for the treatment of hepatocellular carcinoma[J]. Hepatology,2018,67(1):358-380.

[11] Cancer Genome Atlas Research Network. Comprehensive and integrative genomic characterization of hepatocellular carcinoma[J]. Cell,2017,169(7):1327-1341. e23.

[12] QIN G,DANG M,GAO H,et al. Deciphering the proteinprotein interaction network regulating hepatocellular carcinoma metastasis[J]. Biochim Biophys Acta Proteins Proteom,2017,1865(9):1114-1122.

[13] CHEUNG KS,SETO WK,WONG DK,et al. Relationship between HBsAg,HBcrAg and hepatocellular carcinoma in patients with undetectable HBV DNA under nucleos(t)ide therapy[J]. J Viral Hepat,2017,24(8):654-661.

[14] WANG X,OISHI N,SHIMAKAMI T,et al. Hepatitis B virus X protein induces hepatic stem cell-like features in hepatocellular carcinoma by activating KDM5B[J]. World J Gastroenterol,2017,23(18):3252-3261.

[15] YANG Q,LI XP,ZHONG YB,et al. Interferon-αinhibits cell migration and invasion and induces the expression of antiviral proteins in Huh-7 cells transfected with hepatitis B virus X gene-expressing lentivirus[J]. Exp Ther Med,2017,14(6):5924-5930.

[16] BRABLETZ T,KALLURI R,NIETO MA,et al. EMT in cancer[J]. Nat Rev Cancer,2018,18(2):128-134.

[17] SABA NF,WANG Y,FU H,et al. Association of cytoplasmic CXCR4 with loss of epithelial marker and activation of ERK1/2and AKT signaling pathways in non-small-cell lung cancer[J]. Clin Lung Cancer,2017,18(3):e203-e210.

[18] HAI H,TAMORI A,THUY L,et al. Polymorphisms in MICA,but not in DEPDC5,HCP5 or PNPLA3,are associated with chronic hepatitis C-related hepatocellular carcinoma[J]. Sci Rep,2017,7(1):11912.

[19] ARAI J,GOTO K,STEPHANOU A,et al. Predominance of regorafenib over sorafenib:Restoration of membrane-bound MICA in hepatocellular carcinoma cells[J]. J Gastroenterol Hepatol,2018,33(5):1075-1081.

[20] CHEN J,XU H,ZHU XX. Abnormal expression levels of sMICA and NKG2D are correlated with poor prognosis in pancreatic cancer[J]. Ther Clin Risk Manag,2016,12:11-18.

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