β干扰素联合全反式维甲酸通过抑制JAK2/STAT3信号通路抑制HepG2人肝癌细胞增殖并促进其凋亡
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摘要
目的研究β干扰素(IFN-β)和全反式维甲酸(ATRA)联合应用对Hep G2人肝癌细胞增殖和凋亡的影响,并探讨Janus激酶2/信号转导子与转录激活子3(JAK2/STAT3)通路在其中可能的机制。方法分别用1000 U/m L IFN-β、10μmol/L ATRA及1000 U/m L IFN-β联合10μmol/L ATRA处理Hep G2细胞24 h,采用MTT法检测Hep G2细胞的增殖抑制率,流式细胞术检测细胞凋亡。Western blot法检测磷酸化的Janus激酶2(p-JAK2)和磷酸化的信号转导子与转录激活子3(p-STAT3)、维甲酸-干扰素诱导死亡相关基因19(GRIM-19)、Bcl-2、Bcl-xl、Bax蛋白的表达。结果 IFN-β、ATRA作用于细胞后,Hep G2细胞增殖受到抑制,同时诱导细胞发生凋亡,IFN-β联合ATRA联合处理作用更强;Hep G2细胞中p-JAK2和p-STAT3蛋白的表达在IFN-β或ATRA作用下减弱,GRIM-19和Bax蛋白表达升高。IFN-β联合ATRA处理作用更强。结论 IFN-β联合ATRA通过抑制JAK2/STAT3信号通路抑制Hep G2人肝癌细胞的增殖并促进其凋亡。
Objective To investigate the effect of interferon-β( IFN-β) combined with all-trans retinoic acid( ATRA) on the proliferation and apoptosis of Hep G2 human hepatocarcinoma cells and the role of Janus kinase 2 / signal transducer and activator of transcription 3( JAK2 / STAT3) signal pathway in the process. Methods Hep G2 cells were randomly divided intro three groups and treated with 1000 U / m L IFN-β,10 μmol / L ATRA and 1000 U / m L IFN-β combined with 10 μmol / L ATRA,respectively for 24 hours. Cell viability was measured by MTT assay and apoptosis rate was detected by flow cytometry.Western blotting was applied to detect the protein levels of p-JAK2, p-STAT3, gene associated with retinoidinterferon-induced mortality-19( GRIM-19),Bcl-2,Bcl-xl and Bax. Results IFN-β or ATRA inhibited the proliferation and induced the apoptosis of Hep G2 cells. The effect was enhanced when IFN-β was combined with ATRA. The expressions of p-JAK2 and p-STAT3 were down-regulated while the expressions of GRIM-19 and Bax were up-regulated after treated with IFN-β or ATRA on Hep G2 cells,especially the combination of IFN-β and ATRA. Conclusion Combination of IFN-β and ATRA could suppress the proliferation and induced the apoptosis of Hep G2 hepatocarcinoma cells by inhibiting JAK2 / STAT3 signal pathway.
Objective To investigate the effect of interferon-β( IFN-β) combined with all-trans retinoic acid( ATRA) on the proliferation and apoptosis of Hep G2 human hepatocarcinoma cells and the role of Janus kinase 2 / signal transducer and activator of transcription 3( JAK2 / STAT3) signal pathway in the process. Methods Hep G2 cells were randomly divided intro three groups and treated with 1000 U / m L IFN-β,10 μmol / L ATRA and 1000 U / m L IFN-β combined with 10 μmol / L ATRA,respectively for 24 hours. Cell viability was measured by MTT assay and apoptosis rate was detected by flow cytometry.Western blotting was applied to detect the protein levels of p-JAK2, p-STAT3, gene associated with retinoidinterferon-induced mortality-19( GRIM-19),Bcl-2,Bcl-xl and Bax. Results IFN-β or ATRA inhibited the proliferation and induced the apoptosis of Hep G2 cells. The effect was enhanced when IFN-β was combined with ATRA. The expressions of p-JAK2 and p-STAT3 were down-regulated while the expressions of GRIM-19 and Bax were up-regulated after treated with IFN-β or ATRA on Hep G2 cells,especially the combination of IFN-β and ATRA. Conclusion Combination of IFN-β and ATRA could suppress the proliferation and induced the apoptosis of Hep G2 hepatocarcinoma cells by inhibiting JAK2 / STAT3 signal pathway.
引文
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[21]Wung B S,Hsu M C,Wu C C,et al.Resveratrol suppresses IL-6-induced ICAM-1 gene expression in endothelial cells:effects on the inhibition of STAT3 phosphorylation[J].Life Sci,2005,78(4):389-397.
[22]Kalakonda S,Nallar S C,Lindner D J,et al.Tumor-suppressive activity of the cell death activator GRIM-19 on a constitutively active signal transducer and activator of transcription 3[J].Cancer Res,2007,67(13):6212-6220.
[23]Li F,Ren W,Zhao Y,et al.Downregulation of GRIM-19 is associated with hyperactivation of p-STAT3 in hepatocellular carcinoma[J].Med Oncol,2012,29(5):3046-3054.
[24]冯其柱,张超,李方跃,等.肝细胞癌组织GRIM-19和STAT3蛋白的表达[J].安徽医科大学学报,2011,46(5):419-422.
[25]Lippman S M,Parkinson D R,Itri L M,et al.13-cis-retinoic acid and interferon alpha-2a:effective combination therapy for advanced squamous cell carcinoma of the skin[J].J Natl Cancer Inst,1992,84(4):235-241.
[26]Lancillotti F,Giandomenico V,Affabris E,et al.Interferon alpha-2b and retinoic acid combined treatment affects proliferation and gene expression of human cervical carcinoma cells[J].Cancer Res,1995,55(14):3158-3164.
[27]Motzer R J,Schwartz L,Law T M,et al.Interferon alfa-2a and13-cis-retinoic acid in renal cell carcinoma:anti tumor activity in a phraseⅡtrial and interactions in vitro[J].J Clin Oncol,1995,13(8):1950-1957.
[28]Bhatt S,Qin J,Bennett C,et al.All-trans retinoic acid induces arginase-1 and inducible nitric oxide synthase-producing dendritic cells with T cell inhibitory function[J].J Immunol,2014,192(11):5098-5108.
[29]Szabo A,Osman R M,Bacskai I,et al.Temporally designed treatment of melanoma cells by ATRA and poly I∶C results in enhanced chemokine and IFNβsecretion controlled differently by TLR3 and MDA5[J].Melanoma Res,2012,22(5):351-361.
[2]蒋树龙,花宝金.JAK2/STAT3/SOCS3信号通路与肿瘤转移[J].中国肿瘤生物治疗杂志,2014,21(6):698-702.
[3]赵冀安,刘文聪,孙会凤,等.JAK2/STAT3信号通路在肝细胞性肝癌中表达及意义[J].中国普通外科杂志,2016,25(1):83-89.
[4]Kim M S,Lee W S,Jeong J,et al.Induction of metastatic potential by Trk B via activation of IL6/JAK2/STAT3 and PI3K/AKT signaling in breast cancer[J].Oncotarget,2015,6(37):40158-40171.
[5]Li L,Li H,Li M.Curcumin protects against cerebral ischemiareperfusion injury by activating JAK2/STAT3 signaling pathway in rats[J].Int J Clin Exp Med,2015,8(9):14985-14991.
[6]龚代鹏,张竹青,张妍,等.p-STAT3、Survivin和Mcl-1蛋白在胃癌组织中的表达及意义[J].中华临床医师杂志,2015,9(6):915-920.
[7]Wu N,Liu J,Zhao X,et al.Cardamonin induces apoptosis by suppressing STAT3 signaling pathway in glioblastoma stem cells[J].Tumour Biol,2015,36(12):9667-9676.
[8]李永光,朱伟,魏盟.GRIM-19与STAT3蛋白作用及相关机制研究进展[J].医学研究杂志,2013,42(9):157-159.
[9]Li M,Li Z,Liang C,et al.Upregulation of GRIM-19 suppresses the growth of oral squamous cell carcinoma in vitro and in vivo[J].Oncol Rep,2014,32(5):2183-2190.
[10]Zhang W,Du Y,Jiang T,et al.Upregulation of GRIM-19 inhibits the growth and invasion of human breast cancer cells[J].Mol Med Rep,2015,12(2):2919-2925.
[11]Wang Q,Mao-Draayer Y.Interferon beta(IFN-β)treatment exerts potential neuroprotective effects through neurotrophic factors and novel neurotensin/neurotensin high affinity receptor 1 pathway[J].Neural Regen Res,2015,10(12):1932-1933.
[12]Shakado S,Iwata K,Tsuchiya N,et al.Pilot study of hepatic arterial infusion chemotherapy with interferon-beta and 5-fluorouracil:A new chemotherapy for patients with advanced hepatocellular carcinoma[J].Hepatogastroenterology,2014,61(131):557-562.
[13]Kuroda S,Ishikawa T,Houkin K,et al.Incidence and clinical features of disease progression in adult moyamoya disease[J].Stroke,2005,36(10):2148-2153.
[14]Zou C,Ramakumar S,Qian L,et al.Effect of retinoic acid and interferon alpha-2a on transitional cell carcinoma of bladder[J].J Urol,2005,173(1):247-251.
[15]林秀英,韩振东,尹淑红,等.阿维A酸联合干扰素诱导皮肤鳞癌细胞凋亡作用的实验研究[J].黑龙江医药科学,2012,35(5):54-55.
[16]王威,徐旭光,吕智勇,等.全反式维甲酸和α-干扰素对人涎腺腺样囊性癌SACC-83细胞株联合作用的研究[J].临床口腔医学杂志,2004,20(1):7-9.
[17]Murphy D,Detjen K M,Welzel M,et al.Interferon alpha delays S-phase progression in human hepatocellular carcinoma cells via inhibition of specific cyclin-dependent kinases[J].Hepatology,2001,33(2):346-356.
[18]Obora A,Shiratori Y,Okuno M,et al.Synergistic induction of apoptosis by acyclic retinoid and interferon-βin human hepatocellular carcinoma cells[J].Hepatology,2002,36(5):1115-1124.
[19]Rajasekaran D,Srivastava J,Ebeid K,et al.Combination of nanoparticledelivered siRNA for astrocyte elevated gene-1(AEG-1)and all-trans retinoic acid(ATRA):An effective therapeutic strategy for hepatocellular carcinoma(HCC)[J].Bioconjug Chem,2015,26(8):1651-1661.
[20]Booz G W,Day J N,Baker K M.Interplay between the cardiac rennin angiotensin system and JAK-STAT signaling:role in cardiac hypertrophy,ischemia/reperfusion dysfunction,and heart failure[J].J Mol Cell Cardiol,2002,34(11):1443-1453.
[21]Wung B S,Hsu M C,Wu C C,et al.Resveratrol suppresses IL-6-induced ICAM-1 gene expression in endothelial cells:effects on the inhibition of STAT3 phosphorylation[J].Life Sci,2005,78(4):389-397.
[22]Kalakonda S,Nallar S C,Lindner D J,et al.Tumor-suppressive activity of the cell death activator GRIM-19 on a constitutively active signal transducer and activator of transcription 3[J].Cancer Res,2007,67(13):6212-6220.
[23]Li F,Ren W,Zhao Y,et al.Downregulation of GRIM-19 is associated with hyperactivation of p-STAT3 in hepatocellular carcinoma[J].Med Oncol,2012,29(5):3046-3054.
[24]冯其柱,张超,李方跃,等.肝细胞癌组织GRIM-19和STAT3蛋白的表达[J].安徽医科大学学报,2011,46(5):419-422.
[25]Lippman S M,Parkinson D R,Itri L M,et al.13-cis-retinoic acid and interferon alpha-2a:effective combination therapy for advanced squamous cell carcinoma of the skin[J].J Natl Cancer Inst,1992,84(4):235-241.
[26]Lancillotti F,Giandomenico V,Affabris E,et al.Interferon alpha-2b and retinoic acid combined treatment affects proliferation and gene expression of human cervical carcinoma cells[J].Cancer Res,1995,55(14):3158-3164.
[27]Motzer R J,Schwartz L,Law T M,et al.Interferon alfa-2a and13-cis-retinoic acid in renal cell carcinoma:anti tumor activity in a phraseⅡtrial and interactions in vitro[J].J Clin Oncol,1995,13(8):1950-1957.
[28]Bhatt S,Qin J,Bennett C,et al.All-trans retinoic acid induces arginase-1 and inducible nitric oxide synthase-producing dendritic cells with T cell inhibitory function[J].J Immunol,2014,192(11):5098-5108.
[29]Szabo A,Osman R M,Bacskai I,et al.Temporally designed treatment of melanoma cells by ATRA and poly I∶C results in enhanced chemokine and IFNβsecretion controlled differently by TLR3 and MDA5[J].Melanoma Res,2012,22(5):351-361.