Jarid1b对食管鳞癌KYSE-150细胞增殖及分化的影响
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摘要
目的研究去甲基化酶Jarid1b对食管鳞癌细胞增殖及分化的影响,为食管鳞癌治疗提供新的靶点。方法采用实时荧光定量PCR(RT-qPCR)以及Western-blot方法检测低分化食管鳞癌细胞系KYSE-150和高分化食管鳞癌细胞系TE-1中去甲基化酶Jarid1b、细胞角蛋白10(CK10)、细胞角蛋白13(CK13)的表达。采用慢病毒转染的方式获得能够在药物诱导时过表达Jarid1b的KYSE-150稳转细胞系,应用CCK8细胞毒性实验检测过表达Jarid1b对该细胞系增殖能力的影响;采用RT-qPCR和Western-blot方法检测过表达Jarid1b后细胞系CK10和CK13表达量的变化。结果 RT-qPCR检测结果表明,和低分化KYSE-150细胞系相比,高分化TE-1细胞系中CK10和CK13高表达(t=22.07、38.44,P<0.05);Western-blot检测结果表明,和KYSE-150细胞系相比,TE-1细胞系中Jarid1b的蛋白表达水平明显增高。CCK8实验结果显示,与未诱导的KYSE-150细胞系相比,诱导KYSE-150细胞系过表达Jarid1b24、48h后,该细胞的增殖能力明显下降(t=10.94、16.71,P<0.05)。RT-qPCR和Western-blot方法检测结果表明,诱导KYSE-150细胞系过表达Jarid1b以后,与未诱导的KYSE-150细胞系相比CK10和CK13的表达明显增高(t=9.706、5.23,P<0.05)。结论去甲基化酶Jarid1b过表达可以抑制低分化食管鳞癌细胞KYSE-150的增殖并促进其分化。
Objective To investigate the effect of Jarid1b,a demethylase,on the proliferation and differentiation of esophageal squamous cell carcinoma(ESCC)cells,and to provide a new target for the treatment of ESCC. Methods RT-qPCR and Western-blot were used to measure the expression of Jarid1b,cytokeratin 10(CK10),and cytokeratin 13(CK13)in poorly differentiated ESCC cell line KYSE-150 and well-differentiated ESCC cell line TE-1.KYSE-150 cells with stable overexpression of Jarid1b when induced by drugs were obtained by lentivirus transfection.CCK-8 cytotoxicity assay was used to analyze the effect of Jarid1b overexpression on the proliferative capacity of this cell line,and RT-qPCR and Western blot were used to measure the changes in the expression of CK10 and CK13 in this cell line after Jarid1b overexpression. Results RT-qPCR data showed that welldifferentiated TE-1 cells had higher expression of CK10 and CK13 than poorly differentiated KYSE-150 cells(t=22.07,38.44,P<0.05).Western blot showed that compared with KYSE-150 cells,TE-1 cells had a significant increase in the protein expression of Jarid1b.The CCK-8 assay showed that compared with the uninduced KYSE-150 cells,the induced KYSE-150 cells had a significant reduction in proliferative capacity at 24 and 48 hours after Jarid1b overexpression(t=10.94,16.71,P<0.05).RT-qPCR and Western blot showed that after Jarid1b overexpression,the induced KYSE-150 cells had significant increases in the expression of CK10 and CK13 compared with the uninduced KYSE-150 cells(t=9.706,5.23,P<0.05). Conclusion Overexpression of the demethylase Jarid1b can inhibit the proliferation and promote the differentiation of poorly differentiated ESCC KYSE-150 cells.
Objective To investigate the effect of Jarid1b,a demethylase,on the proliferation and differentiation of esophageal squamous cell carcinoma(ESCC)cells,and to provide a new target for the treatment of ESCC. Methods RT-qPCR and Western-blot were used to measure the expression of Jarid1b,cytokeratin 10(CK10),and cytokeratin 13(CK13)in poorly differentiated ESCC cell line KYSE-150 and well-differentiated ESCC cell line TE-1.KYSE-150 cells with stable overexpression of Jarid1b when induced by drugs were obtained by lentivirus transfection.CCK-8 cytotoxicity assay was used to analyze the effect of Jarid1b overexpression on the proliferative capacity of this cell line,and RT-qPCR and Western blot were used to measure the changes in the expression of CK10 and CK13 in this cell line after Jarid1b overexpression. Results RT-qPCR data showed that welldifferentiated TE-1 cells had higher expression of CK10 and CK13 than poorly differentiated KYSE-150 cells(t=22.07,38.44,P<0.05).Western blot showed that compared with KYSE-150 cells,TE-1 cells had a significant increase in the protein expression of Jarid1b.The CCK-8 assay showed that compared with the uninduced KYSE-150 cells,the induced KYSE-150 cells had a significant reduction in proliferative capacity at 24 and 48 hours after Jarid1b overexpression(t=10.94,16.71,P<0.05).RT-qPCR and Western blot showed that after Jarid1b overexpression,the induced KYSE-150 cells had significant increases in the expression of CK10 and CK13 compared with the uninduced KYSE-150 cells(t=9.706,5.23,P<0.05). Conclusion Overexpression of the demethylase Jarid1b can inhibit the proliferation and promote the differentiation of poorly differentiated ESCC KYSE-150 cells.
引文
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[2]李鹏,王拥军,陈光勇,等.中国早期食管鳞状细胞癌及癌前病变筛查与诊治共识(2015年·北京)[J].中国实用内科杂志,2016(1):20-33.
[3]MA J,GUO X,ZHANG S,et al.Trichostatin A,a histone deacetylase inhibitor suppresses proliferation and promotes apoptosis of esophageal squamous cell lines[J].Mol Med Rep,2015,11(6):4525-4531.
[4]LIU J S,WANG L,MAO W M,et al.Progress of surgery for esophageal carcinoma[J].China Cancer,2013,22(12):953-960.
[5]夏娟,刘建平,张永恒,等.食管鳞癌中血管内皮生长因子-C的表达及与淋巴管生成的关系[J].现代肿瘤医学,2016(5):749-753.
[6]张立伟,马骏,金悦,等.KDM5B基因在卵巢癌中的表达及其对卵巢癌耐药能力的影响[J].现代妇产科进展,2017,26(3):161-164.
[7]ZHAO L H,LIU H G.Immunohistochemical detection and clinicopathological significance of JARID1B/KDM5Band P16expression in invasive ductal carcinoma of the breast[J].Genet Mol Res,2015,14(2):5417-5426.
[8]SHEN X,ZHUANG Z,ZHANG Y,et al.JARID1B modulates lung cancer cell proliferation and invasion by regulating p53expression[J].Tumour Biol,2015,36(9):7133-7142.
[9]TANG B,QI G,TANG F,et al.JARID1Bpromotes metastasis and epithelial-mesenchymal transition via PTEN/AKT signaling in hepatocellular carcinoma cells[J].Oncotarget,2015,6(14):12723-12739.
[10]WANG Z,TANG F,QI G,et al.KDM5Bis overexpressed in gastric cancer and is required for gastric cancer cell proliferation and metastasis[J].Am J Cancer Res,2014,5(1):87-100.
[11]TARNOWSKI M,CZEREWATY M,DESKUR A,et al.Expression of cancer testis antigens in colorectal cancer:new prognostic and therapeutic implications[J].Dis Markers,2016(16):1987505.
[12]CATCHPOLE S,SPENCER-DENE B,HALL D,et al.PLU-1/JARID1B/KDM5Bis required for embryonic survival and contributes to cell proliferation in the mammary gland and in ER+breast cancer cells[J].Int J Oncol,2011,38(5):1267-1277.
[13]YAMANE K,TATEISHI K,KLOSE R J,et al.PLU-1is an H3K4 demethylase involved in transcriptional repression and breast cancer cell proliferation[J].Mol Cell,2007,25(6):801-812.
[14]HAYAMI S,YOSHIMATSU M,VEERAKUMARASIVAM A,et al.Overexpression of the JmjC histone demethylase KDM5Bin human carcinogenesis:involvement in the proliferation of cancer cells through the E2F/RB pathway[J].Mol Cancer,2010,9(1):59.
[15]BAMODU O A,HUANG W C,LEE W H,et al.Aberrant KDM5Bexpression promotes aggressive breast cancer through MALAT1overexpression and downregulation of hsa-miR-448[J].BMC Cancer,2016,16(1):160.
[16]LI Q,SHI L,GUI B,et al.Binding of the JmjCdemethylase JARID1Bto LSD1/NuRD suppresses angiogenesis and metastasis in breast cancer cells by repressing chemokine CCL14[J].Cancer Res,2011,71(21):6899-6908.
[17]ROJAS A,AGUILAR R,HENRIQUEZ B,et al.Epigenetic control of the bone-master runx2gene during osteoblast-lineage commitment by the histone demethylase JARID1B/KDM5B[J].J Biol Chem,2015,290(47):28329-28342.
[18]POSPISIL V,VARGOVA K,KOKAVEC J,et al.Epigenetic silencing of the oncogenic miR-17-92cluster during PU.1-directed macrophage differentiation[J].EMBO J,2011,30(21):4450-4464.
[19]ZHOU Q,OBANA E A,RADOMSKI K L,et al.Inhibition of the histone demethylase Kdm5bpromotes neurogenesis and derepresses Reln(reelin)in neural stem cells from the adult subventricular zone of mice[J].Mol Biol Cell,2016,27(4):627-639.
[20]KIDDER B L,HU G,YU Z X,et al.Extended self-renewal and accelerated reprogramming in the absence of Kdm5b[J].Mol Cell Biol,2013,33(24):4793-4810.
[21]WONG S H,GOODE D L,IWASAKI M,et al.The H3K4-methyl epigenome regulates leukemia stem cell oncogenic potential[J].Cancer Cell,2015,28(2):198-209.
[22]KIDDER B L,HU G,ZHAO K.KDM5Bfocuses H3K4 methylation near promoters and enhancers during embryonic stem cell self-renewal and differentiation[J].Genome Biol,2014,15(2):R32.
[23]尹文华,刘素琴,陈志凌,等.细胞角蛋白4和细胞角蛋白13在鼻腔-鼻窦鳞状细胞癌中的表达及临床意义[J].中国耳鼻咽喉头颈外科,2016,23(2):98-101.
[24]ZHANG J,AN X,HAN Y,et al.Overexpression of JARID1Bpromotes differentiation via SHIP1/AKT signaling in human hypopharyngeal squamous cell carcinoma[J].Cell Death Dis,2016,7(9):e2358.