RNA聚合酶Ⅰ抑制剂通过NF-κb调控鼻咽癌细胞系CNE-1的增殖、侵袭与凋亡
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摘要
目的探讨RNA聚合酶Ⅰ抑制剂CX-5461对人鼻咽癌细胞CNE-1增殖、侵袭与凋亡的作用机制。方法免疫组织化学染色检测鼻咽癌与癌旁组织NF-κb的表达。Western blot检测相关效应蛋白在蛋白质水平的表达。CCK-8法检测不同浓度药物处理后CNE-1细胞系的增殖情况。Transwell法比较药物处理后CNE-1细胞系的侵袭能力变化。流式细胞术检测不同浓度药物处理后鼻咽癌细胞系CNE-1细胞周期和凋亡的变化。结果鼻咽癌组织中磷酸化NF-κb表达较癌旁组织明显增高(P<0.01)。NF-κb总蛋白在蛋白水平无明显变化,但磷酸化水平在药物处理后明显降低(P<0.01)。Bax较对照组明显增高(P<0.01),Bcl-2较对照组明显降低(P<0.01)。药物处理后,发现CNE-1细胞系的增殖、侵袭能力较对照组明显降低且差异具有统计学意义(均P<0.01);细胞凋亡较对照组明显增多(P<0.01),药物处理组较对照组细胞周期明显阻滞在G_2期(P<0.01)。结论 RNA聚合酶Ⅰ抑制剂CX-5461能够通过调控NF-κb磷酸化促进鼻咽癌CNE-1细胞系凋亡及周期阻滞,抑制其增殖与侵袭能力。
Objective To investigate the effect of RNA polymerase Ⅰ inhibitor CX-5461 on the proliferation,invasion and apoptosis of human nasopharyngeal carcinoma cell line CNE-1. Methods Immunohistochemic al staining was used to determine the difference in phosphorylated NF-κb expression between nasopharyngeal carcinoma and paracancerous tissues. Western blot was used to detect the expression of related effector proteins at the protein level. The CCK-8 method was used to detect the proliferation of CNE-1 cell line treated with different concentrations of drugs. Transwell method was applied to detect the invasive ability of CNE-1 cell line after drug treatment. Flow cytometry was used to detect cell cycle and apoptosis of nasopharyngeal carcinoma cell line CNE-1 treated with different concentrations of drugs. Results Immunohistochemical staining showed that the expression of NF-κb in nasopharyngeal carcinoma tissues was significantly higher than that in adjacent tissues(P<0.01). The total protein of NF-κb did not change significantly at the protein level, but the phosphorylation level and Bcl-2 were significantly decreased, while Bax was significantly increased after drug treatment(all P<0.01). After drug treatment, the proliferation and invasion abilities of CNE-1 cell line were significantly lower than that of the control group detected by CCK-8(P<0.01) and Transwell(P<0.01); Hoechst fluorescence staining showed that cell apoptosis was significantly higher than that of the control group(P<0.01); flow cytometry showed that cell cycle of the drug-administered group was significantly blocked in the G_2 phase(P<0.01). Conclusion RNA polymerase Ⅰ inhibitor CX-5461 could promote the apoptosis and cycle arrest, and inhibit the proliferation and invasion abilities of nasopharyngeal carcinoma CNE-1 cell line by regulating NF-κb phosphorylation.
Objective To investigate the effect of RNA polymerase Ⅰ inhibitor CX-5461 on the proliferation,invasion and apoptosis of human nasopharyngeal carcinoma cell line CNE-1. Methods Immunohistochemic al staining was used to determine the difference in phosphorylated NF-κb expression between nasopharyngeal carcinoma and paracancerous tissues. Western blot was used to detect the expression of related effector proteins at the protein level. The CCK-8 method was used to detect the proliferation of CNE-1 cell line treated with different concentrations of drugs. Transwell method was applied to detect the invasive ability of CNE-1 cell line after drug treatment. Flow cytometry was used to detect cell cycle and apoptosis of nasopharyngeal carcinoma cell line CNE-1 treated with different concentrations of drugs. Results Immunohistochemical staining showed that the expression of NF-κb in nasopharyngeal carcinoma tissues was significantly higher than that in adjacent tissues(P<0.01). The total protein of NF-κb did not change significantly at the protein level, but the phosphorylation level and Bcl-2 were significantly decreased, while Bax was significantly increased after drug treatment(all P<0.01). After drug treatment, the proliferation and invasion abilities of CNE-1 cell line were significantly lower than that of the control group detected by CCK-8(P<0.01) and Transwell(P<0.01); Hoechst fluorescence staining showed that cell apoptosis was significantly higher than that of the control group(P<0.01); flow cytometry showed that cell cycle of the drug-administered group was significantly blocked in the G_2 phase(P<0.01). Conclusion RNA polymerase Ⅰ inhibitor CX-5461 could promote the apoptosis and cycle arrest, and inhibit the proliferation and invasion abilities of nasopharyngeal carcinoma CNE-1 cell line by regulating NF-κb phosphorylation.
引文
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[2]Wei F,Ding L,Wei Z,et al.Ribosomal protein L34 promotes the proliferation,invasion and metastasis of pancreatic cancer cells[J].Oncotarget,2016,7(51):85259-72.
[3]李敏,周兴涛.真核生物核糖体RNA基因的转录调控[J].江西农业学报,2008,20(11):14-16.[Li M,Zhou XT.Transcriptional regulation of ribosomal RNA genes in eukaryotes[J].Jiangsu Nong Ye Xue Bao,2008,20(11):14-6.]
[4]Zhou X,Hao Q,Liao J,et al.Ribosomal protein S14 unties the MDM2-p53 loop upon ribosomal stress[J].Oncogene,2013,32(3):388-96.
[5]Yang M,Sun H,He J,et al.Interaction of ribosomal protein L22with casein kinase 2α:a novel mechanism for understanding the biology of non-small cell lung cancer[J].Oncol Rep,2014,32(1):139-44.
[6]Scala F,Brighenti E,Govoni M,et al.Direct relationship between the level of p53 stabilization induced by rRNA synthesisinhibiting drugs and the cell ribosome biogenesis rate[J].Oncogene,2016,35(8):977-89.
[7]何薇.鳜NKAP和VIP的初步功能研究[D].中山大学,2009.[He W.Preliminary functional study of Squid NKAP and VIP[D].Sun Yat-Sen University,2009.]
[8]Li L,Li Y,Zhao J,et al.CX-5461 induces autophagy and inhibits tumor growth via mammalian target of rapamycin-related signaling pathways in osteosarcoma[J].Onco Targets Ther,2016,9:5985-97.
[9]DuoY,Yang M,Du Z,et al.CX-5461-loaded nucleolus-targeting nanoplatform for cancer therapy through induction of pro-death autophagy[J].Acta Biomater,2018,79:317-30.
[10]Khan M,Shah SA,Kim MO.17β-Estradiol via SIRT1/Acetyl-p53/NF-κb Signaling Pathway Rescued Postnatal Rat Brain Against Acute Ethanol Intoxication[J].Mol Neurobiol,2018,55(4):3067-78.
[11]Zhang DX,Ma DY,Yao ZQ,et al.ERK1/2/p53 and NF-κBdependent-PUMA activation involves in doxorubicin-induced cardiomyocyte apoptosis[J].Eur Rev Med Pharmacol Sci,2016,20(11):2435-42.
[12]Sen R,Baltimore D.Multiple nuclear factors interact with the immunoglobulin enhancer sequences[J].Cell,1986,46(5):705-16.
[13]Helbig G,Christopherson KW,Bhat-Nakshatri P,et al.NF-kappa Bpromotes breastcancer cell migration andmetastasis by inducing the expression of thechemokine receptor CXCR4[J].J Biol Chem,2003,278(24):21631-8.
[14]Hinz M,Krappmann D,Eichten A,et al.NF-κB Function in Growth Control:Regulation of Cyclin D1 Expression and G0/G1-to-S-Phase Transition[J].Mol Cell Biol,1999,19(4):2690-8.
[15]Rao S,Lee SY,Gutierrez A,et a1.Inactivation of ribosomal protein L22 promotes transformation by induction of the stemness factor,Lin28B[J].Blood,2012,120(18):3764-73.