[1]陈施翰,邓青松,蒋家云,等.LncRNA RNU12上调热休克蛋白72抑制重组人TRAIL蛋白诱导肝癌细胞凋亡[J].第三军医大学学报,2017,39(09):833-839.
 Chen Shihan,Deng Qingsong,Jiang Jiayun,et al.LncRNA RNU12 inhibits apoptosis of hepatocellular carcinoma cells induced by recombinant human TRAIL protein through up-regulating heat shock protein 72[J].J Third Mil Med Univ,2017,39(09):833-839.
点击复制

LncRNA RNU12上调热休克蛋白72抑制重组人TRAIL蛋白诱导肝癌细胞凋亡(/HTML )
分享到:

《第三军医大学学报》[ISSN:1000-5404/CN:51-1095/R]

卷:
39卷
期数:
2017年第09期
页码:
833-839
栏目:
基础医学
出版日期:
2017-05-15

文章信息/Info

Title:
LncRNA RNU12 inhibits apoptosis of hepatocellular carcinoma cells induced by recombinant human TRAIL protein through up-regulating heat shock protein 72
作者:
陈施翰邓青松蒋家云邹孟达谭运华马宽生
第三军医大学西南医院全军肝胆外科研究所
Author(s):
Chen Shihan Deng Qingsong Jiang Jiayun Zou Mengda Tan Yunhua Ma Kuansheng

Institute of Hepatobiliary Surgery, Southwest Hospital, Third Military Medical University, Chongqing, 400038, China

关键词:
肝癌细胞凋亡长链非编码RNA亚致死性热温度HSP72
Keywords:
apoptosis of hepatocellular carcinoma cells long non-coding RNA sub-lethal heat temperature HSP72
分类号:
R394.2; R454.5; R735.7
文献标志码:
A
摘要:

目的      探讨46~50 ℃时抑制重组人肿瘤坏死因子相关凋亡诱导配体(tumor necrosis factor-related apoptosis inducing ligand,TRAIL)蛋白促肝癌细胞凋亡相关分子机制。方法      建立肝癌细胞亚致死性热温度模型,在不同温度(43、46、50 ℃)条件下分别处理10 min,Annexin-V/PI染色检测重组人TRAIL蛋白对肝癌细胞凋亡率的影响,以常温37 ℃作为对照组。根据前期肝癌细胞Agilent Human lncRNA+mRNA Array V4.0芯片表达谱数据,选定50 ℃时特异性高表达的LncRNA RNU12作为研究对象。构建LncRNA RNU12-shRNA慢病毒干扰载体,转染MHCC97-H细胞。qRT-PCR及Western blot检测感染前后不同温度组中热休克蛋白72(heat shock protein 72,HSP72) mRNA和蛋白的表达变化。利用流式细胞技术检测LncRNA RNU12慢病毒干扰载体对TRAIL蛋白诱导 细胞凋亡的影响。采用荧光原位杂交(FISH)技术检测LncRNA RNU12亚细胞定位情况。结果      温度在43 ℃时促进重组人TRAIL蛋白诱导肝癌细胞凋亡的功能,46~50 ℃时抑制重组人TRAIL蛋白诱导肝癌细胞凋亡的功能。HSP72 mRNA在4个温度组中的表达均较干扰前明显降低(P<0.01);在蛋白水平,37、46、50 ℃组较干扰前显著降低(P<0.05)。转染后,46、50 ℃条件下TRAIL蛋白诱导MHCC97-H细胞凋亡分别上升7.36%、8.84%(P<0.01)。利用RNA-FISH技术确定50 ℃时LncRNA RNU12主要在MHCC97-H细胞细胞核中表达。结论      LncRNA RNU12在转录水平上调HSP72表达可能是46~50 ℃时抑制重组人TRAIL蛋白促肝癌细胞凋亡分子机制之一。

Abstract:

Objective      To investigate the molecular mechanism of inhibiting the apoptosis of hepatocellular carcinoma (HCC) cells induced by recombinant human tumor necrosis factor-related apoptosis inducing ligand (TRAIL) protein at 46~50 ℃. Methods       A sub-lethal heat treatment model of HCC cells was established. The Annexin-V/PI staining was used to detect the effect of treatment of recombinant human TRAIL on the apoptotic rate of HCC cells under different temperatures (43, 46 and 50 ℃, and 37 ℃ as control temperature) for 10 min. According to the expression profile from the Agilent human lncRNA+mRNA Array V4.0 microarray data in HCC cells in our previous study, 50 ℃ specific highly expressed LncRNA RNU12 was selected as the research object. Lentiviral vector  LncRNA RNU12 was constructed, and then transfected into MHCC97-H cells. The expression of heat shock protein 72 (HSP72) at mRNA and protein levels were measured in different temperature treatment groups before and after transfection by qRT-PCR and Western blotting. Meanwhile, flow cytometry was used to detect the effect of lentiviral vector LncRNA RNU12 on the apoptosis of MHCC97-H cells induced by TRAIL protein. The fluorescence in situ hybridization (FISH) was used to detect the sub-cellular localization of LncRNA RNU12 in HCC cells. Results      Temperature at 43 ℃ promoted recombinant human TRAIL protein to induce the apoptosis in HCC cells, but it exerted opposite effect  at 46~50 ℃. The mRNA level of HSP72 was significantly decreased in all 4 treatment groups after lentiviral vector infection (P<0.01). And TRAIL protein levels were obviously lower at the 37, 46 and 50 ℃ groups than before lentivirus-mediated interference (P<0.05). After the lentiviral vector infection, the apoptosis of MHCC97-H cells induced by TRAIL protein were increased by 7.36% (P<0.01) and 8.84% (P<0.01), respectively at the 46℃ and 50℃ groups. RNA-FISH showed that LncRNA RNU12 was mainly expressed in the nucleus of MHCC97-H cells at 50℃. Conclusion      LncRNA RNU12 up-regulating HSP72 at transcriptional level may be one of the molecular mechanisms of inhibiting the apoptosis of HCC cells induced by recombinant human TRAIL protein at 46~50℃.

参考文献/References:

[1]Ferlay J, Shin H R, Bray F, et al. Estimates of worldwide burden of cancer in 2008: GLOBOCAN 2008[J]. Int J Cancer, 2010, 127(12): 2893-2917. DOI: 10.1002/ijc.25516.
[2]Forner A, Llovet J M, Bruix J. Hepatocellular carcinoma[J]. Lancet, 2012, 379(9822): 1245-1255. DOI: 10.1016/S0140-6736(11)61347-0.
[3]Nikfarjam M, Muralidharan V, Christophi C. Mechanisms of focal heat destruction of liver tumors[J]. J Surg Res, 2005, 127(2): 208-223. DOI: 10.1016/j.jss.2005.02.009.
[4]Lim B, Allen J E, Prabhu V V, et al. Targeting TRAIL in the treatment of cancer: new developments[J]. Expert Opin Ther Targets, 2015, 19(9): 1171-1185. DOI: 10.1517/14728222.2015.1049838.
[5]Yoo J, Lee Y J. Effect of hyperthermia and chemotherapeutic agents on TRAIL-induced cell death in human colon cancer cells[J]. J Cell Biochem, 2008, 103(1): 98-109. DOI: 10.1002/jcb.21389.
[6]Alcala M J, Park K, Yoo J, et al. Effect of hyperthermia in combination with TRAIL on the JNKBim signal transduction pathway and growth of xenograft tumors[J]. J Cell Biochem, 2010, 110(5): 1073-1081. DOI: 10.1002/jcb.22619.
[7]Song X, Kim H C, Kim S Y, et al. Hyperthermiaenhanced TRAIL- and mapatumumabinduced apoptotic death is mediated  through mitochondria in human colon cancer cells[J]. J Cell Biochem, 2012, 113(5): 1547-1558. DOI: 10.1002/jcb.24023.
[8]Yoo J, Lee Y J. Effect of hyperthermia on TRAIL-induced apoptotic death in human colon cancer cells: development of a novel strategy for regional therapy[J]. J Cell Biochem, 2007, 101(3): 619-630. DOI: 10.1002/jcb.21203.
[9]Wang K C, Chang H Y. Molecular mechanisms of long noncoding RNAs[J]. Mol Cell, 2011, 43(6): 904-914. DOI: 10.1016/j.molcel.2011.08.018.
[10]Evans J R, Feng F Y, Chinnaiyan A M. The bright side of dark matter: lncRNAs in cancer[J]. J Clin Invest, 2016, 126(8): 2775-2782. DOI: 10.1172/JCI84421.
[11]Jolly C, Lakhotia S C. Human sat III and Drosophila hsr omega transcripts: a common paradigm for regulation of nuclear RNA processing in stressed cells[J]. Nucleic Acids Res, 2006, 34(19): 5508-5514. DOI: 10.1093/nar/gkl711.
[12]Lanneau D, Brunet M, Frisan E, et al. Heat shock proteins: essential proteins for apoptosis regulation[J]. J Cell Mol Med, 2008, 12(3): 743-761. DOI: 10.1111/j.1582-4934. 2008.00273.x.
[13]Kennedy D, Jager R, Mosser D D, et al. Regulation of apoptosis by heat shock proteins[J]. IUBMB Life, 2014, 66(5): 327-338. DOI: 10.1002/iub.1274.
[14]Taylor R P, Benjamin I J. Small heat shock proteins: a new classification scheme in mammals[J]. J Mol Cell Cardiol, 2005, 38(3): 433-444. DOI: 10.1016/j.yjmcc.2004.12.014.
[15]Goloudina A R, Demidov O N, Garrido C. Inhibition of HSP70: a challenging anti-cancer strategy[J]. Cancer Lett, 2012, 325(2): 117-124. DOI: 10.1016/j.canlet.2012.06.003.
[16]Beere H M, Wolf B B, Cain K, et al. Heat-shock protein 70 inhibits apoptosis by preventing recruitment of procaspase-9 to the Apaf-1 apoptosome[J]. Nat Cell Biol, 2000, 2(8): 469-475. DOI: 10.1038/35019501.
[17]Zhang B, Rong R, Li H, et al. Heat shock protein 72 suppresses apoptosis by increasing the stability of X-linked inhibitor of apoptosis protein in renal ischemia/reperfusion injury[J]. Mol Med Rep, 2015, 11(3): 1793-1799. DOI: 10.3892/mmr.2014.2939.
[18]Wang X, Chen M, Zhou J, et al. HSP27, 70 and 90, anti-apoptotic proteins, in clinical cancer therapy (Review)[J]. Int J Oncol, 2014, 45(1): 18-30. DOI: 10.3892/ijo.2014.2399.

相似文献/References:

[1]赵志平,李健,王云超,等.LncRNA LOC100506123对胰腺癌细胞增殖及迁移的影响[J].第三军医大学学报,2017,39(09):840.
 Zhao Zhiping,Li Jian,Wang Yunchao,et al.Effect of LncRNA LOC100506123 on proliferation and migration in pancreatic cancer cells[J].J Third Mil Med Univ,2017,39(09):840.

更新日期/Last Update: 2017-05-05