下调SIRT1表达对子宫内膜癌细胞增殖及凋亡的影响
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摘要
目的:研究下调沉默信息调节因子1(SIRT1)表达对子宫内膜癌细胞增殖及凋亡的影响。方法:子宫内膜癌细胞Ishikawa分为3组,空白对照组不处理,另2组分别感染SIRT1 shRNA重组慢病毒(干扰组)和阴性对照shRNA慢病毒(阴性对照组),用qRT-PCR和Western blot检测干扰效果,用MTT法测定细胞增殖,PI单染法检测细胞周期分布,Annexin V-FITC/PI法检测细胞凋亡,Western blot检测细胞中活化Caspase-3(Cleaved Caspase-3)、CCAAT/增强子结合蛋白同源蛋白(CHOP)、活化转录因子6(ATF6)、细胞周期蛋白D1(Cyclin D1)和P21蛋白表达水平。结果:与阴性对照组和空白对照组比较,干扰组细胞SIRT1 mRNA和蛋白表达水平、细胞增殖能力降低,G0/G1期细胞比例升高,细胞凋亡率升高,Cyclin D1蛋白表达水平降低,P21蛋白表达水平升高,Cleaved Caspase-3、CHOP、ATF6蛋白表达水平也升高(P <0. 05)。结论:下调SIRT1可能通过阻滞细胞周期进展抑制子宫内膜癌细胞增殖,同时通过激活内质网应激诱导细胞凋亡。
Aim: To study the effects of down-regulating the expression of SIRT1 on the proliferation and apoptosis of endometrial cancer cells. Methods: The endometrial cancer Ishikawa cells were allocated into 3 groups,the blank control group was not treated,and the other 2 groups were infected with SIRT1 shRNA recombinant lentivirus( interference group)and the negative control shRNA lentivirus( negative control group). The expression of SIRT1 was detected by qRT-PCR and Western blot. Cell proliferation,apoptosis and changes of cycle distribution were determined by MTT,Annexin V-FITC/PI and PI single staining,respectively. The expressions of Cleaved Caspase-3,CCAAT/enhancer-binding protein homologous protein( CHOP),activating transcription factor 6( ATF6),cell cycle protein D1( Cyclin D1) and P21 were detected by Western blot. Results: Compared with the 2 control groups,the levels of SIRT1 mRNA and protein were significantly decreased in Ishikawa cells infected with SIRT1 shRNA recombinant lentivirus,the proliferation capacity of Ishikawa cells and cyclin D1 protein levels were decreased,G0/G1 cell ratio and cell apoptosis rate were increased,the expressions of Cleaved Caspase-3,CHOP,ATF6 and P21 were also increased( P < 0. 05). Conclusion: Down-regulation of SIRT1 could inhibit the proliferation of endometrial cancer cells by blocking cell cycle progression and induce cell apoptosis by activating endoplasmic reticulum stress.
Aim: To study the effects of down-regulating the expression of SIRT1 on the proliferation and apoptosis of endometrial cancer cells. Methods: The endometrial cancer Ishikawa cells were allocated into 3 groups,the blank control group was not treated,and the other 2 groups were infected with SIRT1 shRNA recombinant lentivirus( interference group)and the negative control shRNA lentivirus( negative control group). The expression of SIRT1 was detected by qRT-PCR and Western blot. Cell proliferation,apoptosis and changes of cycle distribution were determined by MTT,Annexin V-FITC/PI and PI single staining,respectively. The expressions of Cleaved Caspase-3,CCAAT/enhancer-binding protein homologous protein( CHOP),activating transcription factor 6( ATF6),cell cycle protein D1( Cyclin D1) and P21 were detected by Western blot. Results: Compared with the 2 control groups,the levels of SIRT1 mRNA and protein were significantly decreased in Ishikawa cells infected with SIRT1 shRNA recombinant lentivirus,the proliferation capacity of Ishikawa cells and cyclin D1 protein levels were decreased,G0/G1 cell ratio and cell apoptosis rate were increased,the expressions of Cleaved Caspase-3,CHOP,ATF6 and P21 were also increased( P < 0. 05). Conclusion: Down-regulation of SIRT1 could inhibit the proliferation of endometrial cancer cells by blocking cell cycle progression and induce cell apoptosis by activating endoplasmic reticulum stress.
引文
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[17]LUO J,ZHANG C,WANG C,et al. Miz-1 promotes the proliferation of esophageal cancer cells via suppression of p21and release of p21-arrested cyclin D1[J]. Oncol Rep,2016,35(6):3532
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[19]SUN L,LI H,CHEN J,et al. PIASy mediates hypoxia-induced SIRT1 transcriptional repression and epithelial-tomesenchymal transition in ovarian cancer cells[J]. J Cell Sci,2013,126(Pt 17):3939
[20]KANG KA,PIAO MJ,RYU YS,et al. Luteolin induces apoptotic cell death via antioxidant activity in human colon cancer cells[J]. Int J Oncol,2017,51(4):1169
[21]MA J,YANG YR,CHEN W,et al. Fluoxetine synergizes with temozolomide to induce the CHOP-dependent endoplasmic reticulum stress-related apoptosis pathway in glioma cells[J]. Oncol Rep,2016,36(2):676
[22]TIAN X,SHI Y,LIU N,et al. Upregulation of DAPK contributes to homocysteine-induced endothelial apoptosis via the modulation of Bcl2/Bax and activation of caspase 3[J]. Mol Med Rep,2016,14(5):4173
[23]吴倩,刘新伟. SIRT1激活ERK1/2通路抑制大鼠心肌缺血再灌注时内质网应激相关蛋白的表达[J].基础医学与临床,2015,35(6):761
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[2] WANG Y,BI Y,CHEN X,et al. Histone deacetylase SIRT1negatively regulates the differentiation of interleukin-9-producing CD4(+)T cells[J]. Immunity,2016,44(6):1337
[3] LAGOUGE M,ARGMANN C,GERHART-HINES Z,et al.Resveratrol improves mitochondrial function and protects against metabolic disease by activating SIRT1 and PGC-1 alpha[J]. Cell,2006,127(6):1109
[4] CHEN IC,CHIANG WF,HUANG HH,et al. Role of SIRT1in regulation of epithelial-to-mesenchymal transition in oral squamous cell carcinoma metastasis[J]. Mol Cancer,2014,13:254
[5] HIDA Y,KUBO Y,MURAO K,et al. Strong expression of a longevity-related protein,SIRT1,in Bowen's disease[J].Arch Dermatol Res,2007,299(2):103
[6]郭宏英,张云良,李志红,等.子宫内膜癌中SIRT1和Survivin的表达及意义[J].临床与实验病理学杂志,2012,28(11):1224
[7] JEONG H,COHEN DE,CUI LB,et al. Sirt1 mediates neuroprotection from mutant huntingtin by activation of the TORC1 and CREB transcriptional pathway[J]. Nat Med,2011,18(1):159
[8] OTA H,TOKUNAGA E,CHANG K,et al. Sirt1 inhibitor,Sirtinol,induces senescence-like growth arrest with attenuated Ras-MAPK signaling in human cancer cells[J]. Oncogene,2006,25(2):176
[9] HONG EH,LEE SJ,KIM JS,et al. Ionizing radiation induces cellular senescence of articular chondrocytes via negative regulation of SIRT1 by p38 kinase[J]. J Biol Chem,2010,285(2):1283
[10]JANG KY,HWANG SH,KWON KS,et al. SIRT1 expression is associated with poor prognosis of diffuse large B-cell lymphoma[J]. Am J Surg Pathol,2008,32(10):1523
[11]HUFFMAN DM,GRIZZLE WE,BAMMAN MM,et al.SIRT1 is significantly elevated in mouse and human prostate cancer[J]. Cancer Res,2007,67(14):6612
[12]HU Q,WANG G,PENG J,et al. Knockdown of SIRT1 suppresses bladder cancer cell proliferation and migration and induces cell cycle arrest and antioxidant response through FOXO3a-mediated pathways[J]. Biomed Res Int,2017,2017:3781904
[13]吴琦,于红,张文卿. siRNA沉默SIRT1基因对宫颈癌细胞He La增殖和凋亡的影响[J].微生物学杂志,2014,34(3):47
[14]魏琳琳,王蕊,关淑梅.子宫内膜癌中ILK、SIRT1和PTEN的表达及意义[J].中国妇幼保健,2014,29(29):4747
[15]ZURYNA,LITWINIEC A,SAFIEJKO-MROCZKA B,et al.The effect of sulforaphane on the cell cycle,apoptosis and expression of cyclin D1 and p21 in the A549 non-small cell lung cancer cell line[J]. Int J Oncol,2016,48(6):2521
[16]ZERJATKE T,GAK IA,KIROVA D,et al. Quantitative cell cycle analysis based on an endogenous all-in-one reporter for cell tracking and classification[J]. Cell Rep,2017,19(9):1953
[17]LUO J,ZHANG C,WANG C,et al. Miz-1 promotes the proliferation of esophageal cancer cells via suppression of p21and release of p21-arrested cyclin D1[J]. Oncol Rep,2016,35(6):3532
[18]李驰,王忠利. RNA干扰沉默信息调节因子2同源蛋白1阻滞前列腺癌PC3细胞周期[J].中国肿瘤临床,2014,41(20):1274
[19]SUN L,LI H,CHEN J,et al. PIASy mediates hypoxia-induced SIRT1 transcriptional repression and epithelial-tomesenchymal transition in ovarian cancer cells[J]. J Cell Sci,2013,126(Pt 17):3939
[20]KANG KA,PIAO MJ,RYU YS,et al. Luteolin induces apoptotic cell death via antioxidant activity in human colon cancer cells[J]. Int J Oncol,2017,51(4):1169
[21]MA J,YANG YR,CHEN W,et al. Fluoxetine synergizes with temozolomide to induce the CHOP-dependent endoplasmic reticulum stress-related apoptosis pathway in glioma cells[J]. Oncol Rep,2016,36(2):676
[22]TIAN X,SHI Y,LIU N,et al. Upregulation of DAPK contributes to homocysteine-induced endothelial apoptosis via the modulation of Bcl2/Bax and activation of caspase 3[J]. Mol Med Rep,2016,14(5):4173
[23]吴倩,刘新伟. SIRT1激活ERK1/2通路抑制大鼠心肌缺血再灌注时内质网应激相关蛋白的表达[J].基础医学与临床,2015,35(6):761
[24]张开阳.硫化氢上调SIRT1拮抗甲醛诱导PC12细胞内质网应激[D].衡阳:南华大学,2013.