RNAi沉默Survivin基因对膀胱癌细胞增殖和凋亡作用的体外研究
摘要
目的初步探讨用RNAi沉默survivin基因对膀胱癌T24细胞增殖和调亡的影响。
方法扩增并鉴定重组质粒;实验分为空白对照组,阴性对照组和实验组,将重组质粒(pTZU6+1-shRNA-survivin)、阴性质粒分别转染至膀胱癌T24细胞,用RT-PCR检测T24细胞survivin基因表达的变化;用MTT法观察T24细胞增殖抑制率;用AO/EB荧光染色检测和膜联蛋白V-PI(annexinV/PI)染色流式细胞技术检测T24细胞调亡的情况;用透射电镜观察T24细胞形态的改变。
结果酶切及测序鉴定结果表明:重组质粒目的片段插入正确; RT-PCR实验显示:转染pTZU6+1-shRNA-survivin 24h后,实验组T24细胞survivin基因的表达受到显著抑制,抑制率为41.77%,与阴性对照组相比有差异显著性,(P<0.05);MTT实验显示:转染48h后,实验组T24细胞增殖抑制率达到最高,为52.53%,与空白对照组及阴性对照组相比有差异显著性,(P<0.05);annexinV/PI染色法流式细胞术显示:转染24h后,实验组T24细胞凋亡率为(16.76±1.31)%,与空白对照组及阴性对照组相比有差异显著性,(P<0.05);转染24h后,实验组透射电镜观察可见各期凋亡细胞及凋亡小体,阴性对照组无明显改变。
结论转染pTZU6+1-shRNA-survivin可以显著下调survivin基因的表达,并可以显著抑制T24细胞增殖并诱导其自发凋亡。RNAi技术有可能应用于膀胱癌的治疗。
Objective To investigate the proliferation supperring and apoptosis inducing effects of RNAi targeted to Survivin on T-24 cells, a bladder cancer cell line.
Methods An siRNA eukaryotic expression vector targeted to Survivin, pTZU6+1-shRNA-survivin, was emplified in system and identitified by sequencing method. It was transfected into T24 cells following lipofectamineTM2000 protocols. The changes of transcriptional level of survivin gene were detected by semi-quantitive RT-PCR.The effects of proliferation suppressing on T24 cell were detected by MTT assay. The effects of apoptosis inducing on T-24 cells were detected by flow cytometry assay.
Rusult The recombined vector was successfully emplified, and was proved to have no mutation sites by sequencing. 24 hours after trancfection the transcriptional level of Survivin gene in T24 cells was significently suppressed at the rate of 41.77%, detected by semi-quantative RT-PCR. The proliferation suppressing effects of RNAi targeted to survivin were observed by MTT assy, 48 hours after transfecion. The proliferation inhibitory rate reached to 52.53%. Meanwhile, 24h after transfection, it was observed that silence survivin by RNAi could significently induce the spontaneous apoptosis of T-24 cells at the rate 16.76%±1.31 , detected by flow cytometry assay and comfirmed by electron microscope.
Conclusion Silencing exogenous Suvivin by RNAi can significantly suppress the proliferation of T-24 cells and induce spontaneous apoptosis. RNAi targeted to survivin has a potential role in gene therapy of bladder cancer.
方法扩增并鉴定重组质粒;实验分为空白对照组,阴性对照组和实验组,将重组质粒(pTZU6+1-shRNA-survivin)、阴性质粒分别转染至膀胱癌T24细胞,用RT-PCR检测T24细胞survivin基因表达的变化;用MTT法观察T24细胞增殖抑制率;用AO/EB荧光染色检测和膜联蛋白V-PI(annexinV/PI)染色流式细胞技术检测T24细胞调亡的情况;用透射电镜观察T24细胞形态的改变。
结果酶切及测序鉴定结果表明:重组质粒目的片段插入正确; RT-PCR实验显示:转染pTZU6+1-shRNA-survivin 24h后,实验组T24细胞survivin基因的表达受到显著抑制,抑制率为41.77%,与阴性对照组相比有差异显著性,(P<0.05);MTT实验显示:转染48h后,实验组T24细胞增殖抑制率达到最高,为52.53%,与空白对照组及阴性对照组相比有差异显著性,(P<0.05);annexinV/PI染色法流式细胞术显示:转染24h后,实验组T24细胞凋亡率为(16.76±1.31)%,与空白对照组及阴性对照组相比有差异显著性,(P<0.05);转染24h后,实验组透射电镜观察可见各期凋亡细胞及凋亡小体,阴性对照组无明显改变。
结论转染pTZU6+1-shRNA-survivin可以显著下调survivin基因的表达,并可以显著抑制T24细胞增殖并诱导其自发凋亡。RNAi技术有可能应用于膀胱癌的治疗。
Objective To investigate the proliferation supperring and apoptosis inducing effects of RNAi targeted to Survivin on T-24 cells, a bladder cancer cell line.
Methods An siRNA eukaryotic expression vector targeted to Survivin, pTZU6+1-shRNA-survivin, was emplified in system and identitified by sequencing method. It was transfected into T24 cells following lipofectamineTM2000 protocols. The changes of transcriptional level of survivin gene were detected by semi-quantitive RT-PCR.The effects of proliferation suppressing on T24 cell were detected by MTT assay. The effects of apoptosis inducing on T-24 cells were detected by flow cytometry assay.
Rusult The recombined vector was successfully emplified, and was proved to have no mutation sites by sequencing. 24 hours after trancfection the transcriptional level of Survivin gene in T24 cells was significently suppressed at the rate of 41.77%, detected by semi-quantative RT-PCR. The proliferation suppressing effects of RNAi targeted to survivin were observed by MTT assy, 48 hours after transfecion. The proliferation inhibitory rate reached to 52.53%. Meanwhile, 24h after transfection, it was observed that silence survivin by RNAi could significently induce the spontaneous apoptosis of T-24 cells at the rate 16.76%±1.31 , detected by flow cytometry assay and comfirmed by electron microscope.
Conclusion Silencing exogenous Suvivin by RNAi can significantly suppress the proliferation of T-24 cells and induce spontaneous apoptosis. RNAi targeted to survivin has a potential role in gene therapy of bladder cancer.
引文
[1] Borden LS Jr,Clark PE,Hall MC.Bladder cancer[J].Curr Opin Oncol,2003, 15 (30):227-223.
[2] 张睿,张建军等。膀胱癌相关基因的研究进展[J].癌症,2003,22(1):104-107。
[3] Ait-Si-Ait S,guasconi V,harel-Bellan A.RNA interference and its possible use in cancer therapy[J].Bull cancer, 2004;91:15-18。
[4] Ambrosini G,AItieri DC,et al.A nevel anti-apoptosisgene,Survivn,expressed in cancer and lymphoma[J].Nature MED,1997,3;917-921
[5] 曾柯,吴小候等,Survivin 与基因治疗[J].重医学报,2007,32(3):331-333
[6] 邓凯,贤钟玲,姜梅贤等。RNA 干扰技术沉默 survivin 基因逆转卵巢癌细胞株SKOV3/ADM 耐药性的研究[J].中华妇产科杂志,2005,40(12):836-839
[7] 闫歌,黄爱龙,唐霓,等.短发夹状 RNA抑制 survivin基因在肝癌细胞中的表达[J].中华肝脏病杂志,2003,11:712-715
[8] [xw00331]闫歌,RNA 干扰沉寂 survivin 表达及其对肝癌细胞凋亡和相关信号传导通路的影响[D],重庆医科大学,2004
[9] Jaattela M.Multiple cell death pathways as regulator of tumor inintiation and progression[J]. Oncogene 2004;23:2746-2756
[10]Suzuki A,Ito T,Kawano H,et al,Suevivin initiates procaspase/P21 complex formation as aresult of interration wath CDK4 to resist FAS-mediated cell death[J].ONcogene,2000,19(10);1346-1353
[11]Uren A G,Wang l.Pakusch M,et al.Survivin and the inner centromere protein INCENP show similar cell-cycle localization and gene knockout phenotype[J].Curr Biol,2002,10(21):1319
[12]Deveraux QL, Reed JC.IAP family proteins-suppressors of apoptosis[J]. Genes,1999,13(3):239-252
[13]陆才德,戴北坚,等,Survivin-值得关注的抗癌治疗靶[J].世界华人消化杂志,2005,3(15):165-166
[14]Alticri DC.Validating surviving as a cancer therapecutic target[J].Cancer,2003,3:46- 54
[15]Chang Q,Liu ZR,Wang DY,Kumar M,Chen YB,Qin RY.survivin expression in- duced by doxorubicin in cholangiocarcinoma[J].World J Gastroenterol 2004; 10:415-418
[16]高晓康,李清,王禾,等。反义 survivin 基因对 786-0 细胞的体外作用及其对表阿霉素诱导细胞凋亡作用的观察[J].中华肿瘤杂志,200,27:468-470
[17]Lu CD,Altieri DC.Tanigawa N.Expression of a novel anti-apoptosis gene, Sur- vivin,correlated with tumor cell apoptosis and P53 accumuiation in gastric carcinomas[J].Cancer Res,1998,58(9):1808
[18]13.Endoh A,Asanuma K,Moriai R,et al.Expression of surviving mRNA in CD 34-positive cells[J].Clin Chim AcTa,2001,306,149-151
[19]Swana HS,Grossman D,Anthony JN,et al.Tumor content of the antiapoplosis molecule Survivin and recurrence of bladder cancer[J].N Engl J MED,1999,341:452-453
[20][410008] ZHANG Xiang-yang, ZENG Qiang, QI Fan Department of Urology,[D],Xiangya Hospital, Central South University, 2004
[21]Monzo M,Rosell R,Felip E,et al.A novel anti-apoptosis qene:Reexpression of Survivin messenger RNA as a prognosis marker in nonsmall-cell lung cancers.J Clin Oncol.1999,17:2100-2104
[22][430030] Hou Yan, Research laboratory of pedia.tric hematology [D] Tongji medical college, 2002
[23]曹贵华,吴小侯,张尧,等。膀胱癌患者尿脱落细胞存活素表达的临床意义[J].中华泌尿外科杂志,2004,25:377-379
[24][etd-0721106-145320] 苏文娸，黃芩素抑制人类膀胱癌細胞中 survivin 表达的分子机制[D],台湾慈济大学,2006
[25]Ansell SM, Arendt BK, Grote DM. Inhibition of lymphoma [J].Leukemia 2004,18(3):616-623.
[26]Gossman D,Mc-Niff JM, Li F,Kamiya J,Altieri DC.Expression and targeting of thethe apoptosis inhibitor ,survivin,in hhuman melanoma. [J].J Invest Dermatol.1999,133(20):1076-81.
[27]Erantl S.Antisense-rna regulation and RNA interference[J].Biochim Biophys Actn 2002,1575:15-25
[28]Downward J,RNA interference.BMJ[J].2004;328:1245-1248
[29]Scherr M,Morgen MA,Eder M.Gene silencing mediated by small interfering RNAs in mammalian cells[J].Curr Med Chem 2003;10:245-246
[30]Caplen NJ,RNAi as a gene therapy approach[J].Expert Opin Biol Ther. 2003; 3:575-586.
[31]陈道荣,王丕龙,等,RNA 干扰技术的研究进展,胃肠病学和肝病学杂志[J].2005,14:104-110
[32]Sui C,Soohoo,C Affar el B,et al.A DNA vector-based RNAi technology to suppress geneexpression in mammalian cells[J].Proc Natl Acad Sci USA,2002,99(80):5515-5520.
[33]Miyagishi M,Talra K.U6 promoter driverm siRNA with four uridine 3’ overhangs efficiently suppress targeted gene erpression in mannnalian cells[J].Nature biotechnol,2002,20(5):497-500
[34]郑骏年,谢叔良,等,流式细胞术定量检测细胞凋亡 3 种方法的比较研究[J].中国免疫学杂志, 1999,10,(15):127-129
[1] Ambrosini G, AItieri DC, et al. A nevel anti-apoptosisgene, Survivn, expressed in cancer and lymphoma[J]. Nature MED,1997;3:917-921.
[2] Badran A, yoshida A, Ishikawa K, et al, Identification of anovel splice variant of the human anti-apoptopsis gene surviving[J]. Biochem Biophys Res Commum,2004;314(3):902.
[3] Uren A G, Wang l. Pakusch M, et al. Survivin and the inner centromere protein INCENP show similar cell-cycle localization and gene knockout phenotype[J]. Curr Biol,2002;10(21):1319.
[4] Deveraux QL, Reed JC. IAP family proteins- suppressors of apoptosis[J].Genes, 1999; 13 (3): 239 -252.
[5] Suzuki A, Hayashida M, Ito T, et al, Survivin initiates cell cycle entry by the complex activation with CDK4 /p16(INK4a)and CDK2, cyclinE complex activation[J]. Oncogene. 2000; 19(29): 3225-3234.
[6] Suzuki A, Ito T, Kawano H, et al, Suevivin initiates procaspase/P21 complex formation as a result of interaction with CDK4 to resist FAS-mediated cell death[J].Oncogene,2000; 19(10):1346-1353.
[7] Wheatley SP, Kandels L, Adams RR, et al. INCENP binds directly to tubulin, and requires dynamic microtubules to target to the cleavage furrow[J]. Exp, Cell Res,2001b;262:122-127.
[8] Oconnor DS, Schechner JS, Adida C, et al. Angioptosis during angiogenesis by surviving express in endothelial cell[J].Am J Pathol,2000;156(6):393.
[9] Papapetropoulos A, Fulton D, Mubboubi K, et al. Angiopioetin-1 inhibits endothelial cell[J]. J Biol Chem, 2000;275(13):9102.
[10]Alticri DC. Validating surviving as a cancer therapeutic target[J]. Nat.Rev. Cancer,2003;3:46-54.
[11]O'Connor D S, Schechner J S, Adida C. Control of apoptosis during angiogenesis by Survivin expression in endothelial cells[J].Am J Pathol,2000; 156(2): 393-398.
[12]Lu CD, Altieri DC. Tanigawa N. Expression of a novel anti-apoptosis gene, Survivin, correlated with tumor cell apoptosis and P53 accumulation in gastric carcinomas[J]. Cancer Res,1998;58(9):1808.
[13]Endoh A, Asanuma K, Moriai R, et al. Expression of surviving mRNA in CD34-positive cells[J]. Clin Chim AcTa,2001;306:149-151.
[14] Kawasaki H, Altieri DC, LuCD, et al. Unhibition of apoptosis by Survivin predicts Shorter Survivin rates in colorectal cancer[J].Cancer Res,1998;56(22):5071.
[15]Swana HS, Grossman D, Anthony JN, et al. Tumor content of the anti-apoplosis molecule Survivin and recurrence of bladder cancer[J]. N Engl J MED,1999;341:452-453.
[16] ZHANG Xiang-yang, ZENG Qiang, QI Fan,et al. Expression of Survivin protein in prostate cancer and its clinical sign ificance[J]. China Journl Of Modern Medicine, 2006;16:1060-1062.
[17]Mono M, Rosell R, Felip E, et al. A novel anti-apoptosis gene Rexpression of Survivin messenger RNA as a prognosis marker in nonsmall-cell lung cancers[J]. J Clin Oncol. 1999; 17: 2100-2104.
[18]Hou Yan, Hu Qun, Liu Aiguo, Zhang Liuqing, et al. Expression of survivin and its location in cell and clinical significance in pediatric acute leukemia[J]. Journal Of China PediaTric Blood And Cancer, 2006; 1:6-8.
[19]Smith SD, Wheeler MA, Plescia J, et al. Urine detection of Survivin and diagnosis of bladder cancer[J]. JAMA, 2001;17; 285(3):324-8.
[20]Ansell SM, Arendt BK, Grote DM. Inhibition of surviving expression suppresses the growth of aggressive non-hodgkin’s lymphoma[J].Leukemia,2004; 18(3):616-623.
[21]Fonaro M, Plescia J, Chheng S, et al. Fibronectin protects prostate cancer cells from tumor mecrosisi factor-alpha induced apoptosisi via the AKT/surviving pathway[J]. J boil chem, 2003;278(50):50402-50411.
[22]Fire A, Xu S, Montgomery MK, et al. Potent and specific genetic interference bydouble-stranded RNA in Caenorhabditis elegans[J].Nature,1998:391(6669)806-811
[23]Kappler M, Bache M, Bartel F, et al. Knockdown of survivin expression by small interfering RNA reduces the clonogenic surviving of human sarcoma cell lines independentlyof p53[J]. Cancer Gene Ther, 2004; 11(3): 186-193.
[24]Marzia P, Mara B, Micheiandrea DC. Ribozyme-mediated down-regulation of surviving expression sensitizes human melanoma cells to potecan in vitro and in[J]. Carcinogenesis, 2004;10(5):1093.
[25]Jiangguo W, Xiang L, Dalin P, et al. Molecular Mechanism of inhibition of surviving transcription by the GC-rich sequence-selective DNA binding anti-tumor agent, Hedamycin[J]. J Boil Chem,2005; 280(10):116-122
[2] 张睿,张建军等。膀胱癌相关基因的研究进展[J].癌症,2003,22(1):104-107。
[3] Ait-Si-Ait S,guasconi V,harel-Bellan A.RNA interference and its possible use in cancer therapy[J].Bull cancer, 2004;91:15-18。
[4] Ambrosini G,AItieri DC,et al.A nevel anti-apoptosisgene,Survivn,expressed in cancer and lymphoma[J].Nature MED,1997,3;917-921
[5] 曾柯,吴小候等,Survivin 与基因治疗[J].重医学报,2007,32(3):331-333
[6] 邓凯,贤钟玲,姜梅贤等。RNA 干扰技术沉默 survivin 基因逆转卵巢癌细胞株SKOV3/ADM 耐药性的研究[J].中华妇产科杂志,2005,40(12):836-839
[7] 闫歌,黄爱龙,唐霓,等.短发夹状 RNA抑制 survivin基因在肝癌细胞中的表达[J].中华肝脏病杂志,2003,11:712-715
[8] [xw00331]闫歌,RNA 干扰沉寂 survivin 表达及其对肝癌细胞凋亡和相关信号传导通路的影响[D],重庆医科大学,2004
[9] Jaattela M.Multiple cell death pathways as regulator of tumor inintiation and progression[J]. Oncogene 2004;23:2746-2756
[10]Suzuki A,Ito T,Kawano H,et al,Suevivin initiates procaspase/P21 complex formation as aresult of interration wath CDK4 to resist FAS-mediated cell death[J].ONcogene,2000,19(10);1346-1353
[11]Uren A G,Wang l.Pakusch M,et al.Survivin and the inner centromere protein INCENP show similar cell-cycle localization and gene knockout phenotype[J].Curr Biol,2002,10(21):1319
[12]Deveraux QL, Reed JC.IAP family proteins-suppressors of apoptosis[J]. Genes,1999,13(3):239-252
[13]陆才德,戴北坚,等,Survivin-值得关注的抗癌治疗靶[J].世界华人消化杂志,2005,3(15):165-166
[14]Alticri DC.Validating surviving as a cancer therapecutic target[J].Cancer,2003,3:46- 54
[15]Chang Q,Liu ZR,Wang DY,Kumar M,Chen YB,Qin RY.survivin expression in- duced by doxorubicin in cholangiocarcinoma[J].World J Gastroenterol 2004; 10:415-418
[16]高晓康,李清,王禾,等。反义 survivin 基因对 786-0 细胞的体外作用及其对表阿霉素诱导细胞凋亡作用的观察[J].中华肿瘤杂志,200,27:468-470
[17]Lu CD,Altieri DC.Tanigawa N.Expression of a novel anti-apoptosis gene, Sur- vivin,correlated with tumor cell apoptosis and P53 accumuiation in gastric carcinomas[J].Cancer Res,1998,58(9):1808
[18]13.Endoh A,Asanuma K,Moriai R,et al.Expression of surviving mRNA in CD 34-positive cells[J].Clin Chim AcTa,2001,306,149-151
[19]Swana HS,Grossman D,Anthony JN,et al.Tumor content of the antiapoplosis molecule Survivin and recurrence of bladder cancer[J].N Engl J MED,1999,341:452-453
[20][410008] ZHANG Xiang-yang, ZENG Qiang, QI Fan Department of Urology,[D],Xiangya Hospital, Central South University, 2004
[21]Monzo M,Rosell R,Felip E,et al.A novel anti-apoptosis qene:Reexpression of Survivin messenger RNA as a prognosis marker in nonsmall-cell lung cancers.J Clin Oncol.1999,17:2100-2104
[22][430030] Hou Yan, Research laboratory of pedia.tric hematology [D] Tongji medical college, 2002
[23]曹贵华,吴小侯,张尧,等。膀胱癌患者尿脱落细胞存活素表达的临床意义[J].中华泌尿外科杂志,2004,25:377-379
[24][etd-0721106-145320] 苏文娸，黃芩素抑制人类膀胱癌細胞中 survivin 表达的分子机制[D],台湾慈济大学,2006
[25]Ansell SM, Arendt BK, Grote DM. Inhibition of lymphoma [J].Leukemia 2004,18(3):616-623.
[26]Gossman D,Mc-Niff JM, Li F,Kamiya J,Altieri DC.Expression and targeting of thethe apoptosis inhibitor ,survivin,in hhuman melanoma. [J].J Invest Dermatol.1999,133(20):1076-81.
[27]Erantl S.Antisense-rna regulation and RNA interference[J].Biochim Biophys Actn 2002,1575:15-25
[28]Downward J,RNA interference.BMJ[J].2004;328:1245-1248
[29]Scherr M,Morgen MA,Eder M.Gene silencing mediated by small interfering RNAs in mammalian cells[J].Curr Med Chem 2003;10:245-246
[30]Caplen NJ,RNAi as a gene therapy approach[J].Expert Opin Biol Ther. 2003; 3:575-586.
[31]陈道荣,王丕龙,等,RNA 干扰技术的研究进展,胃肠病学和肝病学杂志[J].2005,14:104-110
[32]Sui C,Soohoo,C Affar el B,et al.A DNA vector-based RNAi technology to suppress geneexpression in mammalian cells[J].Proc Natl Acad Sci USA,2002,99(80):5515-5520.
[33]Miyagishi M,Talra K.U6 promoter driverm siRNA with four uridine 3’ overhangs efficiently suppress targeted gene erpression in mannnalian cells[J].Nature biotechnol,2002,20(5):497-500
[34]郑骏年,谢叔良,等,流式细胞术定量检测细胞凋亡 3 种方法的比较研究[J].中国免疫学杂志, 1999,10,(15):127-129
[1] Ambrosini G, AItieri DC, et al. A nevel anti-apoptosisgene, Survivn, expressed in cancer and lymphoma[J]. Nature MED,1997;3:917-921.
[2] Badran A, yoshida A, Ishikawa K, et al, Identification of anovel splice variant of the human anti-apoptopsis gene surviving[J]. Biochem Biophys Res Commum,2004;314(3):902.
[3] Uren A G, Wang l. Pakusch M, et al. Survivin and the inner centromere protein INCENP show similar cell-cycle localization and gene knockout phenotype[J]. Curr Biol,2002;10(21):1319.
[4] Deveraux QL, Reed JC. IAP family proteins- suppressors of apoptosis[J].Genes, 1999; 13 (3): 239 -252.
[5] Suzuki A, Hayashida M, Ito T, et al, Survivin initiates cell cycle entry by the complex activation with CDK4 /p16(INK4a)and CDK2, cyclinE complex activation[J]. Oncogene. 2000; 19(29): 3225-3234.
[6] Suzuki A, Ito T, Kawano H, et al, Suevivin initiates procaspase/P21 complex formation as a result of interaction with CDK4 to resist FAS-mediated cell death[J].Oncogene,2000; 19(10):1346-1353.
[7] Wheatley SP, Kandels L, Adams RR, et al. INCENP binds directly to tubulin, and requires dynamic microtubules to target to the cleavage furrow[J]. Exp, Cell Res,2001b;262:122-127.
[8] Oconnor DS, Schechner JS, Adida C, et al. Angioptosis during angiogenesis by surviving express in endothelial cell[J].Am J Pathol,2000;156(6):393.
[9] Papapetropoulos A, Fulton D, Mubboubi K, et al. Angiopioetin-1 inhibits endothelial cell[J]. J Biol Chem, 2000;275(13):9102.
[10]Alticri DC. Validating surviving as a cancer therapeutic target[J]. Nat.Rev. Cancer,2003;3:46-54.
[11]O'Connor D S, Schechner J S, Adida C. Control of apoptosis during angiogenesis by Survivin expression in endothelial cells[J].Am J Pathol,2000; 156(2): 393-398.
[12]Lu CD, Altieri DC. Tanigawa N. Expression of a novel anti-apoptosis gene, Survivin, correlated with tumor cell apoptosis and P53 accumulation in gastric carcinomas[J]. Cancer Res,1998;58(9):1808.
[13]Endoh A, Asanuma K, Moriai R, et al. Expression of surviving mRNA in CD34-positive cells[J]. Clin Chim AcTa,2001;306:149-151.
[14] Kawasaki H, Altieri DC, LuCD, et al. Unhibition of apoptosis by Survivin predicts Shorter Survivin rates in colorectal cancer[J].Cancer Res,1998;56(22):5071.
[15]Swana HS, Grossman D, Anthony JN, et al. Tumor content of the anti-apoplosis molecule Survivin and recurrence of bladder cancer[J]. N Engl J MED,1999;341:452-453.
[16] ZHANG Xiang-yang, ZENG Qiang, QI Fan,et al. Expression of Survivin protein in prostate cancer and its clinical sign ificance[J]. China Journl Of Modern Medicine, 2006;16:1060-1062.
[17]Mono M, Rosell R, Felip E, et al. A novel anti-apoptosis gene Rexpression of Survivin messenger RNA as a prognosis marker in nonsmall-cell lung cancers[J]. J Clin Oncol. 1999; 17: 2100-2104.
[18]Hou Yan, Hu Qun, Liu Aiguo, Zhang Liuqing, et al. Expression of survivin and its location in cell and clinical significance in pediatric acute leukemia[J]. Journal Of China PediaTric Blood And Cancer, 2006; 1:6-8.
[19]Smith SD, Wheeler MA, Plescia J, et al. Urine detection of Survivin and diagnosis of bladder cancer[J]. JAMA, 2001;17; 285(3):324-8.
[20]Ansell SM, Arendt BK, Grote DM. Inhibition of surviving expression suppresses the growth of aggressive non-hodgkin’s lymphoma[J].Leukemia,2004; 18(3):616-623.
[21]Fonaro M, Plescia J, Chheng S, et al. Fibronectin protects prostate cancer cells from tumor mecrosisi factor-alpha induced apoptosisi via the AKT/surviving pathway[J]. J boil chem, 2003;278(50):50402-50411.
[22]Fire A, Xu S, Montgomery MK, et al. Potent and specific genetic interference bydouble-stranded RNA in Caenorhabditis elegans[J].Nature,1998:391(6669)806-811
[23]Kappler M, Bache M, Bartel F, et al. Knockdown of survivin expression by small interfering RNA reduces the clonogenic surviving of human sarcoma cell lines independentlyof p53[J]. Cancer Gene Ther, 2004; 11(3): 186-193.
[24]Marzia P, Mara B, Micheiandrea DC. Ribozyme-mediated down-regulation of surviving expression sensitizes human melanoma cells to potecan in vitro and in[J]. Carcinogenesis, 2004;10(5):1093.
[25]Jiangguo W, Xiang L, Dalin P, et al. Molecular Mechanism of inhibition of surviving transcription by the GC-rich sequence-selective DNA binding anti-tumor agent, Hedamycin[J]. J Boil Chem,2005; 280(10):116-122