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荧光内参在校正Caspase-3／7活性检测偏差中的应用 被引量：1

Application of Internal Control Reniila Luciferase in Unbiased Estimation of Caspase-3/7 Activity Assay for Apoptosis
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摘要 目的改进Caspase-3／7活性检测方法。方法在顺铂诱导的HeLa细胞凋亡模型中，分别利用传统的和改良的实验方法检测Caspase-3／7的活性变化，并与Western印迹实验结果进行方法学比较。结果改良的实验方法显示不同剂量顺铂诱导下，HeLa细胞Caspase-3／7活性有剂量依赖性增高，与Western印迹实验结果相一致，但传统实验方法显示HeLa细胞Caspase-3／7活性呈现先增高后降低的趋势。结论由于参测细胞数不同，导致这种Caspase-3／7活性检测方法不能真实反映细胞凋亡程度。本研究成功建立了一种新的改良型Caspase-3／7活性检测方式，这种检测方法可以排除不同凋亡诱导方式诱导细胞凋亡时所产生的因参测细胞数不同所造成的Caspase-3／7活性检测的误差。 Objective To improve the accuracy of Caspase-3/7 activity assay. Methods In cisplatin induced apoptotic Hela cell model, Caspase-3/7 activity of Hela cells was determined by traditional method or modified method. Meanwhile, cleaved PARP was detected by Western blot to determine apoptosis induction. Results The results of modified assay showed that Caspase-3/7 activity was increased in a cisplatin dose dependent manner, which was consistent with the Western blot result. However, the results of traditional assay showed that Caspase-3/7 activity of Hela cells was firstly increased but then declined, displaying a bias due to decreased cell numbers in the high dose of eisplatin. Conclusion Because the cell numbers involved in the assay were different, the traditional Caspase-3/7 activity assay is not reliable to reflect the apoptotic extent of tested samples. Our newly established Caspase-3/7 activity assay with internal control may correct the deviations of Caspase-3/7 activity assay caused by difference in cell numbers among samples, thus providing more accurate and reliable results.
作者 晋亮 杨国栋 傅海燕 卢晓昭 韦梦影 于芳 卢兹凡
机构地区 国家肿瘤生物学重点实验室第四军医大学基础部生物化学与分子生物学教研室
出处 《医学分子生物学杂志》 CAS CSCD 2010年第1期6-10,共5页 Journal of Medical Molecular Biology
基金 国家杰出青年科学基金B类（No.30628025）
关键词 凋亡 Caspase-3/7 内参 顺铂 PARP apoptosis caspase-3/7 internal control cisplatin PARP
分类号 Q291 [生物学—细胞生物学]
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参考文献11

  • 1KUROKAWA M,KORNBLUTH S. Caspases and kinases in a death grip [ J ]. Cell,2009,138 ( 5 ) : 838-854.
  • 2O' BRIEN M A, DAILY W J, HESSELBERTH P E, et al. Homogeneous, bioluminescent protease assays: caspase-3 as a model [J]. J Biomol Screen,2005,10(2) :137-148.
  • 3MARTHA O' BRIEN P D P C. Technically Speaking-Cell-Based Caspa-se Assays: Analyzing the Data [ J ]. Promega Notes, 2004, 87:33-36.
  • 4YANG G, FU H,ZHANG J, et al. RNA-binding protein quaking, a critical regulator of colon epithelial differentiation and a suppressor of colon cancer[ J]. Gastroenterology,2010,138( 1 ) :231-240.
  • 5CALL J A,ECKHARDT S G,CAMIDGE D R. Targeted manipulation of apoptosis in cancer treatment [ J ]. Lancet Oncol, 2008,9 (10) :1002-1011.
  • 6POP C, SALVESEN G S. Human caspases : activation, specificity, and regulation[J]. J Biol Chem,2009,284(33):21777-21781.
  • 7DE BRUIN E C, MEDEMA J P. Apoptosis and non-apoptotic deaths in cancer development and treatment response [ J ]. Cancer Treat Rev,2008,34 ( 8 ) :737-749.
  • 8KERR J F, WYLLIE A H, CURRIE A R. Apoptosis : a basic biological phenomenon with wide-ranging implications in tissue kinetics [J]. Br J C 1972,26(4) :239-257.
  • 9COTTER T G. Apoptosis and cancer:the genesis of a research field [ J ]. Nat Rev Cancer, 2009,9 ( 7 ) :501-507.
  • 10徐智芳,王建祥.Caspase 8与肿瘤的发生[J].医学分子生物学杂志,2007,4(4):331-334. 被引量：3

二级参考文献11

  • 1[1]BIENKO M,GREEN C M,CROSETTO N,et al.Ubiquitin-binding domains in Y-family polymerases regulate tranalesion synthesis[J].Science.2005,310(5755):1821-1824.
  • 2[2]MAGA G,HUBSCHER U.Proliferating cell nuclear antigen (PC-NA):a dancer with many partners[J].J Cell Sci,2003,116 (Pt15):3051-3060.
  • 3[3]WATANABE K,TATEISHI S,KAWASUJI M,et al.Rad18 guides poleta to replication stalling sites through physical interaction and PCNA monoubiquitination[J].EMBO J,2004,23(19):3886-3896.
  • 4[4]TORRES-RAMOS C A,PRAKASH S,PRAKASH L.Requirement of RAD5 and MMS2 for postreplication repair of UV-damaged DNA in Saccharomyces cerevisiae[J].Mol Cell Biol,2002,22 (7):2419-2426.
  • 5[5]KRISHNA T S,KONG X P,GARY S,et al.Crystal stracture of the eukaryotic DNA polymerase proeessivity factor PCNA[J].Cell,1994,79(7):1233-1243.
  • 6[6]HARACSKA L,TORRES-RAMOS C A,JOHNSON R E,et al.Opposing effects of ubiquitin onjugation and SUMO modification of PCNA on replicational bypass of DNA lesions in saccharomyces cerevisiae[J].Mol Cell Biol,2004,24(10):4267-4274.
  • 7[7]HOEGE C,PFANDER B,MOLDOVAN G L,et al.RAD6-dependent DNA repair is linked to modification of PCNA by ubiquitin and SUMO[J].Nature,2002,419(6903):135-141.
  • 8[8]DEMPKE W,VOIGT W,GROTHEY A,et al.Cisplatin resistance and oncogenes[J].Anticancer Drugs,2000,11(4):225-236.
  • 9J. F. de Vries,L. J. Wammes,I. Jedema,L. van Dreunen,B. A. Nijmeijer,M. H. M. Heemskerk,R. Willemze,J. H. F. Falkenburg,R. M. Y. Barge. Involvement of caspase-8 in chemotherapy-induced apoptosis of patient derived leukemia cell lines independent of the death receptor pathway and downstream from mitochondria[J] 2007,Apoptosis(1):181～193
  • 10S. Cagnol,E. Obberghen-Schilling,J.-C. Chambard. Prolonged activation of ERK1,2 induces FADD-independent caspase 8 activation and cell death[J] 2006,Apoptosis(3):337～346

共引文献3

同被引文献20

  • 1HICKS C W,ROME E S. Chronic pelvic: pain in the ado- lescent[ J ]. Endocr Dev,2012,22 : 230-250.
  • 2BERKLEY K J. Sex differences in pain [ J ]. Behav Brain Sci, 1997,20 ( 3 ) : 371-380.
  • 3LEE O Y, MAYER E A, SCHMULSON M, et al. Gender- related differences in IBS symptoms [ J ]. Am J Gastroen- terol,2001,96(7 ) :2184-2193.
  • 4BERKI,EY K J. A life of pelvic pain [ J ]. Physiol Behav, 2005,86(3 ) :272-280.
  • 5BRINKERT W, DIMCEVSKI G, ARENDT-NIELSEN L, et al. Dysmenorrhoea is associated with hypersensitivily in the sigmoid colon and rectum [ J ]. Pain, 2007,132 ( Suppl 1 ) :46-51.
  • 6CHABAN V V. Visceral sensory neurons that innervate both uterus and colon express nocieeptive TRPvl and P2X3 re- ceptors in rats[J]. Ethn Dis,2008,18: 20-24.
  • 7LI J, MICEVYCH P, MCDONALD J, et al. Inflammation in the uterus induces phosphorylated extracellular signal- regulated kinase and substance P immunoreactivity in dor- sal root ganglia neurons innervating both uterus and colon in rats[J]. J Neurosci Res,2008,86(12) :2746-2752.
  • 8DREW I, J,MACDERMOTT A B. Neuroscience: Unbear- able lightness of touch [ J ] . Nature, 2009,462 ( 7273 ) : 580-581.
  • 9SEAL R P, AKIL O, YI E, et al. Sensorineural deafness and seizures in mice lacking vesicular glutamate transport- er 3 [ J ]. Neuron, 2008,57 ( 2 ) : 263-275.
  • 10DANIEl.S R W, GELFAND M V, COLLINS C A, et al. Visualizing glutamatergic cell bodies and synapses in Drosophila larval and adult CNS [ J ]. J Comp Neurol, 2008,508( 1 ) :131-152.

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医学分子生物学杂志
2010年 第1期

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