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THE EFFECT OF Antisense OLIGONUCLEOTIDE ON THE INTERLEUKIN-5 IN THE SUPERNATANTS OF SPLEEN CELL CULTURES OF ASTHMATIC MICE

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China Medical Abstracts(Internal Medicine) 2001年第3期18卷 183-188页

作者：王美琴白春学钮善福方晓惠陈常庆陈波 Institute of Respiratory Disease Zhongshan Hospital Fudan University Shanghai 200032 China Department of Respiratory Medicine Huashan Hospital Fudan University Shanghai 200040 China Institute of Respiratory Disease Zhongshan Hospital Fudan University Shanghai 200032 China Central Biological Engineering of Shanghai Affiliated to China Academy of Sciences Shanghai 200233 China

To explore the effect of Antisense oligonucleotide on the production of IL-5 by mouse spleen T lymphocytes.Methods Based on the IL-5 cDNA sequence of mouse, a segment of Antisense oligonucleotide was designed and synthesized. 5’-labeling of Antisense oligonucleotide was signed by T4 PNK in order that the efficiency of stearylamine liposome in transfecting Antisense oligonucleotide can be evaluated. Asthma model was duplicated with ovalbumin（OVA） absorbed to aluminum hydroxide. T lymphocytes of mice were separated by nylon fiber method, then T lymphocytes transfected with different concentration of Antisense oligonucleotide with cation stearylamine liposme were incubated respectively in order to observe the effect of Antisense oligonucleotide on Il-5 production by T lymphocytes. IL-5 levels in the supernatants of T lymphocyte cultures were determined by ELISA.Results Stearylamine liposome could markedly increase the efficiency of Antisense oligonucleotide transfection. The transfection efficiency of Antisense oligouncleotide increased approximately 12 times at a ratio of 1: 15m/m （Antisense oligonucleotide to SA liposome）. In healthy and asthma Balb/c mice, IL-5 was not detectable in the supernatants of T lymphocyte cultures without stimulated with OVA; however, IL-5 was increased markedly in the supernatants of T lymphocyte cultures stimulated with OVA. After transfection with different concentrations of Antisense oligonucleotide, IL-5 levels in the supernatants of T lymphocyte cultures were significantly lower than those in control cultured without Antisense oligonucleotide transfection. IL-5 levels decreased from 44.60±6.23 pg/ml to 30.70±7.362 pg/ml, 17.20±6.181 pg/ml and 8.16±2.34 pg/ml respectively. And IL-5 synthesis was inhibited by 31.17%, 61.43% and 81.7% respectively.Conclusion IL-5 synthesis could be obviously inhibited by Antisense oligonucleotide and showed a markedly correlation between dose and effectiveness. It suggests the production of IL-5 is inhibited by preventing the transcription

关键词： Antisense asthma lymphocyte transfection ovalbumin inhibited liposome markedly stimulated detectable

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Distribution of c-erbB2 Antisense Probe Labeled with Superparamagnetic Iron Oxide Nanoparticles in the Major Organs of Mice on MR Imaging

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Advances in Nanoparticles 2013年第1期2卷 16-20页

作者： Zhipeng Wen Haiyan Liu Hairong He Shude Tan Ming Wen Shaolin Li Department of Radiology The First Affiliated Hospital Chongqing Medical University Chongqing China

Background: The aim of this study was to investigate the distribution of the c-erbB2 Antisense probe labeled with superparamagnetic iron oxide nanoparticles in the major organs of mice by MR imaging. Methods: Sixty BALB/c mice were randomly divided into experimental and control groups. MR scans were performed in each mouse of the experimental group at five different time points (10, 30, 60, 180 and 360 min) after injection of the Antisense probe. The signal from each major organ (liver, spleen, heart, kidney and muscle tissue) in comparison with the background signal (signal to noise ratio) was determined at each time point as a measure of the distribution of the Antisense probe. Six control mice were killed at each of the same time points and the organs immediately removed for determination of their iron content. Results: After injection of the Antisense probe, the highest enrichment of the probe was seen in the spleen, reaching a peak at 180 min, followed by the liver, muscle, heart and kidney. Conclusions: MR imaging can visualize the distribution of c-erbB2 Antisense probe labeled with superparamagnetic iron oxide nanoparticles in the major organs of mice, and this may provide the basis for further in vivo studies of MR imaging time and dose selection.

关键词： Antisense Oligodeoxynucleotide Superparamagnetic Iron Oxide Mr Imaging Iron Concentration Vivo Distribution

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Construction of Antisense hTR gene in retroviral vector

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中国实验血液学杂志 1997年第3期5卷 332-333页

作者： Zheng-Sheng Li, Guan-Hua Lai, Zheng-GangZhu, Hao-Ran Yin, Yan-Zhen Lin, Shi-ShuChenRuijin Affiliated Hospital and Research Center forHuman Gene Therapy, Shanghai Second MedicalUniversity, Shanghai 200025

Human chromosomes terminate with severalkilobases of the simple telomere repeat (TTAGGG) n.Telomeres protect the chromosomes from DNAdegradation, end-to-end fusion, rearrangements andchromosome losses. Since DNA polymerasessynthesize DNA in the 5’ to 3’ direction and require aRNA primer for initiation, telomeric DNA could belost at chromosome ends unless the termini arespecifically extended by telomerase. Telomerase is aspecialized ribonucleoprotein polymerase that containsan integral RNA with a short template element

关键词： chromosomes telomerase unless termini Antisense telomere template RNA primer DNA

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Inhibition of HIV-1 replication by intracellular expression of Antisense polymeric TAR-Core RNAs

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中国实验血液学杂志 1997年第3期5卷 319-320页

作者： Longchuan Bai, Jiangang Yuan, Quanbi Zhao, Yiming Shao, Boqin QiangNational Lalxmtory of Medical Molecular Biology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100005 Laboratory of AIDS Research and Reference, Chinese Academy of Preventive Medicine, Beijing 100051

Objectives: To construct retroviral vectorsexpressing Antisense polymeric RNAs targeted at the coresequence (+13-+47) of HIV-l TAR RNA, and to testthe anti-HIV properties of this construct in transducedCD4^+T cells. M... 详细信息

关键词： Antisense polymeric replication intracellular template targeted RNA Inhibition inhibited inserted

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Effect of Antisense human telomerase RNA on malignant behaviors of gastric carcinoma cell line SGC-7901

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Journal of Medical Colleges of PLA(China) 2001年第4期16卷 255-259页

作者：杨金亮房殿春杨仕明罗元辉罗昆仑鲁荣刘为纹 Department of Gastroenterology Southwest Hospital Third Military Medical University Chongqing 400038 China

Objective:To studytheeffectsof AntisensehumantelomeraseRNA(ahTR)transfectionon themalignantbeha-viorsof gastriccarcinomacelllineSGC-7901anditspotentialroleingenetherapyfortumor.Methods:AnAntisensehTR eukaryoticexpressionvectorcontainingthesequenceof templateregionof telomererepeatswas transfectedintogastric carcinomacelllineSGC-7901withliposomeDOTAP.Theexpressionsof hTRRNAandAntisensehTRRNAwereob-servedwithRT-PCR,telomeraseactivitywithPCR-ELISA.Telomerelengthwas measuredwithSouthernblot.Cellmor-phologyandcellularproliferationcapacitywerestudiedwithMTTassay.Cellcycledistributionandapoptoticstatewere observedwithflowcytometry.Efficiencyof cloneformationin softagarandtumorigencityin nudemicewereexamined andevaluatedinahTR-transfected7901cells,andplasmidpCL-neotransfected7901cellsandparental7901cellsserved as control.Results:AnAntisensehTReukaryoticexpressionvectorwastransfectedinto7901cellssuccessfully.Thetelom-eraseactivityin ahTR-transfected7901cellswas decreasedfrom100%to about25%,andtelomerelengthin thecells shortenedfrom4.08kb to3.35kb at60populationdoublings(PDs).Comparedwithparental7901andpCL-neotransfect-ed7901cells,ahTR-transfected7901cellsdisplayedsomemorphologicalchanges,includingdecreasedcellatypiaandnu-cleus/cytoplasmratiounderlightmicroscope.Furthermore,ahTR-transfected7901cellsdisplayedgrowthinhibition,de-creasedinvasivecapacityin Borden’schamberinvasivemodel,increasedG 0 /G 1 phaserateandapoptoticrate,andrestored contactinhibitionanddensityinhibition.Surprisingly,ahTR-transfected7901cellslosttheircapacityof cloneformationin softagarandcarcinogensisinnudemice.Conclusion:AntisensehTRtransfectioncaninduce7901celldifferentiationand reverseitsmalignantphenotype.Thisstudyprovidesan excitingapproachfor cancertherapythroughtheinhibitionof telomeraseactivitywithAntisensegeneandothertelomeraseinhibitors.

关键词： human telomerase RNA components Antisense gene telomerase eukaryotic expression vector, gene therapy gastric carcinoma cell line

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AntisenseGAD_(67)ODN对戊四氮所致慢性癫癇大鼠的作用

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中国临床神经科学 2001年第1期9卷 1-5页

作者：王伟何小华阮旭中张梁王玉同济医科大学附属同济医院神经内科 430030

目的 :探讨谷氨酸脱羧酶 (GAD)的抗癫疒间作用。方法 :采用戊四氮制备大鼠慢性癫疒间模型 ,利用反义寡脱氧核苷酸技术 ,通过选择性地抑制海马GAD基因的表达来观察其对慢性癫疒间大鼠行为、发作阈值及脑电图的影响。结果 :给予Antisense... 详细信息

目的 :探讨谷氨酸脱羧酶 (GAD)的抗癫疒间作用。方法 :采用戊四氮制备大鼠慢性癫疒间模型 ,利用反义寡脱氧核苷酸技术 ,通过选择性地抑制海马GAD基因的表达来观察其对慢性癫疒间大鼠行为、发作阈值及脑电图的影响。结果 :给予AntisenseGAD67ODN处理后 ,GADmRNA表达下降、GABA浓度下降、疒间性发作潜伏期缩短、发作程度加重及脑电图疒间波频率增加。结论 :从反面进一步说明GAD基因可能是抗疒间基因。

关键词：癫痫谷氨酸脱羧酶 GAD 寡脱氧核苷酸戊四氮 Antisense GAD67ODN

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Inhibition of metastasis to lung of a human nasopharyngeal carcinoma cell line CNE-2L2 transfected with pRc/CMV-Antisense 6A8 cDNA in nude mice

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Science China(Life Sciences) 1999年第2期42卷 209-215页

作者：张立新刘玉琴马凤蓉顾蓓史耕先赵雪梅李波高进赵方萄张淑珍李国燕王讯朱立平

The growth of CNE-2L2 cell, a cloned line of human nasopharyngeal carcinoma with a high potentiality of metastasis to lung was inhibited to a certain extent after transfection with a recombinant Antisense expression vector of a cDNA encoding a human α-mannosidase (pRc/CMV-Antisense 6A8 cDNA)( the Genbank accession number of 6A8 cDNA is U37248) in comparison with that of the cell transfected with the Mock and of the wild cell. Two months after a subcutaneous inoculation of CNE-2L2 cell into the axilla of nude mice metastatic lesions in the lung were observed in 9/10 mice (90%) with grade Ⅲ in 8 mice and grade Ⅱ in one mouse in the wild cell group, in 6/8 mice (75%) with grade Ⅲ in one mouse, grade Ⅱ in 2 mice and grade Ⅰ in 3 mice in the Mock-transfection group, in only 3/10 mice (30%) with all grade Ⅰ in pRc/CMV-Antisense 6A8 cDNA-transfection group.

关键词： Antisense 6A8 cDNA transfection metastasis nasopharyngeal carcinoma cell line α-mannosidase.

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Inhibitory effects of Antisense oligodeoxynucleotide on replication and expression of HDV genome in vitro and in vivo

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Journal of Medical Colleges of PLA(China) 2001年第4期16卷 239-243页

作者：毛青李奇芬李宏文王升启郑红丁健吴纯清朱宝珍 Center of Infectious Diseases Southwest Hospital Third Military Medial University Chongqing 400038 China Institute of Beijing Radiation A cademy of Military Medici Beijing 100850 China

Objective:To studytheinhibitoryeffectsof Antisenseoligodeoxynucleotide(ASODN)anditsthiophosphate(S-ASODN)on thereplicationandexpressionof hepatitisD virus(HDV)in H1δ9cellsandin tupaiabody.Methods:After15mer-ASODNandS-ASODNweresynthesized,differentconcentrationsof ASODNandS-ASODNwereaddedto theculturemediumof H1δ9celllineandthenHDAgin thesupernatantof theculturewas examinedwithELISAand HDV-RNAinthecellsdeterminedwithdotblothybridization.SixteentupaiaeweresuccessfullyinfectedwithHDVand thenequallyrandomizedinto2groups.In thetreatedgroup,theanimalsreceivedintravenousinjectionsof S-ASODN3mg everyotherdayfor7timesandthecontrolgroupwereinjected withsamevolumeof normalsaline.On the5th,10th,15thand20thdayaftertheadministration,samplesof bloodandlivertissueswereexaminedwithimmunohistochemical methodforHDAganddotblothybridizationandin situ hybridizationforHDV-RNA.Results:Twenty-fourhoursafter theadditionof a finalconcentrationof6μmol/Lof S-ASODN,thereplicationof HDV-RNAandthereleaseof HDAgin theH1δ9cellsweresuppressedby84.5%and76.14%respectively.Theinhibitionwas dose-dependentwhenthefinal concentrationsof2,4and6μmol/Lof S-ASODNweregiven.WhenASODNandS-ASODNwereadministeredinthe samedosage,theinhibitionshowedno significantlydifferencebetweenthe2agents.On thelastdayof theadministration of S-ASODN,7outof the8tupaiaeof thetreatedgroupshowednegativeHDAgandHDV-RNAinthelivertissuewhile only1outof the8tupaiaeof thecontrolgroupwasnegative.Tendaysafterthecessationof drugadministration,3tupaiae of thetreatedgroupand7of controlwerepositiveof HDAgandHDV-RNA.Conclusion:OurfindingsshowthatS-ASODNefficientlyinhibitesthereplicationandexpressionof HDVgeneinH1δ9cellsandin thebodyof tupaia,which providesanexperimentalbasisfortheanti-HDVapplicationof Antisenseoligonucleotides.

关键词： Antisense oligodeoxynucleotide thiophosphoate hepatic D virus H1δ9 cells tupaiae

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Effects of CIITA Antisense RNA on the expression of HLA class Ⅱ molecules

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Chinese Science Bulletin 2000年第22期45卷 2068-2071页

作者： ZHOU Caihong, LU Daru, ZHU Qiquan, QIU Xinfang & XUE JinglunInstitute of Genetics, Fudan University, Shanghai 200433, China Institute of Genetics Fudan University Shanghai China

To study the effect of the major histocompatibility complex class Ⅱ (MHC Ⅱ ) transactivator (CIITA) Antisense RNA on the expression of the human leukemia (HLA) class Ⅱ molecules, 5’ end cDNA sequence of CIITA gene was cloned, and Antisense RNA expression vector pcDNA-Ⅱ was constructed. HeLa cells transfected with pcDNA-Ⅱ and pcDNA3 were induced by IFN-γ for 3 d. The expression of HLA class Ⅱ molecules on HeLa/pcDNA-Ⅱ cells was significantly decreased, while it has no effect on the expression of HLA class Ⅰ molecules. This result suggests that the CIITA Antisense RNA can inhibit the expression of HLA class Ⅱ molecules in HeLa cells. It also implies a promising approach to generate immune tolerance in qraft transplantation.

关键词： CIITA Antisense RNA HLA class Ⅱ molecules.

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Altered Oncogene Activity Contributes to Compensation for Antisense Suppression of Bcl-2 and Tumor Resistance

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Open Journal of Apoptosis 2015年第3期4卷 62-70页

作者： Marvin Rubenstein Courtney M. P. Hollowell Patrick Guinan Division of Cellular Biology Hektoen Institute for Medical Research Chicago IL USA Division of Urology Stroger Hospital of Cook County Chicago IL USA Department of Biochemistry Rush University Medical Center Chicago IL USA Department of Urology Rush University Medical Center Chicago IL USA Department of Urology University of Illinois at Chicago Chicago IL IL USA

Antisense oligonucleotides (oligos) have targeted growth regulatory proteins in prostate cancer models. To identify compensatory alterations in the expression of non-targeted genes we evaluate mono- and bispecific oligos targeting and equally suppressing the expression of the apoptosis inhibitory protein bcl-2. Bcl-2 is chosen because oligos directed towards it have entered clinical trials to restore apoptosis in cancer patients. Treated LNCaP cells compensate for the diminished bcl-2 by suppressing caspase-3 (an apoptosis promoter) while enhancing expression of AKT-1 (another apoptosis inhibitor), androgen receptor (AR) and its (p300 and IL-6) coactivators. Additional proteins are enhanced including PD-1, its ligand PD-L1 (immune checkpoint blockade markers) and fas-ligand, which activate apoptosis through the signal transduction, along with suppressor protein p53, polymerase transcription mediator MED-12 and signal transducer STAT-3. These alterations in expression may contribute to a greatly enhanced expression of the proliferation marker KI-67. This suggests that therapeutic approaches to restore apoptosis through suppression of bcl-2 lead to an altered expression in non-targeted genes involving apoptosis, androgen sensitivity, transcriptional activity and immune responsiveness, leads to an increase in proliferation (and a more androgen driven aggressive phenotype). In this study we evaluate the expression of two oncogenes (v-myc and K-ras) and find a large and significant enhancement of v-myc activity, which is produced by oligos targeting bcl-2 at the 5’ position. For K-ras, although significant suppression is produced by the bispecific targeting bcl-2 at the 3’ position, the percent change is relatively small compared with other compensatory alterations we have measured, and much less than in v-myc. Therefore, for the two oncogenes being evaluated, only increased v-myc activity is probably large enough to contribute to increased tumor aggressiveness in compensation for bcl-2 suppression.

关键词： Antisense Oligonucleotides Prostate Cancer Bcl-2 Gene Compensation Therapy

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