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Long-term stable expression of antisense cDNA of cyclin b1 profoundly inhibits the proliferation of tumor cells and suppresses tumorigenicity in implanted mice

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Chinese Medical Journal 2008年第15期121卷 1433-1438页

作者： ZHANG Tao ZHANG Ling LI Ji-cheng WEI Dong WEI Yu-quan ZHANG Ru CHENG Peng CHEN Xian-cheng LIU Huan-yi SU Xiao-mei Department of Oncology General Hospital of Chengdu Military Command of PLA Chengdu Sichuan 610083 China Institute of Cell biology Medical College of Zhejiang University Hangzhou Zhejiang 310058 China State Key Laboratory of biotherapy and Cancer Center West China Hospital West China Medical School Sichuan University Chengdu Sichuan 610041 Chin

background cyclin b1 （CLb1） is necessary for mitotic initiation in mammalian cells and plays important roles in cancer development. Therefore, a potential strategy in cancer therapy is to suppress the activity of CLb1 by delivering antisense constructs of CLb1 into tumor cells. In previous CLb1 studies, antisense constructs with a short half life were often used and these constructs might not persistently inhibit CLb1. Methods We successfully created a recombinant plasmid encoding the full-length antisense cDNA of mouse cyclin b1 （AS-mCLb1） and transfected this construct to the murine Lewis lung carcinoma （LL/2） and CT-26 colon carcinoma （CT-26） cells. We isolated clones of LL/2 and CT-26 transfectants with stable expression of AS-mCLb1. Reverse transcriptional polymerase chain reaction （RT-PCR） and Western blot were applied to detect the expression of the mRNA and protein levels of CLb1. To further test the efficacy of this strategy in vivo, AS-mCLbl-expressing LL/2 and CT-26 transfectants were implanted into mice. Results We found the expression of the mRNA and protein levels of CLb1 decrease in these transfectants. The inhibition of CLb1 caused prominent G1 arrest, abnormal morphology, retarded cell growth and an increase in apoptosis. In AS-mCLbl-expressing LL/2 and CT-26 transfectants implanted mice, tumorigenicity was effectively suppressed compared with the controls. In addition, the expression of AS-mCLb1 also significantly increases the survival duration of implanted animals. Conclusion AS-mCLb1 is likely to be useful in future cancer therapy, which may be associated with its ability to down-regulate the expression of CLb1 and then induce G1 arrest and apoptosis in tumor cells.

关键词： tumorigenicity cyclin b1 cancer therapy antisense cDNA

The expression and significance of cyclin b1 and survivin in human non-small cell lung cancer

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The Chinese-German Journal of Clinical Oncology 2011年第4期10卷 192-197页

作者： Guosheng Li Xuhan Liu Daorong Zhang Dong Liu Zhiyong Li Department of Pathology The First Affiliated Hospital of Dalian Medical University Dalian 116011 China Department of Endocrinology Dalian Municipal Central Hospital Dalian 116003 China Department of Pathology College of Basic Medical Sciences China Medical University Shenyang 110001 China Department of Vathology Tumor Hospital （Institute） of Liaoning Province Shenyang 110042 China Department of Radiology The First Affiliated Hospital of DalianMedical University.Dalian 116011 China

Objective:We studied the expression of cyclin b1 and survivin in human non-small cell lung cancer(NSCLC),and the relationship between such expression and clinicopathological features of ***:One hundred cases of tissue specimen including NSCLC,neighboring noncancerous tissue and normal lung tissue were collected at *** specimens were detected by immunohistochemical ***:The expression of cyclin b1 and survivin showed significant difference(P between NSCLC tissues,proliferating epithelial cells of bronchioles and small bronchi in neighboring noncancerous tissues,and normal lung *** with normal lung tissues,there was an overexpression of cyclin b1 and survivin in NSCLC and an enhancing expression of cyclin b1 and survivin in proliferating epithelial cells of bronchioles and small bronchi in neighboring noncancerous *** positive correlation was found between the overexpression of cyclin b1 and that of survivin in 100 NSCLC cases(P between the enhancing expression of cyclin b1 and that of survivin in proliferating epithelial cells of bronchioles and small bronchi in neighboring noncancerous tissues(P between the different histological types,the differentiated degree,lymphatic metastasis and the expression of cyclin b1 and survivin(P > 0.05) in *** significance was marked between different clinical stages of NSCLC and the expression of cyclin b1 and survivin(P cyclin b1 and survivin was found in *** expression of cyclin b1 and survivin might be up-regulated during an early step of tumorigenesis and during the development of *** progression of cell cycle could be efficiently connected with the control of apoptosis by the interrelations between the overexpression of cyclin b1 and that of survivin in NSCLC during the G2/M *** ove

关键词： non-small cell lung cancer cell cycle regulation apoptosis cyclin b1 survivin

cyclin b1对头颈部鳞状细胞癌生物学特性及细胞焦亡的影响

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中国医科大学学报 2022年第3期51卷 209-215页

作者：罗水妹张琦黄谊强谢贤和福建医科大学附属第一医院肿瘤内科福建医科大学分子肿瘤研究所福州350005 福建省癌症精准医学重点实验室福州350005

目的通过大数据挖掘与实验相结合,探讨cyclin b1表达对头颈部鳞状细胞癌(HNSC)生物学特性及细胞焦亡的影响。方法从GEO、UALCAN及Human Protein Atlas数据库分析cyclin b1在HNSC组织及正常组织的表达及其与临床特征及预后的关系;通过GEO... 详细信息

目的通过大数据挖掘与实验相结合,探讨cyclin b1表达对头颈部鳞状细胞癌(HNSC)生物学特性及细胞焦亡的影响。方法从GEO、UALCAN及Human Protein Atlas数据库分析cyclin b1在HNSC组织及正常组织的表达及其与临床特征及预后的关系;通过GEO、OncoLnc及GEPAI数据库筛选差异表达及与cyclin b1表达相关的细胞焦亡基因。通过转录组测序、蛋白印迹、ELISA等方法探讨cyclin b1对细胞焦亡的影响。结果UALCAN及GEO数据库分析提示cyclin b1表达在HNSC组织中显著上调(P1),与不良组织分级、分期及无病生存期(DFS)相关(Pcyclin b1表达水平相关(P1细胞株为对象的研究显示,与对照组相比,沉默cyclin b1表达可显著增加caspase 1的mRNA表达(P1β释放(P>0.05)。蛋白印迹实验结果提示2组焦亡通路关键蛋白caspase 1、caspase 3、GSDMD、GSDME的裂解片段未见显著差异。结论cyclin b1在HNSC组织中呈显著高表达,与HNSC的不良分级、分期及DFS显著相关。cyclin b1表达可能与细胞焦亡生物学过程相关,其分子机制有待进一步研究。

关键词：细胞焦亡头颈部鳞状细胞癌 cyclin b1

The significance of cyclin b1 expression in colorectal cancers

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The Chinese-German Journal of Clinical Oncology 2006年第6期5卷 399-401页

作者： Haocheng Long Xia Gao Xiaoyan Chen Xiaolan Li Xiaojun He Deding Tao Jianping Gong The Cancer Research Institute / Molecular Medical Center Tongji Hospital Tongji Medical College Huazhong University of Science and Technology Wuhan 430030 China

Objective: To study the relationship between the expression of human cyclin b1 in colorectal carcinomas and the pathological characters. Methods: The Expression of cyclin b1 in 66 cases of colorectal carcinomas were detected by flow cytometry and immunohistochemistry. Then the relationship between the expression of cyclin b1 in colorectal carcinomas and pathological characters was analyzed with statistics. Results: The expression of cyclin b1 in colorectal carcinomas had associa- tivity with the cancer cell differentiation (Pcyclin b1 in colorectal carcinomas had no obvious associativity with cancer cell infiltrate depth and lymph nodes metastasis (P>0.05). Conclusion: In the colorectal cancers with high expression of cyclin b1, the cancer cells would present high differentiation; with low expression of cyclin b1 the cancer cells would present low differentiation. Along with the expression of cyclin b1 from high to low, the cancer cells differentiation has the tendency from high to low too.

关键词： cyclin b1 colorectal cancers immunohistochemistry flow cytometry cell differentiation

cyclin b1、Cdc2和Cdc25C在生门上型喉鳞状细胞癌中的表达及意义

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Cyclin B1、Cdc2和Cdc25C在生门上型喉鳞状细胞癌中的表达及意义

作者：张颉河北医科大学

学位级别：硕士

目的：喉癌（Carcinoma of larynx）是耳鼻喉--头颈部常见的恶性肿瘤，以鳞状细胞癌最为多见。喉癌分为声门上型、声门型、声门下型及跨声门型，其中声门上型和声门型占绝大多数，在我国部分地区声门上型喉癌常见，而声门上型喉癌有易... 详细信息

目的：喉癌（Carcinoma of larynx）是耳鼻喉--头颈部常见的恶性肿瘤，以鳞状细胞癌最为多见。喉癌分为声门上型、声门型、声门下型及跨声门型，其中声门上型和声门型占绝大多数，在我国部分地区声门上型喉癌常见，而声门上型喉癌有易转移、预后较声门型喉癌差的特点。研究喉癌发生发展机制，尤其对喉癌的基础研究，对于临床有效防治具有重要的意义。如癌基因研究有助于筛选可靠的靶向治疗靶控基因。一般认为，细胞周期调控紊乱在肿瘤的发生发生发展过程中发挥重要作用。细胞周期调控机制复杂，其中G1/S、G2/M两个关键限制点在细胞周期调控中发挥重要作用，G1/S期的主要调控因子有cyclin D、cyclin E、CDK4、P16、P21、P27等，G2/M期的主要调控因子有cyclin b、Cdc2、CDK2、Cdc25C等。细胞经过S期后，DNA复制完成，细胞进入G2期，为细胞分裂做准备。M期为细胞分裂期，在M期，细胞一分为二，遗传物质也随之平均分配到两个子代细胞中，完成整个细胞周期。cyclinb1/Cdc2复合物是G2/M检查点的监督机制，是有丝分裂中最关键的因子。Cdc25C主要的作用靶点是cyclin b1/Cdc2复合物。Cdc25C发生抑制性磷酸化后其磷酸酶活性丧失，使得cyclin b1/Cdc2复合物处于磷酸化的失活状态，阻止细胞G2/M期转换，使细胞周期进程停止。研究表明，cyclinb1、Cdc2及Cdc25C在多种肿瘤组织中的表达明显增高，与肿瘤的发生发展密切相关，但尚未见上述因子在声门上型喉癌中的表达情况及其临床病理意义的研究报道。本研究拟采用免疫组织化学方法观察声门上型喉鳞状细胞癌组织中cyclin b1、 Cdc2及Cdc25C的表达，探讨cyclin b1、Cdc2及Cdc25C在声门上型喉癌发生发展中的作用及相关性，为喉癌的治疗和评估预后提供新的方向。方法：采用免疫组织化学SP法，选择声门上型喉鳞状细胞癌组织96例，正常喉粘膜组织15例作为对照。检测癌组织及正常对照组中cyclinb1、Cdc2及Cdc25C的表达情况。结果：1cyclin b1的表达cyclin b1的阳性染色部位定位于细胞质，以胞质内出现棕黄色或棕褐色颗粒定义为阳性细胞。cyclin b1在声门上型鳞状细胞癌中的阳性表达率（51.0%）明显高于正常喉粘膜组织（13.3%，Pcyclin b1在高、中、低分化癌组织组中的阳性表达率分别为11.1%、51.4%和76.9%，cyclin b1的表达随喉癌分化程度降低而升高（Pcyclin b1在伴有淋巴结转移的癌组织中阳性表达率明显高于不伴有淋巴结转移的癌组织（73.2%vs34.5%，Pcyclin b1表达与临床分期密切相关，临床分期越晚，cyclin b1的表达越强，Ⅲ、Ⅳ期喉癌组的阳性率（59.2%）明显高于I、Ⅱ期喉癌组（28.0%，Pcyclin b1在癌组织中的表达与病人年龄、性别、吸烟量和肿瘤大小无明显相关性（P>0.05）。2Cdc2的表达Cdc的阳性染色部位定位于细胞核与细胞质，以胞核或胞质内出现棕黄色及棕褐色颗粒为阳性细胞。Cdc2在声门上型鳞状细胞癌的阳性表达率明显高于正常喉粘膜组织（61.5%vs6.7%），差异有统计学意义（P0.05）。Cdc2在癌组织中的表达与年龄、性别、吸烟量、肿瘤大小无明显相关性（P>0.05）。3Cdc25C的表达Cdc25C的阳性染色部位为细胞核，以细胞核内出现棕黄色颗粒为阳性细胞。Cdc25C在声门上型鳞状细胞癌及正常喉粘膜组织中的阳性表达率分别为81.3%和13.3%，声门上型鳞状细胞癌组中明显高于正常喉粘膜组，差异有统计学意义（P0.05）。4声门上型喉鳞状细胞癌中cyclin b1、 Cdc2及Cdc25C表达量之间的关系cyclin b1与Cdc2的表达存在正相关关系（r=0

关键词：声门上型喉鳞状细胞癌 cyclin b1 Cdc2 Cdc25C 细胞周期

cyclin b1与TGF-β1在非小细胞肺癌中的表达及意义

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Cyclin B1与TGF-β1在非小细胞肺癌中的表达及意义

作者：张楠福建医科大学

学位级别：硕士

目的：研究细胞周期蛋白b1（cyclin b1）与转化生长因子β1（TGF-β1）在非小细胞肺癌（NSCLC）组织中的表达及其与组织学类型、分期、淋巴结转移及预后的关系，并分析两者的关系。方法：采用免疫组化SP法检测cyclin b1与TGF-β1在62例N... 详细信息

目的：研究细胞周期蛋白b1（cyclin b1）与转化生长因子β1（TGF-β1）在非小细胞肺癌（NSCLC）组织中的表达及其与组织学类型、分期、淋巴结转移及预后的关系，并分析两者的关系。方法：采用免疫组化SP法检测cyclin b1与TGF-β1在62例NSCLC及30例癌旁正常肺组织中表达。对所有NSCLC患者进行术后随访，了解术后存活时间，随访时间截止至2010年6月。结合临床病理和随访资料，运用统计学方法，回顾性分析cyclin b1和TGF-β1在NSCLC和正常肺组织表达的差异;cyclin b1与TGF-β1的表达和各项临床病理指标及预后之间的关系。计数资料采用χ2检验，相关性运用Spearman分析，生存率计算使用Kaplan-Meier法，并绘制累积生存曲线，用Log-rank检验各组间差异;COX法多因素分析筛选NSCLC预后的独立危险因素。所有数据的统计分析应用PASW Statistics18.0统计软件进行，所有统计结果均以Pb1与TGF-β1在NSCLC组织中的表达阳性率分别为46.77％（29/62）和56.45％（35/62），在正常肺组织中的表达阳性率分别为16.67％（5/30）和10％（3/30）。*** b1的表达与年龄、性别、组织类型、淋巴结转移无关（P>0.05），而与肿瘤大小、分化程度、TNM分期有关（P0.05），与淋巴结转移、TNM分期有关（Pcyclin b1表达和TGF-β1表达相关（Pcyclin b1与TGF-β1的联合表达也是影响NSCLC预后的独立危险因素（Pb1与TGF-β1在NSCLC组织中的表达具有正相关性（Pcyclin b1在NSCLC组织中过表达，且与肿瘤大小、分化程度、TNM分期有关。2. TGF-β1在NSCLC组织中过表达，且与淋巴结转移、TNM分期有关。3. cyclin b1与TGF-β1在NSCLC组织中的表达具有正相关性。4.分化程度、淋巴结转移、TNM分期和TGF-β1的表达是影响NSCLC预后的独立危险因素， cyclin b1的过表达和/或TGF-β1的过表达提示预后不良，两者联合应用意义更大。*** b1与TGF-β1在NSCLC的发生发展中具有协同作用，促进NSCLC的发生、发展及侵袭、转移，可作为判断预后的生物学指标。

关键词： cyclin b1 TGF-β1 免疫组化非小细胞肺癌预后

Alkaloids-rich fraction of Crotalaria sessiliflora induces cell growth arrest and apoptosis by up-regulating cyclin b1 signaling in pancreatic cancer cells

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Alkaloids-rich fraction of Crotalaria sessiliflora induces c...

Alkaloids-rich fraction of Crotalaria sessiliflora induces cell growth arrest and apoptosis by up-regulating cyclin b1 signaling in pancreatic cancer cells

湖南省生理科学会2013年度学术年会

作者：曾军英李胜华伍贤进民族药用植物资源研究与利用湖南重点实验室湘西药用植物与民族植物学湖南省高校重点实验室怀化学院生命科学系

ETHNOPHARMACOLOGICAL RELEVANCE:A medicinal herb Crotalaria Sessiliflora has been used to treat various diseases including diuretic,cardiotonic,analgesic,and anticancer in folk traditional medicine in Southern of *** aim of the present study was to investigate the anticancer activity of Crotalaria Sessiliflora in pancreatic cancer *** AND METHODS:The cytotoxic and apoptotic effect of alkaloids-rich fraction of Crotalaria sessiliflora（ACS） on human pancreatic cancer cells 13.6pl and bxpc-3 cells were evaluated using MTT and TUNEL assays,and Annexin V/Propidium iodide detection by flow *** effect of ACS on cell cycle was analyzed by flow cytometry with propidium *** apoptotic and cell cycle effects were also evaluated by RT-PCR and western blot ***:Alkaloids-rich fraction of Crotalaria sessiliflora repressed clonogenecity and proliferation,induced apoptosis,and enhanced accumulation in the G2 phase of pancreatic cancer *** addition,a second population of cells expressing cyclin bl had developed polyploidy and DNA content consistent with 8 N *** with adriamycin,a known inducer of p53,ACS-treated pancreatic cancer cells expressed p53 only at high concentrations and p53 expression was not coordinated with G2 arrest or ***,ACS suppressed the transcription of genes associated with cell cycle regulation,anti-apoptosis,and ***:These data showed that ACS blocks proliferation and induces apoptosis in human pancreatic cancer cells through G2 arrest in association with persistent cyclin bl expression,failure to completely activate cdc2,and continued DNA synthesis through a pathway that is unrelated to altered expression of *** can be used as an anticancer drug for pancreatic carcinoma patients.

关键词： Alkaloids Crotalaria sessiliflora pancreatic cancer cell growth arrest apoptosis cyclin b1

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Alkaloids-rich fraction of Crotalaria sessiliflora induces c...

白桦脂酸对人AGS胃腺癌细胞Hiwi和cyclin b1基因表达的调控作用

生态文明建设中的植物学：现在与未来——中国植物学会第十五届会员代表大会暨八十周年学术年会

作者： Jun-ying Zeng Sheng-hua Li Xian-jin WU The Department of Life Science Huaihua University Key Laboratory of Hunan Province for Study and Utilization of Ethnic Medicinal Plant Resources Key Laboratory of Hunan Higher Education for Hunan-Western Medicinal Plant and Ethnobotany

ETHNOPHARMACOLOGICAL RELEVANCE:A medicinal herb Crotalaria Sessiliflora has been used to treat various diseases including diuretic,cardiotonic,analgesic,and anticancer in folk traditional medicine in Southern of *** aim of the present study was to investigate the anticancer activity of Crotalaria Sessiliflora in pancreatic cancer *** AND METHODS:The cytotoxic and apoptotic effect of alkaloids-rich fraction of Crotalaria sessiliflora(ACS) on human pancreatic cancer cells l3.6pl and bxpc-3 cells were evaluated using MTT and TUNEL assays,and Annexin V/Propidium iodide detection by flow *** effect of ACS on cell cycle was analyzed by flow cytometry with propidium *** apoptotic and cell cycle effects were also evaluated by RT-PCR and western blot ***:Alkaloids-rich fraction of Crotalaria sessiliflora repressed clonogenecity and proliferation,induced apoptosis,and enhanced accumulation in the G2 phase of pancreatic cancer *** addition,a second population of cells expressing cyclin b1 had developed polyploidy and DNA content consistent with 8 N *** with adriamycin,a known inducer of p53,ACS-treated pancreatic cancer cells expressed p53 only at high concentrations and p53 expression was not coordinated with G2 arrest or ***,ACS suppressed the transcription of genes associated with cell cycle regulation,anti-apoptosis,and ***:These data showed that ACS blocks proliferation and induces apoptosis in human pancreatic cancer cells through G2 arrest in association with persistent cyclin b1 expression,failure to completely activate cdc2,and continued DNA synthesis through a pathway that is unrelated to altered expression of *** can be used as an anticancer drug for pancreatic carcinoma patients.

关键词： Alkaloids Crotalaria sessiliflora pancreatic cancer cell growth arrest apoptosis cyclin b1

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白桦脂酸对人AGS胃腺癌细胞Hiwi和Cyclin B1基因表达的调控作用

作者：杨立靖华中科技大学

学位级别：硕士

目的:本实验以人胃腺癌AGS细胞为研究对象,观察白桦脂酸对AGS细胞增殖、凋亡及周期的影响,并探讨其分子机制。方法:用噻唑蓝（MTT）法检测来白桦脂酸对AGS细胞增殖的影响,用AnnexinV-PI双标流式细胞术来检测AGS细胞凋亡率,用Hoechst... 详细信息

目的:本实验以人胃腺癌AGS细胞为研究对象,观察白桦脂酸对AGS细胞增殖、凋亡及周期的影响,并探讨其分子机制。方法:用噻唑蓝（MTT）法检测来白桦脂酸对AGS细胞增殖的影响,用AnnexinV-PI双标流式细胞术来检测AGS细胞凋亡率,用Hoechst 33258染色分析AGS细胞发生凋亡后的形态学变化。经白桦脂酸处理后的AGS细胞,其细胞周期的变化经PI染色后用流式细胞仪来检测。Hiwi和cyclin b1蛋白的变化用单标流式细胞术检测,而其mRNA表达水平的相对高低用逆转录PCR（RT-PCR）来检测。结果:在体外实验中,白桦脂酸能以时间和剂量依赖方式显著抑制AGS细胞的增殖,其24 h的IC50值为18.25 ug/mL （95%可信区间:15.16-27.31 ug/mL）。当白桦脂酸的浓度达到12 ug/mL以上时,AGS细胞出现明显的凋亡,经Hoechst 33258染色可见典型的凋亡小体。白桦脂酸还能使AGS细胞周期阻滞在G2/M期,而处在S期的细胞数量减少。经白桦脂酸处理24 h的AGS细胞,其Hiwi和cyclin b1蛋白及m RNA的表达水平均以剂量依赖的方式下调。结论:Hiwi基因在AGS细胞中高水平异常表达。白桦脂酸在体外实验中,具有抑制AGS细胞增殖,诱导其凋亡并阻滞其于G2/M期的活性作用。而这种作用可能与白桦脂酸能在基因水平上抑制Hiwi及cyclin b1的表达有关。白桦脂酸强大的抗肿瘤细胞活性使其可能成为一种新的抗胃腺癌肿瘤药物。

关键词：白桦脂酸 AGS细胞 Hiwi cyclin b1 细胞周期