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国家自然科学基金(30670811)

作品数:11 被引量:40H指数:3
相关作者:任国胜田甜王小毅李凡李佳更多>>
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REGγ基因沉默对甲状腺髓样癌细胞凋亡的影响被引量:2
2009年
目的:研究小干扰RNA(small interfering RNA,siRNA)方法抑制蛋白酶体激活剂REGγ的基因表达对人甲状腺髓样癌TT细胞株凋亡的影响。方法:培养人甲状腺髓样癌TT细胞株,构建真核表达载体pREGγ-shRNA及阴性对照质粒pNeg—shRNA,在脂质体Lipofectamine2000的介导下转染人甲状腺髓样癌TT细胞株,采用RT—PCR和Westem Blot法检测pREGγ-shRNA转染和pNeg—shRNA转染的TT细胞之间REGγ mRNA及REGγ蛋白质表达之间的差别;然后采用TUNEL法检测REGγ shRNA转染和阴性质粒转染的TT细胞的凋亡率之间的差别,以及Western Blot法检测pREGγ-shRNA转染和pNeg—shRNA转染的TT细胞凋亡因子Caspase-3表达之间的差别。结果:真核表达载体pREGγ-shRNA转染TT细胞的REGγ mRNA和REGγ蛋白表达水平明显低于pNeg—shRNA转染的TT细胞(P〈0.05),pREGγ-shRNA转染的TT细胞的凋亡水平明显高于pNeg—shRNA转染的TT细胞(P〈0.05)。结论:应用siRNA技术能有效的抑制人甲状腺髓样癌细胞REGγ基因的表达,抑制蛋白酶体激活剂REGγ基因的表达能明显地促进TT细胞的凋亡,提示REGγ可阻止细胞凋亡。REGγ在人甲状腺髓样癌细胞凋亡中的地位,为肿瘤的生物学治疗提供了新的思路。
张矛任国胜甘露
关键词:小干扰RNA甲状腺髓样癌
甲状腺乳头状癌组织REGγ、p21的蛋白表达及意义被引量:1
2010年
目的研究REGγ和p21在甲状腺乳头状癌组织和正常甲状腺组织中的表达及相互关系。方法标本取自20例甲状腺乳头状癌组织,10例正常甲状腺组织。Western blot检测甲状腺乳头状癌组织和正常甲状腺组织中REGγ和p21蛋白的表达。结果Western blot检测显示,甲状腺乳头状癌组织的REGγ表达明显高于正常甲状腺组织(P<0.05),甲状腺乳头状癌组织的p21表达明显低于正常甲状腺组织(P<0.05)。相关性分析显示,甲状腺乳头状癌组织REGγ和p21表达之间程显著的负相关关系(P<0.05)。结论REGγ蛋白的高表达和p21蛋白的低表达与甲状腺乳头状癌的形成有关,甲状腺乳头状癌组织中REGγ可能促进p21蛋白的降解。
张矛甘露任国胜
关键词:甲状腺乳头状癌P21蛋白酶体
人REG γ cDNA克隆及其稳定转染细胞株的建立
2009年
目的:构建真核表达重组体PcDNA3.1-REGγ的稳定转染细胞株,为进一步深入研究REGγ的功能奠定基础。方法:提取MCF-7细胞的总RNA为模板,通过RT-PCR获取全长REGγcDNA,经限制性内切酶EcoRⅠ、EcoRⅤ酶切后与PcDNA3.1连接构建重组体,再经内切酶酶切及DNA序列分析鉴定重组体构建正确。以Lipofectamine2000将重组体导入HBL-100细胞,由G418筛选出稳定转染细胞株。结果:经免疫细胞化学和RT-PCR检测并证实稳定转染细胞株中有REGγ的高表达。结论:真核表达重组体PcDNA3.1-REGγ构建成功,并经抗生素筛选获得了稳定高表达REGγ的细胞株,为进一步研究REGγ的功能奠定了基础。
田甜王小毅李凡任国胜
关键词:CDNA克隆转染
蛋白酶体激活因子REGgamma基因对乳腺癌MDA-MB-231细胞生长的影响被引量:3
2008年
目的观察蛋白酶体激活因子REGgamma(REGγ)基因对乳腺癌MDA-MB-231细胞生长的影响。方法构建真核表达重组体PcDNA3.1-REGγ并将其以脂质体转染法导人细胞,以600mg/L浓度的G418连续筛选转染细胞6周,获得稳定高表达该基因的细胞株。分别以噻唑蓝(MTT)、流式细胞仪(FCM)、软琼脂集落形成试验检测其对细胞生长的影响。免疫细胞化学检测增殖细胞核抗原(PCNA)的表达。结果转染REGγ的细胞有外源REGγ基因的整合及表达,经Westernblot检测其表达较未转染组和仅转染空载体组明显增加。MTT法检测发现转染REGγ细胞生长加速;FCM检测提示转染REGγ组、未转染组和仅转染空载体组在S+G2+M增殖期的细胞比例分别为55.91%、44.09%、43.69%;软琼脂集落形成试验显示其平均集落形成率分别为10.23%、3.67%、4.06%。转染REGγ基因后PCNA表达明显增强。结论转染外源野生型REG7基因对乳腺癌MDA-MB-231细胞具有加速其生长、促进其增殖的作用。
田甜王小毅李凡任国胜李佳
关键词:乳腺癌细胞周期增殖
Breast cancer development and progression: Risk factors, cancer stem cells, signaling pathways, genomics, and molecular pathogenesis被引量:1
2018年
As the most commonly occurring cancer in women worldwide,breast cancer poses a formidable public health challenge on a global scale.Breast cancer consists of a group of biologically and molecularly heterogeneous diseases originated from the breast.While the risk factors associated with this cancer varies with respect to other cancers,genetic predisposition,most notably mutations in BRCA1 or BRCA2 gene,is an important causative factor for this malignancy.Breast cancers can begin in different areas of the breast,such as the ducts,the lobules,or the tissue in between.Within the large group of diverse breast carcinomas,there are various denoted types of breast cancer based on their invasiveness relative to the primary tumor sites.It is important to distinguish between the various subtypes because they have different prognoses and treatment implications.As there are remarkable parallels between normal development and breast cancer progression at the molecular level,it has been postulated that breast cancer may be derived from mammary cancer stem cells.Normal breast development and mammary stem cells are regulated by several signaling pathways,such as estrogen receptors(ERs),HER2,and Wnt/b-catenin signaling pathways,which control stem cell proliferation,cell death,cell differentiation,and cell motility.Furthermore,emerging evidence indicates that epigenetic regulations and noncoding RNAs may play important roles in breast cancer development and may contribute to the heterogeneity and metastatic aspects of breast cancer,especially for triple-negative breast cancer.This review provides a comprehensive survey of the molecular,cellular and genetic aspects of breast cancer.
Yixiao FengMia SpeziaShifeng HuangChengfu YuanZongyue ZengLinghuan ZhangXiaojuan JiWei LiuBo HuangWenping LuoBo LiuYan LeiScott DuAkhila VuppalapatiHue H.LuuRex C.HaydonTong-Chuan HeGuosheng Ren
关键词:BRCA1/2
REGgamma在胃癌组织与不同分化胃癌细胞株中的表达及其临床意义
2009年
目的:探讨REGgamma(REGγ)在胃癌组织与不同分化细胞株中的表达水平及其临床意义。方法:应用免疫组化SP法检测70例胃癌组织和30例正常胃组织中REGγ/蛋白的表达水平,并分析其与胃癌生物学行为的关系:分别采用逆转录-聚合酶链反应(RT-PCR)及Western blot检测正常胃粘膜细胞株(GES-1)、胃癌高分化(MKN-28)、中分化(SGC-7901)和低分化(BGC823)细胞株中REGγ mRNA转录情况和蛋白表达水平。结果:免疫组化结果表明在胃癌组织中,REGγ蛋白的表达阳性率(74.3%)显著高于正常胃粘膜组织(40.0%,P<0.01),且表达与肿瘤的大小(P<0.01)、淋巴结转移(P<0.05)、分化程度(P<0.01)、浸润程度(P<0.01)及远处转移(P<0.05)相关;RT-PCR结果表明在正常胃粘膜细胞株及胃癌高、中、低分化细胞株中,REGγ mRNA表达水平逐渐增强,分别为0.459±0.079、0.588±0.118、0.715±0.066、0.873±0.099(P<0.01),Western blot结果表明在正常胃粘膜细胞株及胃癌高、中、低分化细胞株中,REGγ蛋白表达水平逐渐增强,分别为0.712±0.065、1.176±0.185、1.533±0.127、2.061±0.398,结果差别有统计学意义(P<0.05)。结论:REGγ在正常胃粘膜组织和胃癌组织中呈一定比例的阳性表达,其表达水平与胃癌癌肿大小、是否有淋巴结转移、分化程度、浸润程度及是否远处转移等肿瘤的恶性生物学行为有关。REGγ表达与胃癌的分化程度相关,随着细胞分化程度的降低其表达逐渐增强,检测REG7 mRNA及蛋白的表达可望成为胃癌早期诊断和判断预后的分子指标之一。
李佳田甜王小毅李凡任国胜
关键词:胃肿瘤免疫组织化学蛋白质
REGgamma基因对乳腺癌细胞MCF-7增殖和凋亡的影响被引量:4
2008年
目的:探讨蛋白酶体激活因子REGgamma(REGγ)基因对乳腺癌细胞MCF-7增殖和凋亡的作用。方法:构建真核表达重组体PcDNA3.1-REGγ,通过脂质体转染将REGγ基因导入MCF-7,以G418(800mg/L)连续筛选获得稳定高表达REGγ基因的细胞株,以转染空载体和未转染细胞作为对照。分别利用MTT法及免疫细胞化学方法检测各组细胞PCNA的表达分析REGγ基因对细胞增殖的影响;以Caspase-3分光光度法及Annexin V-FITC的流式细胞仪(FCM)来检测细胞凋亡的变化;以透射电镜(TEM)观察细胞超微结构的改变。结果:获得稳定转染且高表达REGγ的细胞株。经Western Blot检测转染REGγ基因的细胞(实验组),其REGγ蛋白表达明显高于对照组细胞(P<0.05);未转染组、空载体组和实验组细胞的倍增时间分别为34.6、35.1、26.7h,提示实验组细胞倍增时间缩短;MTT法绘制生长曲线提示实验组细胞生长明显加速;免疫细胞化学检测PCNA结果显示,未转染组、空载体组和实验组细胞的染色灰度值分别为89.61±14.32、87.95±16.38、133.47±8.14,表明实验组细胞PCNA表达明显增强(P<0.01);Caspase-3分光光度法检测显示,实验组细胞的OD值普遍低于对照组(P<0.05);Annexin V-FITC的FCM检测显示,实验组细胞的凋亡率明显低于对照组(P<0.01);TEM观察实验组细胞表现为核仁肥大,线粒体、高尔基体丰富或扩张,未见凋亡小体形成。结论:REGγ基因具有促进乳腺癌MCF-7细胞增殖并抑制其凋亡的作用。
田甜王小毅李佳李凡任国胜
关键词:乳腺癌MCF-7细胞细胞增殖细胞凋亡
Breast cancer development and progression:Risk factors,cancer stem cells,signaling pathways,genomics,and molecular pathogenesis被引量:28
2018年
As the most commonly occurring cancer in women worldwide,breast cancer poses a formidable public health challenge on a global scale.Breast cancer consists of a group of biologically and molecularly heterogeneous diseases originated from the breast.While the risk factors associated with this cancer varies with respect to other cancers,genetic predisposition,most notably mutations in BRCA1 or BRCA2 gene,is an important causative factor for this malignancy.Breast cancers can begin in different areas of the breast,such as the ducts,the lobules,or the tissue in between.Within the large group of diverse breast carcinomas,there are various denoted types of breast cancer based on their invasiveness relative to the primary tumor sites.It is important to distinguish between the various subtypes because they have different prognoses and treatment implications.As there are remarkable parallels between normal development and breast cancer progression at the molecular level,it has been postulated that breast cancer may be derived from mammary cancer stem cells.Normal breast development and mammary stem cells are regulated by several signaling pathways,such as estrogen receptors(ERs),HER2,and Wnt/b-catenin signaling pathways,which control stem cell proliferation,cell death,cell differentiation,and cell motility.Furthermore,emerging evidence indicates that epigenetic regulations and noncoding RNAs may play important roles in breast cancer development and may contribute to the heterogeneity and metastatic aspects of breast cancer,especially for triple-negative breast cancer.This review provides a comprehensive survey of the molecular,cellular and genetic aspects of breast cancer.
Yixiao FengMia SpeziaShifeng HuangChengfu YuanZongyue ZengLinghuan ZhangXiaojuan JiWei LiuBo HuangWenping LuoBo LiuYan LeiScott DuAkhila VuppalapatiHue H.LuuRex C.HaydonTong-Chuan HeGuosheng Ren
关键词:BRCA1/2
转染REGγ基因对乳腺癌细胞裸鼠移植瘤生长的影响被引量:1
2009年
目的:研究转染蛋白酶体激活因子REGγ基因的乳腺癌MDA-MB-231细胞在裸鼠体内的成瘤作用及其机制。方法:以脂质体转染法将构建的重组质粒pcDNA3.1-REGγ导入MDA-MB-231细胞,以G418(600mg/L)筛选获得稳定高表达该基因的细胞株(实验组)。以转染空载体及未施加处理因素的细胞作为对照组。将此3组细胞接种于裸鼠皮下,观察移植瘤的生长情况并计算抑瘤率。RT-PCR检测REGγ基因在移植瘤中的表达,FCM检测移植瘤的肿瘤浸润淋巴细胞(tumor-infiltra-tinglymphocyte,TIL)中CD16的表达以及肿瘤细胞周期和细胞凋亡,免疫组织化学法检测移植瘤中p21的表达。结果:与对照组相比,实验组的移植瘤生长速度较快、体积较大,瘤体质量明显增加(P<0.05);REGγ基因在移植瘤中的表达增加(P<0.01);FCM检测提示CD16阳性率明显降低(P<0.05);G0/G1和G2/M期细胞减少,S期细胞明显增多,肿瘤细胞的凋亡率明显降低(P<0.05);p21的表达明显降低(P<0.05)。结论:REGγ基因在体内具有促进乳腺癌发生、发展的作用,其机制可能与加速细胞周期、抑制细胞凋亡、抑制自然杀伤细胞活化以及对p21的特异性降解有关。
田甜任国胜李晓涛
关键词:基因转染技术异种移植模型抗肿瘤试验
Expression and Clinical Significance of REGy in Gastric Cancer Tissue and Variously Differentiated Gastric Cancer Cell Lines
2009年
OBJECTIVE To evaluate the REGγ expression in gastric cancer tissue and gastric cancer cell lines of various differentiation levels and its clinical significance. METHODS Immunohistochemistry was used to detect the expression of REGγ protein in 70 specimens of gastric cancer and 30 specimens of normal gastric mucosa. The relationship between the expression of REGγ protein and the biological behaviors of gastric cancer was analyzed. RT-PCR and Western blot were used to detect the mRNA level and the protein expression of REGγ in normal gastric cell line GES-1, well differentiated gastric cancer cell line MKN-28, moderately differentiated gastric cancer cell line SGC-7901 and poorly differentiated gastric cancer cell line BGC-823. RESULTS The expression rate of REGγ protein in gastric cancer tissue (52/70, 74.29%) was significantly higher than that in normal gastric tissue (12/30, 40%) (P 〈 0.01). The expression rate of REGγ was correlated with tumor size (P 〈 0.01), lymph node metastasis (P 〈 0.05), differentiation degree (P 〈 0.01), infiltration depth (P 〈 0.01) and distant metastasis (P 〈 0.05). RT-PCR analysis showed that the expression of REGγ mRNA was 0.459 ± 0.079 in the normal gastric mucosa cell ling 0.588 ±0.118 in the well differentiated gastric cancer cell line, 0.715±0.066 in the moderately differentiated gastric cancer cell line, and 0.873 ± 0.099 in the poorly differentiated gastric cancer cell line, showing a negative correlation between REGγ mRNA expression and differentiation level (P 〈 0.05). Western blot analysis showed that the expression of REGγ protein was 0.712±0.065 in the normal gastric mucosa cell line, 1.176±0.185 in the well differentiated gastric cancer cell line, 1.533 ± 0.127 in the moderately differentiated gastric cancer cell line, and 2.061± 0.398 in the poorly differentiated gastric cancer cell line, showing a negative correlation between REGγ protein expression and differentiation level (P 〈 0.05). CONCLUSIO
Jia Li Tian Tian Xiaoyi Wang Fan Li Guosheng Ren
关键词:IMMUNOHISTOCHEMISTRYGENE
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