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目的:研究小干扰RNA(small interf ering RNA,siRNA )抑制轴突导向蛋白分子(Semaphorin 4C,Sema4C)基因的表达对人乳腺癌细胞MDA-MB-231 体外迁移、侵袭及增殖的影响,并初步探讨其作用机制。方法:根据Sema4C 基因设计序列特异性的siRNA(Sema4C-siRNA ),在脂质体介导下转染MDA-MB-231 细胞,Western blot方法检测基因封闭效应,利用细胞划痕实验、Tran ?swell小室侵袭实验及CFSE流式细胞仪方法检测细胞转染前后迁移、侵袭及增殖能力的变化,Western blot检测磷酸化AKT(p-AKT )在转染前后细胞中表达的变化。结果:转染Sema4C-siRNA 72小时后,乳腺癌MDA-MB-231 细胞株(MDA-MB-231/Si)Sema4C 蛋白表达明显下降,与未转染细胞MDA-MB-231 相比,MDA-MB-231/Si细胞体外迁移能力减弱,侵袭及增殖能力明显下降;p-AKT 表达水平在MDA-MB-231/Si细胞中明显降低。结论:Sema4C-SiRNA 转染人乳腺癌MDA-MB-231 细胞可下调细胞中Sema4C 蛋白表达水平,Sema4C-SiRNA 对MDA-MB-231 细胞的体外迁移、侵袭和增殖有抑制作用,这可能与p-AKT 的下调有关。 相似文献
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目的:探索乳腺癌细胞MDA-MB-231及MCF-7中CD44分子的表达水平差异及沉默CD44对乳腺癌细胞MDA-MB-231增殖、侵袭和迁移的影响。方法:利用qRT-PCR及Western blot技术检测细胞中CD44基因表达水平;设计并合成CD44的siRNA片段(CD44-siRNA)转染乳腺癌细胞,利用qRT-PCR、Western blot技术检测细胞中CD44基因表达水平的变化;MTT检测MDA-MB-231细胞增殖;Transwell侵袭实验检测MDA-MB-231细胞的迁移与侵袭能力变化。结果:CD44在侵袭性乳腺癌细胞MDA-MB-231中的表达高于非侵袭性乳腺癌细胞MCF-7,CD44-siRNA下调了 MDA-MB-231细胞中CD44 mRNA与蛋白水平的表达,并抑制了细胞的增殖和侵袭转移能力。结论:CD44-siRNA能够下调CD44的表达,并有效抑制乳腺癌细胞MDA-MB-231的增殖及其侵袭迁移力。 相似文献
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目的:探讨HIF-1α通过调节TP53INP1蛋白对乳腺癌细胞迁移及侵袭能力的影响。方法:通过免疫组化检测60例乳腺癌标本中TP53INP1和HIF-1α的表达情况及分析其临床病理资料。通过体外构建HIF-1α及共转染HIF-1α和TP53INP1细胞模型,迁移侵袭实验和划痕实验检测HIF-1α和TP53INP1对乳腺癌细胞迁移侵袭能力的影响;Western blot实验研究共同下调HIF-1α和 TP53INP1对MMP2、VEGF蛋白表达水平的影响。结果:免疫组化实验显示HIF-1α在人乳腺癌组织中高表达,TP53INP1呈低表达,两者之间具有相关性且均与淋巴结转移有关(P<0.05)。迁移侵袭实验及划痕实验显示在乳腺癌MDA-MB-231细胞和MCF-7细胞中下调HIF-1α抑制了迁移侵袭及愈合能力,MCF-7细胞中共同转染下调HIF-1α和TP53INP1质粒之后逆转了乳腺癌细胞的迁移侵袭能力及愈合能力(P<0.05)。Western blot实验显示shHIF-1α和TP53INP1共转染后在MDA-MB-231细胞中MMP2表达水平高于下调HIF-1α表达组。共转染shHIF-1α和shTP53INP1质粒后在MCF-7细胞中,VEGF和MMP2表达高于下调shHIF-1α组(P<0.05)。结论:HIF-1α可能通过下调TP53INP1的表达促进乳腺癌细胞的迁移侵袭能力。 相似文献
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目的 探讨UHRF1对乳腺癌MDA-MB-231细胞增殖以及侵袭的影响及其相关机制。方法 采用四甲基偶氮唑盐微量酶反应比色法(MTT)检测沉默UHRF1基因后对乳腺癌MDA-MB-231细胞活力的影响;应用克隆形成实验检测沉默UHRF1后对乳腺癌MDA-MB-231细胞存活的影响;吖啶橙-溴乙锭(AO/EB)检测沉默UHRF1后对乳腺癌MDA-MB-231细胞凋亡的影响;Caspase-3活性试剂盒检测沉默UHRF1后乳腺癌细胞Casapse-3活性的变化;Western blot法检测细胞中凋亡相关蛋白Bcl-2、Bax、Bad、p-Bad、XIAP、p53、p21Cip1/Waf1和p16INK4a的表达;应用Transwell实验研究沉默UHRF1对MDA-MB-231细胞侵袭能力的影响;Wound Healing实验研究沉默UHRF1后对其迁移能力的影响。结果 沉默UHRF1使乳腺癌MDA-MB-231细胞活力降低;克隆形成实验结果显示沉默UHRF1后MDA-MB-231细胞存活能力降低,AO/EB染色显示沉默UHRF1促进MDA-MB-231细胞凋亡。同时Caspase-3活性实验结果显示沉默UHRF1后乳腺癌MDA-MB-231细胞Caspase-3的活性增加;Western blot结果显示,沉默UHRF1后,能够使凋亡蛋白Bad、XIAP和Bax的表达上调,同时抗凋亡蛋白p-Bad,Bcl-2的表达下调,也使p53,p21Cip1/Waf1,p16INK4a蛋白表达升高;Transwell以及Wound Healing实验证明沉默UHRF1能够抑制乳腺癌MDA-MB-231细胞侵袭和迁移。结论 沉默UHRF1能够抑制乳腺癌MDA-MB-231细胞活力和存活,并抑制乳腺癌MDA-MB-231的侵袭和迁移。沉默UHRF1通过调控p53,p21Cip1/Waf1,p16INK4a信号发挥作用。 相似文献
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目的:研究干扰赖氨酰氧化酶(LOX)基因表达对乳腺癌细胞MDA-MB-231体外侵袭迁移能力的影响,并探讨相关机制及意义。方法:设计合成特异性慢病毒干扰载体(LOX-RNAi-LV)转染MDA-MB-231细胞,实时定量PCR检测LOX mRNA和侵袭转移相关因子MMP-2、MMP-9和HIF-1α的mRNA表达,蛋白质印迹法检测LOX蛋白表达,细胞侵袭迁移实验(Transwell)检测细胞侵袭迁移能力。结果:转染LOX-RNAi-LV后,乳腺癌MDA-MB-231细胞LOXmRNA和蛋白表达水平明显降低,与空白组相比,抑制率分别为89.2%和82.97%。MMP-2(F=225.271,P=0.0000)、MMP-9(F=35.373,P=0.000 01)和低氧诱导因子HIF-1α(F=341.081,P=0.0000)的mRNA表达均明显降低。细胞侵袭迁移实验显示,干扰组穿过Transwell小室滤过膜的细胞数明显少于对照组(侵袭实验F=269.557,P=0.000 5;迁移实验F=164.321,P=0.000 3)。结论:转染LOX-RNAi-LV能有效地干扰LOX在乳腺癌细胞MDA-MB-231中表达,抑制肿瘤细胞的侵袭和迁移能力,其作用机制可能与下调MMP-2、MMP-9和HIF-1α表达有关。 相似文献
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《Asian Pacific journal of cancer prevention》2014,15(14):5747-5751
Background: Coptisine, an isoquinoline alkaloid extracted from Coptidis rhizoma, has many biologicalactivities such as antidiabetic, antimicrobial and antiviral actions. However, whether coptisine exerts anti-cancermetastasis effects remains unknown. Materials and Methods: Effects of coptisine on highly metastatic humanbreast cancer cell MDA-MB-231 proliferation were evaluated by trypan blue assay and on cell adhesion, migrationand invasion by gelatin adhesion, wound-healing and matrigel invasion chamber assays, respectively. Expressionof two matrix metalloproteinases (MMPs), MMP-9, MMP-2 and their specific inhibitors tissue inhibitor ofmetalloproteinase 1 (TIMP-1) and tissue inhibitor of metalloproteinase 2 (TIMP-2) were analyzed by RT-PCR.Results: Coptisine obviously inhibited adhesion to an ECM-coated substrate, wound healing migration, andinvasion through the matrigel in MDA-MB-231 breast cancer cells. RT-PCR revealed that coptisine reduced theexpression of the ECM degradation-associated gene MMP-9 at the mRNA level, and the expression of TIMP-1was up-regulated in MDA-MB-231 cells, while the expression of MMP-2 and its specific inhibitor TIMP-2 wasnot affected. Conclusions: Taken together, our data showed that coptisine suppressed adhesion, migration andinvasion of MDA-MB-231 breast cancer cells in vitro, the down-regulation of MMP-9 in combination with theincrease of TIMP-1 possibly contributing to the anti-metastatic function. Coptisine might be a potential drugcandidate for breast cancer therapy. 相似文献
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[摘要] 目的:探讨环加氧酶-2(COX-2)在乳腺癌转移中的作用及其可能的机制。方法:收集从2015 年10 月至2018 年4 月在云南省肿瘤医院接受乳腺切除术的患者中获得的原发乳腺癌组织和脑转移乳腺癌组织临床病理样本共45 例,其中原发30 例、脑转移15 例。采用qPCR检测COX-2 在原位乳腺癌和脑转移乳腺癌组织中的表达。将COX-2 过表达重组病毒(LV6-COX2)或敲减COX-2 重组病毒(LV3-COX2 shRNA1、LV3-COX2 shRNA2)感染人乳腺癌MDA-MB-231 细胞并获得稳转细胞株后,CCK-8法检测COX-2 表达对MDA-MB-231 细胞增殖的影响,划痕实验和Transwell 法检测对MDA-MB-231 细胞迁移和侵袭的影响。qPCR和WB实验分析各组细胞中COX-2 mRNA和蛋白的表达水平,qPCR检测COX-2 表达对MDA-MB-231 细胞内EMT相关基因表达的影响。结果:COX-2 表达水平在脑转移乳腺癌患者组织中显著高于原位乳腺癌组织(P<0.01);并且与乳腺癌患者肿瘤TMN分期有关。成功构建稳定过表达/敲减COX-2 的MDA-MB-231 细胞株。过表达COX-2 促进MDA-MB-231 细胞的迁移和侵袭(均P<0.01),同时显著提高MMP2、MMP1、N-cadherin 和vimentin 的表达(均P<0.01),但对细胞增殖无明显影响;而沉默COX-2 则有相反的作用,且可促进细胞增殖(P<0.05)。结论:COX-2 在脑转移乳腺癌组织中高表达,其可能通过调控EMT过程促进乳腺癌MDA-MB-231 细胞的迁移和侵袭。 相似文献
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Rebeca López-Marure Estrella Zapata-Gómez Leticia Rocha-Zavaleta María Cecilia Aguilar Magali Espinosa Castilla Jorge Meléndez Zajgla 《Cancer biology & therapy》2016,17(9):915-924
Dehydroepiandrosterone (DHEA), an adrenal hormone, has a protective role against cancer. We previously shown that DHEA inhibits the proliferation and migration of cell lines derived from breast cancer; however, the role of DHEA in others events related with these effects are unknown. We hypothesized that DHEA inhibits the expression of proteins and some events related with cell migration and metastasis. We determined the migration in Boyden chambers, the invasion in matrigel, anchorage-independent growth and the formation of spheroids in 3 cell lines (MCF-7, MDA-MB-231, ZR-75-30) derived from breast cancer exposed to DHEA. The secretion of metalloproteinases (MMPs), tissue inhibitors of metalloproteinases (TIMPs), and several pro-inflammatory molecules in the secretome of these cells was also evaluated. DHEA inhibited the migration in transwells and the invasion in matrigel of MCF-7 and MDA-MB-231 cells. Besides, DHEA inhibited the anchorage-independent growth on agar and decreased the size of spheroids, and also reduced the secretion of IL-1α, IL-6, IL-8, and TNF-α in all cell lines. Metalloproteinase-1 (MMP-1) secretion was slightly decreased by DHEA treatment in MDA-MB-231 cells. Our results also showed that inhibition of migration and invasion induced by DHEA in breast cancer cells is correlated with the decrease of cytokine/chemokine secretion and the diminution of tumor cells growth. MCF-7 cells were the most responsive to the exposure to DHEA, whereas ZR-75-30 cells responded less to this hormone, suggesting that DHEA could be used in the treatment of breast cancer in early stages. 相似文献