Caprin-1 is a novel microRNA-223 target for regulating the proliferation and invasion of human breast cancer cells

MicroRNAs (miRNAs) are 21–22 nucleotides regulatory small non-coding RNAs that inhibit gene expression by binding to complementary sequences especially the 3’ untranslated region (3’UTR) of mRNA. One miRNA can target many messenger RNAs, leading to a complex metabolic network. Previous studies have shown that miRNA-223 regulates migration and invasion of tumor cells and targets cytoplasmic activation/proliferation-associated protein-1 (Caprin-1). In the present study, we detected the expression of miRNA-223 and Caprin-1 in MCF-7, T-47D and MDA-MB-231 cancer cell lines, and MCF-10A normal breast cell line, and analyzed the role of miRNA-223 in Caprin-1-induced proliferation and invasion of human breast cancer cells. We found that miRNA-223 expression levels are significantly lower in MCF-7, T-47D and MDA-MB-231 cancer cells than in MCF-10A normal breast cells, while Caprin-1 expression is higher in cancer cells than in normal breast cells. The most malignant cancer cell line MDA-MB-231 has the lowest expression of miR-223, but the highest expression of Caprin-1. Further, we found that miR-223 targets the 3’UTR of Caprin-1 miRNA and down-regulates the expression of Caprin-1. We also found that over-expression of Caprin-1 can promote the proliferation and the invasion of breast cancer cells, but miRNA-223 can inhibit the proliferation and the invasion. miRNA-223-induced inhibition can be reversed by ectopic over-expression of Caprin-1. These findings suggest that miR-223 may suppress the proliferation and invasion of cancer cells by directly targeting Caprin-1. Our study also indicates that expression levels of miR-223 and Caprin-1 can be used to predict the state of cancer in breast cancer patient.     

硫酸乙酰肝素蛋白聚糖对乳腺癌细胞的抑制与诱导凋亡作用

目的：观察两种硫酸乙酰肝素蛋白聚糖（HSPG）对MDA-MB-231乳腺癌细胞生长的影响及机制。方法：利用MTT 法和生长曲线的绘制检测不同浓度的HSPG在不同时间对MDA-MB-231细胞的抑制作用, 流式细胞术结合Annexin V-FITC 及PI双标记染色检测分析HSPG 对MDA-MB-231细胞凋亡的影响。结果： MTT法测得不同浓度MCF-7 type HSPG作用于MDA-MB-231后24h、48h、72h均可明显抑制肿瘤细胞生长，并且呈现出剂量依赖性；生长曲线结果显示不同浓度MCF-7 type HSPG可明显抑制MDA-MB-231的生长；流式细胞术显示MCF-7 type HSPG诱导MDA-MB-231细胞凋亡增加；MDA-MB-231 type HSPG对其源细胞（MDA-MB-231）则无明显的生长抑制作用，对其凋亡率无明显影响。结论：MCF-7 type HSPG对MDA-MB-231细胞生长具有显著的抑制作用，其机制可能与诱导细胞凋亡有关。     

Stathmin表达水平与乳腺癌细胞侵袭能力相关性研究

目的 探讨乳腺癌细胞MDA-MB-231和MCF-7中Stathmin基因表达水平与细胞生长、黏附、侵袭等生物学行为之间的关系,为进一步研究乳腺癌转移机制奠定实验基础。方法 应用RT-PCR和Western Blot方法检测MDA-MB-231和MCF-7细胞中Stathmin基因的表达水平,同时利用细胞增殖试验、细胞黏附试验和细胞侵袭试验检测MDA-MB-231和MCF-7细胞的生长、黏附、侵袭能力,分析Stathmin表达与细胞的生长、黏附、侵袭能力之间的关系。结果 RT-PCR和Western Blot检测结果显示,Stathmin基因在MDA-MB-231和MCF-7细胞中表达均高于正常对照细胞(F=10.173,P<0.05),且MDA-MB-231细胞中的表达水平明显高于MCF-7细胞中的表达水平(t=4.562,P<0.05)。而MDA-MB-231细胞在生长、黏附和侵袭能力方面均强于MCF-7细胞(P<0.05)。结论 Stathmin表达水平高的乳腺癌细胞相应的生长、黏附、侵袭能力较强,Stathmin表达水平与细胞侵袭能力密切相关。     

Tamoxifen-resistant human breast cancer cell growth: inhibition by thioridazine, pimozide and the calmodulin antagonist, W-13.

Estrogen receptor (ER)-negative human breast cancer cell lines (MDA-MB-231 and MDA-MB-435) and ER-positive derivatives of the MCF-7 cell line selected for growth in the presence of antiestrogens (LY2 and RR) were used as in vitro models of tamoxifen-resistant human breast cancer in this study. The sensitivity of the tamoxifen-sensitive (MCF-7) and tamoxifen-resistant human breast cancer cell growth to two noncytotoxic neuroleptic drugs, pimozide and thioridazine, and the anticalmodulin agent, W-13, were compared. Inhibition of cell growth was measured as a decrease in cell number following a 72-h incubation with drug. Growth of the ER-negative cell lines MDA-MB-231 and MDA-MB-435 was inhibited by all three drugs. The average Ki values in these two lines were 6.3 and 3.8 microM for pimozide and 4.1 and 15 microM for thioridazine, respectively. Both ER-negative cell lines were more sensitive than MCF-7 cells to growth inhibition by W-13. MCF-7 cells selected for antiestrogen resistance were sensitive to growth inhibition by W-13 and thioridazine (LY2, average Ki = 10.4 microM; RR, average Ki = 5.2 microM). LY2 and RR cells were resistant to pimozide except when treated with estradiol (Ki = 4.6 and 7.9 microM, respectively). Pimozide, thioridazine and W-13 all exerted different effects on the distribution of human breast cancer cells within the cell cycle, suggesting that each drug may utilize a distinct pathway for inhibition of cell growth. We conclude that all three drugs are potential noncytotoxic alternatives to tamoxifen for the treatment of tamoxifen-resistant human breast cancer.     

Cyclin E2 overexpression is associated with endocrine resistance but not insensitivity to CDK2 inhibition in human breast cancer cells

Cyclin E2, but not cyclin E1, is included in several gene signatures that predict disease progression in either tamoxifen-resistant or metastatic breast cancer. We therefore examined the role of cyclin E2 in antiestrogen resistance in vitro and its potential for therapeutic targeting through cyclin-dependent kinase (CDK) inhibition. High expression of CCNE2, but not CCNE1, was characteristic of the luminal B and HER2 subtypes of breast cancer and was strongly predictive of shorter distant metastasis-free survival following endocrine therapy. After antiestrogen treatment of MCF-7 breast cancer cells, cyclin E2 mRNA and protein were downregulated and cyclin E2-CDK2 activity decreased. However, this regulation was lost in tamoxifen-resistant (MCF-7 TAMR) cells, which overexpressed cyclin E2. Expression of either cyclin E1 or E2 in T-47D breast cancer cells conferred acute antiestrogen resistance, suggesting that cyclin E overexpression contributes to the antiestrogen resistance of tamoxifen-resistant cells. Ectopic expression of cyclin E1 or E2 also reduced sensitivity to CDK4, but not CDK2, inhibition. Proliferation of tamoxifen-resistant cells was inhibited by RNAi-mediated knockdown of cyclin E1, cyclin E2, or CDK2. Furthermore, CDK2 inhibition of E-cyclin overexpressing cells and tamoxifen-resistant cells restored sensitivity to tamoxifen or CDK4 inhibition. Cyclin E2 overexpression is therefore a potential mechanism of resistance to both endocrine therapy and CDK4 inhibition. CDK2 inhibitors hold promise as a component of combination therapies in endocrine-resistant disease as they effectively inhibit cyclin E1 and E2 overexpressing cells and enhance the efficacy of other therapeutics.     

2-Methoxy-5((3,4,5-trimethosyphenyl)seleninyl) phenol (SQ0814061), a novel microtubule inhibitor,evokes G2/M cell cycle arrest and apoptosis in human breast cancer cells

Breast cancer is the leading cause of cancer death in women worldwide, and novel chemotherapeutic drugs with high activity and no drug resistance for treating breast cancer are needed urgently. In this study, we investigated the antitumor effect of 2-methoxy-5((3,4,5-trimethosyphenyl)seleninyl) phenol (SQ0814061), which has a strong inhibition of cell growth in MCF-7 and MDA-MB-231 cells. We demonstrated that SQ0814061 (SQ) time-dependently induced cell cycle arrest at G2/M phase and subsequently progressed into apoptosis, which is associated with microtubule depolymerization. Western blot analysis revealed that up-regulation of cyclin B1 and Aurora A was related with G2/M phase arrest in MCF-7 and MDA-MB-231 cells treatment with SQ. However, the formation of multinucleated cells after a long time exposed to SQ of MCF-7 cells delayed the cell death. In addition, apoptosis induced by SQ is correlated with the down-regulation of the PI3K-Akt-MDM2 pathway in MCF-7 and MDA-MB-231 cells. Treatment with the PI3K specific inhibitor, LY294002, increased SQ-induced cell growth inhibitory rate and apoptosis rate of MCF-7 and MDA-MB-231 cells. Moreover, SQ induced MCF-7 and MDA-MB-231 cells to generate reactive oxygen species (ROS), and the SQ-induced cell death was ROS dependent. In conclusion, all the data demonstrated that SQ exhibited its antitumor activity through disrupting the microtubule assembly, inducing cell cycle arrest and eventually apoptosis which is associated with PI3K-Akt-MDM2 pathway in MCF-7 and MDA-MB-231 cells. Therefore, the novel compound SQ is a promising microtubule inhibitor that has tremendous potentials for therapeutic treatment of human mastocarcinoma.     

Drug efflux by breast cancer resistance protein is a mechanism of resistance to the benzimidazole insulin-like growth factor receptor/insulin receptor inhibitor, BMS-536924

Preclinical investigations have identified insulin-like growth factor (IGF) signaling as a key mechanism for cancer growth and resistance to clinically useful therapies in multiple tumor types including breast cancer. Thus, agents targeting and blocking IGF signaling have promise in the treatment of solid tumors. To identify possible mechanisms of resistance to blocking the IGF pathway, we generated a cell line that was resistant to the IGF-1R/InsR benzimidazole inhibitors, BMS-554417 and BMS-536924, and compared expression profiles of the parental and resistant cells lines using Affymetrix GeneChip Human Genome U133 arrays. Compared with MCF-7 cells, breast cancer resistance protein (BCRP) expression was increased 9-fold in MCF-7R4, which was confirmed by immunoblotting and was highly statistically significant (P = 7.13E-09). BCRP was also upregulated in an independently derived resistant cell line, MCF-7 924R. MCF-7R4 cells had significantly lower intracellular accumulation of BMS-536924 compared with MCF-7 cells. Expression of BCRP in MCF-7 cells was sufficient to reduce sensitivity to BMS-536924. Furthermore, knockdown of BCRP in MCF-7R4 cells resensitized cells to BMS-536924. Four cell lines selected for resistance to the pyrrolotriazine IGF-1R/InsR inhibitor, BMS-754807, did not have upregulation of BCRP. These data suggest that benzimidazole IGF-1R/InsR inhibitors may select for upregulation and be effluxed by the ATP-binding cassette transporter, BCRP, contributing to resistance. However, pyrrolotriazine IGF-1R/InsR inhibitors do not appear to be affected by this resistance mechanism.     

印记基因SLC22A18的表达与乳腺癌侵袭能力的关系

目的:研究印记基因SLC22A18(solute carrier family 22,member 18)在乳腺癌中的表达情况及其与乳腺癌侵袭能力的关系。方法:采用Transwell方法评估2种不同恶性程度的乳腺癌细胞株MDA-MB-231(恶性程度高)和MCF-7(恶性程度低)的侵袭转移能力。分别采用实时荧光定量逆转录聚合酶链反应(RT-PCR)和蛋白质印迹法检测SLC22A18的mRNA和蛋白在这2种乳腺癌细胞株中的表达情况。结果:MDA-MB-231细胞株恶性程度高,穿过膜的细胞多,侵袭能力强;MCF-7细胞株恶性程度低,穿过膜的细胞少,侵袭能力弱;SLC22A18在MCF-7中的mRNA和蛋白表达水平高于MDA-MB-231;差异有统计学意义(P0.01)。结论:印记基因SLC22A18的表达与乳腺癌细胞的侵袭能力相关,该基因有望作为一个抑癌基因抑制乳腺癌的转移。     

Rab27A在4种不同人乳癌细胞株中的表达

目的检测Rab27A在4种人乳癌细胞中的定位、表达情况,初步研究Rab27A表达对乳癌细胞生物学特性的影响。方法采用RT-PCR技术,检测Rab27A mRNA在4种人乳癌细胞MCF-7、MDA-MB-231、MDA-MB-435和MDA-MB-435HM中的表达并进行半定量分析。结果Rab27A mRNA在人乳癌细胞MDA-MB-231、MDA-MB-435及MDA-MB-435HM中的表达水平分别是MCF-7中的2.1、3.4和6.9倍,差异有显著意义（F=17.74～136.23,P〈0.05、0.01）;Rab27A蛋白在人乳癌细胞MDA-MB-231、MDA-MB-435及MDA-MB-435HM中的表达分别是MCF-7中的2.8、4.9和9.2倍,差异有显著性（F=37.74～154.29,P〈0.05、0.01）。Rab27A蛋白弥散性分布于MCF-7、MDA-MB-231和MDA-MB-435细胞浆中,MDA-MB-231和MDA-MB-435中又可见Rab27A蛋白在核周凝聚。结论Rab27A表达可能与乳癌细胞侵袭转移能力有关。     

The selective retinoid X receptor agonist bexarotene (LGD1069, Targretin) prevents and overcomes multidrug resistance in advanced breast carcinoma

Acquired drug resistance represents a major challenge in the therapeutic management of breast cancer patients. We reported previously that the retinoid X receptor-selective agonist bexarotene (LGD1069, Targretin) was efficacious in treating animal models of tamoxifen-resistant breast cancer. The goal of this study was to evaluate the effect of bexarotene on development of acquired drug resistance and its role in overcoming acquired drug resistance in advanced breast cancer. Paclitaxel, doxorubicin, and cisplatin were chosen as model compounds to determine the effect of bexarotene on the development of acquired drug resistance. Human breast cancer cells MDA-MB-231 were repeatedly treated in culture with a given therapeutic agent with or without bexarotene for 3 months. Thereafter, cells were isolated and characterized for their drug sensitivity. Compared with parental cells, cells treated with a single therapeutic agent became resistant to the therapeutic agent, whereas cells treated with the bexarotene combination remained chemosensitive. Cells with acquired drug resistance, when treated with the combination, showed increased sensitivity to the cytotoxic agent. Furthermore, cells treated with the combination regimen had reduced invasiveness and angiogenic potential than their resistant counterparts. These in vitro findings were further confirmed in an in vivo MDA-MB-231 xenograft model. Our results suggest a role for bexarotene in combination with chemotherapeutic agents in prevention and overcoming acquired drug resistance in advanced breast carcinoma.     

5,7,4⿲-Trihydroxy-6,8-diprenylisoflavone and lupalbigenin,active components of Derris scandens,induce cell death on breast cancer cell lines

BackgroundNatural products are a potential source for cancer chemotherapeutic development. This current study was performed to investigate the anti-tumor potential of 5,7,4⿲-trihydroxy-6,8-diprenylisoflavone (TD) and lupalbigenin (LB), plant flavonoids found in Derris scandens Benth (family: Leguminosae), in cancer and normal cell lines.MethodsThe human breast cancer cell lines MCF-7, MDA-MB-231 and MDA-MB-468, the human colon cancer cell line SW-620, and the mouse fibroblast cell line L-929 were used to test their anti-cancer activity. Apoptotic cell levels were measured by staining with annexin-V and propidium iodide and Western blot analysis was performed to confirm the apoptotic mechanism.ResultsThe results revealed that TD and LB showed specific cytotoxicity against MDA-MB-231 and MCF-7 cells. To elucidate mode of cell death via cytotoxic activities, breast cancer cell lines were treated. TD and LB induced MDA-MB-231 and MCF-7 cells to apoptosis, with the highest number of apoptotic cells at 24 and 72 h, respectively. Furthermore, TD and LB inhibited cell cycle progression via up-regulation of p21. Both compounds stimulated apoptosis through down-regulation of bcl-2, up-regulation of bax and releasing of cytochrome C proteins.ConclusionsTD and LB have significant anti-cancer effects against human breast cancer cells via cell cycle arrest and the induction of apoptosis through mitochondria signaling pathways, and may be potential anti-cancer agents for the treatment of breast cancer.     

Cyclin A-associated kinase activity is needed for paclitaxel sensitivity

Cyclin A-associated kinases, such as cyclin-dependent kinase 2 (CDK2), participate in regulating cellular progression from G(1) to S to G(2), and CDK2 has also been implicated in the transition to mitosis. The antitumor properties of CDK inhibitors, alone or in combination with taxanes, are currently being examined in clinical trials. Here, we examined whether the activity of kinases associated with cyclin A (such as CDK2) is important in determining cellular sensitivity to paclitaxel, a taxane and mitotic inhibitor used in chemotherapy for breast and ovarian cancer. We used adenoviral suppression or overexpression to manipulate the expression of CDK2 and cyclin A in one breast cancer and three ovarian cancer cell lines with different sensitivities to paclitaxel and assessed protein expression, kinase activity, cell cycle distribution, and sensitivity to paclitaxel. Transfection of a dominant-negative (DN)-CDK2 evoked resistance to paclitaxel by preventing cellular progression to mitosis through loss of CDK1 activity. Reexpression of wild-type CDK2 in DN-CDK2-transfected cancer cells restored CDK2 activity but not paclitaxel sensitivity. However, expression of cyclin A in DN-CDK2-transfected cells restored their sensitivity to paclitaxel. Although CDK2 activity was not directly involved in paclitaxel sensitivity, cyclin A-associated kinases did up-regulate CDK1 via phosphorylation. We conclude that cyclin A-associated kinase activity is required for these cells to enter mitosis and undergo paclitaxel-induced cell death. Combining taxane chemotherapy with any drug targeting cyclin A-associated kinases (e.g., pure CDK2 inhibitors) should be done with caution, if at all, because of the potential for enhancing taxane resistance.     

Genes commonly upregulated by hypoxia in human breast cancer cells MCF-7 and MDA-MB-231.

Hypoxia is a stress that causes alterations in signal transduction and gene instability. In the cancer microenvironment, hypoxia plays a significant role in forming a tumor phenotype and tumor progression. We aimed to identify the genes upregulated by hypoxia in human breast cancer cell lines, a hormone-dependent MCF-7 and a hormone-independent MDA-MB-231, using microarray analysis. These cells were exposed to two oxygen concentrations such as 21% and 1% in a time-course. Out of 12625 genes, 26 genes were identified as commonly upregulated in both MCF-7 and MDA-MB-231 cells. Some of these genes were already reported as hypoxia-related, but some of those were identified newly. These commonly upregulated genes between hormone-dependent and hormone-independent cells would be a clue to study hypoxia-related events and to explore the novel therapeutic targets in human breast cancer.     

表没食子儿茶素没食子酸酯对乳腺癌细胞生长及迁移的影响

目的：探讨表没食子儿茶素没食子酸酯（EGCG）对乳腺癌细胞MCF-7生长的影响及对乳腺癌细胞MDA-MB-231迁移的影响。方法：MCF-7细胞培养贴壁之后,加入EGCG处理,2d后收集蛋白,采用Western Blot检测磷酸化p38丝裂原活化蛋白激酶（phospho-p38MAPK）的表达;同样处理后收集活细胞,用细胞计数法检测细胞的存活;取对数生长期的MDA-MB-231细胞,分至6孔板培养,使用EGCG处理后,采用细胞划线法探测乳腺癌细胞的迁移。结果：使用EGCG处理乳腺癌细胞后,phospho-p38MAPK的表达降低,EGCG处理乳腺癌细胞4d后其增殖率降低50%,迁移活性降低。结论：EGCG处理乳腺癌细胞能抑制肿瘤细胞的生长以及迁移,这与p38 MAPK信号通路相关。     

TNF-α可诱导乳腺癌细胞凋亡

目的研究TNF-α对乳腺癌的影响。方法采用RT-PCR和WesternBlotting分析30例乳腺浸润性导管癌及癌旁正常乳腺组织,乳腺正常上皮细胞系及乳腺癌细胞系中TNF-α的表达情况;采用流式细胞术观察TNF-α对乳腺癌细胞凋亡的影响。结果RT-PCR和Western Blotting结果碌示,TNF-αmRNA和蛋白在乳腺癌组织中表达都明显低于配对的癌旁正常组织（P〈0．05）,在乳腺上皮细胞系中表达均高于乳腺癌三利一细胞系,流式细胞术检测结果显示与未处理组桐比．经TNF-α处理的MDA-MB-435S（16．7±0．31）和MCF7（18．6±0．42）细胞的凋亡率明显增加,差异均有统计学意义（P〈0．05）。结论TNF-α可促进乳腺癌细胞的凋亡,TNF-α可为乳腺癌的治疗提供新靶点。     

Administration with hyperoside sensitizes breast cancer cells to paclitaxel by blocking the TLR4 signaling

Breast cancer is a malignancy and one of the most frequent causes of cancer death among women worldwide. Paclitaxel is a common chemotherapeutic drug and has recently been shown to facilitate tumor cell escape during cytotoxic chemotherapy by inducing inflammatory mediators and pro-survival protein expression. Hyperoside is a flavonoid glycoside compound and exerts anti-inflammation, and anti-tumor growth properties. However, its function in breast cancer chemosensitivity remains poorly elucidated. In this study, hyperoside exhibited little cytotoxicity to normal human breast mammary epithelial cell lines, and also protected against paclitaxel-induced cytotoxicity in MCF-10A. Importantly, treatment with hyperoside engendered not only inhibition of cell viability, but also potentiated cancer cell sensitivity to paclitaxel in TLR4-positive breast cancer MDA-MB-231 cells by suppressing cell viability, and increasing cell apoptosis and caspase-3 activity. Nevertheless, although hyperoside exposure restrained cell viability, its treatment presented little effects to paclitaxel sensitivity in TLR4-null HCC1806 cells. Intriguingly, paclitaxel stimulation activated the TLR4-NF-κB signaling, which was reversed after hyperoside administration. Concomitantly, hyperoside also attenuated paclitaxel-mediated anti-apoptotic Bcl-2 expression, but enhanced the effects of paclitaxel on pro-apoptotic Bax expression, and pro-inflammatory cytokine IL-6 and IL-6 levels in MDA-MB-231 cells. Importantly, restoring the TLR4 pathway overturned hyperoside-evoked chemosensitivity to paclitaxel in MDA-MB-231 cells. Thus, hyperoside may elevate breast cancer cell sensitivity to paclitaxel by blocking TLR4 activation-mediated pro-inflammatory and pro-survival approaches, thereby endorsing its usefulness as a promising therapeutic combination to overcome chemosensitivity in breast cancer.     

TMPRSS4通过上皮间质转化促进乳腺癌细胞的增殖、侵袭和转移

目的通过检测Ⅱ型跨膜丝氨酸蛋白酶4（TMPRSS4）在乳腺癌细胞中的表达情况,分析其在乳腺癌细胞增殖、侵袭和转移过程中的作用及其与上皮间质转化（EMT）的关系。 方法采用实时荧光定量PCR（qRT-PCR）和Western blot法检测5种不同乳腺癌细胞系中TMPRSS4 mRNA和蛋白水平的表达情况。将过表达质粒转染至乳腺癌细胞中,采用qRT-PCR方法检测转染效率,并通过MTS和EdU细胞增殖实验、Transwell和Matrigel细胞迁移和侵袭实验,研究过表达TMPRSS4对乳腺癌细胞增殖、侵袭和迁移的作用。采用qRT-PCR和Western blot法检测过表达TMPRSS4后细胞EMT相关基因E-cadherin、Vimentin、Claudin-1、Slug、ZEB1的表达变化。 结果TMPRSS4在乳腺癌MDA-MB-468和MDA-MB-231细胞系中高表达。MTS实验显示TMPRSS4过表达组乳腺癌细胞MDA-MB-468和MDA-MB-231的增殖能力与阴性对照组相比明显增加,差异具有统计学意义（P=0.039和0.038）,EdU实验进一步证实过表达TMPRSS4促进乳腺癌细胞的增殖,MDA-MB-468细胞和MDA-MB-231细胞的增殖率与阴性对照组比较差异具有统计学意义（P=0.001和0.008）。Transwell实验显示,TMPRSS4过表达组MDA-MB-468细胞和MDA-MB-231细胞的迁移能力与阴性对照组比较明显提高,差异具有统计学意义（p均=0.001）。Matrigel实验显示,TMPRSS4过表达组MDA-MB-468细胞和MDA-MB-231细胞的侵袭浸润能力与阴性对照组比较明显提高,差异具有统计学意义（P=0.012和0.000）。与阴性对照组比较,过表达TMPRSS4后,EMT相关基因包括上皮性标记E-cadherin和Claudin-1的表达均显著下降,而间质标记Vimentin和Slug的表达均明显提高,差异具有统计学意义（P分别=0.024,0.003,0.002和0.012）。 结论TMPRSS4参与了乳腺癌EMT的发生过程,并通过EMT促进乳腺癌细胞的增殖、侵袭和转移。     

Do wine polyphenols modulate p53 gene expression in human cancer cell lines?

BACKGROUND: The p53 gene is an established tumor suppressor and an inducer of apoptosis. We here attempt to determine whether the putative anticarcinogenic properties attributed to red wine and its polyphenolic constituents depend, at least in part, upon their ability to modulate p53 expression in cancer cells. METHODS: Three human breast cancer cell lines (MCF-7, T47D; MDA-MB-486) and one human colon cancer cell line [Colo 320 HSR (+)] were treated for 24-h with each of four polyphenols [quercetin; (+)-catechin, trans-resveratrol; caffeic acid] at concentrations ranging from 10(-7) M to 10(-4) M, after which, p53 concentrations were measured in cell lysates by a time-resolved fluorescence immunoassay. RESULTS: None of the polyphenols tested affected p53 expression in the breast cancer cell lines T-47D and MDA-MB-486. p53 content of MCF-7 breast cancer cells (wild-type) was increased by caffeic acid, decreased by resveratrol, and showed a twofold increase with catechin, that reached borderline statistical significance; however, none of these effects were dose-responsive. Colo 320 HSR (+) cells (with a mutant p53 gene) had lower p53 content upon stimulation, reaching borderline statistical significance, but without being dose-responsive, in the presence of caffeic acid and resveratrol. Apart from toxicity at 10(-4) M, quercetin had no effect upon these four cell lines. CONCLUSIONS: The observed p53 concentration changes upon stimulation by polyphenols are relatively small, do not follow a uniform pattern in the four cell lines tested, and do not exhibit a dose-response effect. For these reasons, we speculate that the putative anticarcinogenic properties of wine polyphenols are unlikely to be mediated by modulation of p53 gene expression.     

Benzyl isothiocyanate-induced apoptosis in human breast cancer cells is initiated by reactive oxygen species and regulated by Bax and Bak

Epidemiologic studies have revealed an inverse correlation between dietary intake of cruciferous vegetables and the risk of breast cancer. We now show that cruciferous vegetable constituent benzyl isothiocyanate (BITC) effectively suppresses growth of cultured human breast cancer cells (MDA-MB-231 and MCF-7) by causing G(2)-M phase cell cycle arrest and apoptosis induction. On the other hand, a normal mammary epithelial cell line (MCF-10A) is significantly more resistant to growth arrest and apoptosis by BITC compared with breast cancer cells. The BITC-mediated cell cycle arrest was associated with a decrease in levels of proteins involved in regulation of G(2)-M transition, including cyclin B1, cyclin-dependent kinase 1, and cell division cycle 25C. The BITC-induced apoptosis correlated with induction of proapoptotic proteins Bax (MCF-7) and Bak (MDA-MB-231 and MCF-7) and down-regulation of antiapoptotic proteins Bcl-2 and Bcl-xL (MDA-MB-231). The SV40-immortalized mouse embryonic fibroblasts derived from Bax and Bak double knockout mice were significantly more resistant to BITC-induced DNA fragmentation compared with wild-type mouse embryonic fibroblasts. The BITC treatment caused rapid disruption of the mitochondrial membrane potential, leading to cytosolic release of apoptogenic molecules, which was accompanied by formation of autophagosome-like structures as revealed by transmission electron microscopy. The BITC-mediated apoptosis was associated with generation of reactive oxygen species and cleavage of caspase-9, caspase-8, and caspase-3. Apoptosis induction by BITC was significantly attenuated in the presence of a combined superoxide dismutase and catalase mimetic EUK134 as well as caspase inhibitors. In conclusion, the present study reveals a complex signaling leading to growth arrest and apoptosis induction by BITC.     

Identification of magnetic resonance detectable metabolic changes associated with inhibition of phosphoinositide 3-kinase signaling in human breast cancer cells

Phosphoinositide 3-kinase (PI3K) is an attractive target for novel mechanism-based anticancer treatment. We used magnetic resonance (MR) spectroscopy (MRS) to detect biomarkers of PI3K signaling inhibition in human breast cancer cells. MDA-MB-231, MCF-7, and Hs578T cells were treated with the prototype PI3K inhibitor LY294002, and the (31)P MR spectra of cell extracts were monitored. In every case, LY294002 treatment was associated with a significant decrease in phosphocholine levels by up to 2-fold (P < 0.05). In addition, a significant increase in glycerophosphocholine levels by up to 5-fold was also observed (P 相似文献