姜黄素诱导乳腺癌MCF-7细胞凋亡

目的 研究姜黄素对人乳腺癌细胞株MCF-7细胞增殖抑制和诱导凋亡作用.方法 MTT法检测姜黄素对MCF-7细胞的增殖抑制作用;流式细胞术(FCM)PI单染检测细胞周期;Annexin V/PI双染法检测细胞凋亡;Western blot法检测Bcl-2和Bax蛋白的表达.结果 姜黄素对MCF-7细胞生长有明显抑制作用,并呈剂量、时间依赖性;姜黄素能使MCF-7细胞阻滞在G1/S期,可以诱导细胞凋亡,Bax蛋白表达上调,而Bcl-2的表达减少.结论 姜黄素对人乳腺癌MCF-7细胞的增殖具有显著的抑制作用并可诱导细胞凋亡.其分子作用机制可能与其上调Bax基因表达水平的同时下调Bcl-2基因表达水平,从而诱导细胞凋亡有关.     

Induction of apoptosis and autophagy by sodium selenite in A549 human lung carcinoma cells through generation of reactive oxygen species

Selenium in the form of sodium selenite has been reported to exert anti-tumor effects in several cancer cell types by inducing autophagic cell death and apoptosis mediated by reactive oxygen species (ROS). However, the exact molecular pathways underlying these effects have not been fully established. The present study used A549 human lung carcinoma cells for further investigation of the anti-cancer mechanism of sodium selenite. We showed that sodium selenite modulated both the extrinsic and intrinsic apoptotic pathways, which were interconnected by Bid truncation. We used z-VAD-fmk, a pan-caspase inhibitor, to demonstrate that sodium selenite-induced apoptosis was dependent on the activation of caspases. Sodium selenite also increased autophagy, as indicated by an increase in microtubule-associated protein light chain-3 (LC3) puncta, accumulation of LC3II, and elevation of autophagic flux. Pretreatment with bafilomycin A1 enhanced sodium selenite-induced apoptosis, indicating that sodium selenite-induced autophagy functioned as a survival mechanism. Sodium selenite treatment also resulted in generation of ROS, which abrogated mitochondrial membrane potential (MMP) and regulated both apoptosis and autophagy. Phospho-nuclear factor erythroid 2-related factor 2 (p-Nrf2) showed a ROS-dependent translocation to the nucleus, which suggested that Nrf2 might increase cell survival by suppressing ROS accumulation and apoptosis mediated by oxidative stress. Sodium selenite treatment of A549 cells therefore appeared to trigger both apoptosis and cytoprotective autophagy, which were both mediated by ROS. The data suggest that regulation of ROS generation and autophagy can be a potential strategy for treating lung cancer that is resistant to pro-apoptotic therapeutics.     

氧化苦参碱诱导人乳腺癌细胞MCF-7凋亡的实验研究

目的　研究氧化苦参碱对MCF 7细胞的诱导凋亡作用机制。方法　用光学显微镜、电子显微镜、激光共聚焦显微镜、流式细胞仪和DNA凝胶电泳等技术观察细胞凋亡。结果　实验显示氧化苦参碱作用于体外培养的MCF 7细胞可诱导发生凋亡 ,凋亡细胞表现为细胞固缩 ,核染色质聚集或碎裂、胞质空泡化等 ;DNA电泳可见DNA梯形条带 ;激光共聚焦显微镜示DNA含量下降 ,流式细胞仪检测sub G1峰在G1期前出现 ,S期细胞比例增高。结论　氧化苦参碱对体外培养MCF 7细胞生长有抑制作用 ,机制与通过阻止细胞周期的进程 ,启动细胞自身调控程序 ,诱导肿瘤细胞凋亡有关     

Metallothionein induction by sodium selenite at two different ambient temperatures in mice

The induction of metallothionein (MT) synthesis by sodium selenite was investigated in mice with regard to the hypothermic response known to be caused by sodium selenite. Mice received a subcutaneous injection of sodium selenite at two doses (20 and 45 mol/kg) under two ambient temperature (22 and 33° C) conditions. Hepatic MT concentration was significantly increased by an injection of sodium selenite compared to the control, whereas no significant effect of ambient temperature was observed. The distribution of radiolabeled selenium was examined in vivo and in vitro. When sodium selenite was injected into mice, radiolabeled selenium was mostly eluted in a fraction larger in molecular weight than MT and was not found in a fraction corresponding to MT. When sodium selenite was added to the hepatic supernatant of the mice that had been injected with zinc sulfate, zinc in zinc-thionein was not displaced by radiolabeled selenium.     

沙棘油和亚硒酸钠对汞诱导的大鼠肝肾氧化损伤的影响

目的通过汞诱导大鼠肝肾氧化损伤,观察沙棘油(Sea Buckthorn Oil,SBO)和亚硒酸钠(Na2SeO3)干预效应。方法Wister大鼠随机分成对照组、单纯染汞组、SBO和Na2SeO3处理组。测定肝、肾及尿中汞含量,肝、肾中GSH、MDA、蛋白含量和SOD、GSH-PX活力。结果与单纯染汞组相比,SBO处理组尿汞含量增加(P<0.05),肝GSH-PX活力升高(P<0.01);Na2SeO3处理组肝汞增加,肾和尿汞含量下降,肝GSH-PX活力、GSH含量均升高(P<0.01),肝SOD活力和肾GSH-PX活力均增加(P<0.05),肾MDA含量下降(P<0.05)。结论沙棘油具有促进汞从肾脏排出的作用,对汞诱导的肝氧化损伤具有一定保护作用,对肾氧化损伤未表现出明显的保护作用。Na2SeO3可有效拮抗汞诱导的肝肾氧化损伤作用。     

N-乙酰半胱氨酸和亚硒酸钠对镉亚慢性毒性的影响

目的探讨N-乙酰半胱氨酸(N-acetyl cysteine,NAC)和亚硒酸钠(Na2SeO3)对亚慢性染镉大鼠肝肾毒性的影响及其机制。方法32只Wistart大鼠随机分为4组,每组8只,第1组为对照组,第2组为单位染镉组,第3、4组为干预组。大鼠连续6周皮下注射7μmol/kg氯化镉,然后干预组分别腹腔注射1 mmol/kg NAC和10μmol/kg Na2SeO3,共2周;测定大鼠尿N-乙酰-β-苷酶(NAG)、碱性磷酸酶活力(ALP)和肝、肾皮质谷胱甘肽(GSH)、丙二醛(MDA)含量及谷胱甘肽过氧化物酶(GSH-Px)活力。结果亚慢性染镉使大鼠肝、肾皮质和尿镉含量显著升高,尿ALP、NAG和蛋白含量显著升高,肝、肾皮质GSH含量显著升高,GSH-Px活力显著降低。与单纯染镉组比较,NAC处理组尿镉、NAG和蛋白含量显著下降,肝、肾皮质GSH显著降低;Na2SeO3处理组尿镉、ALP及肝、肾皮质GSH含量显著下降,GSH-Px活力显著升高。结论NAC和Na2SeO3对镉致肾损伤的恢复具有促进作用,其机制可能与NAC或Na2SeO3改变体内GSH含量和GSH-Px活力有关。     

苦参碱对人乳腺癌细胞MCF-7增殖、凋亡的影响

目的探讨苦参碱对人乳腺癌细胞MCF-7的增殖、凋亡的影响。方法实验组MCF-7细胞加入苦参碱溶液,对照组加入等量培养液。MTT比色法检测苦参碱对MCF-7细胞的生长抑制率;镜下观察细胞形态学变化;流式细胞术检测MCF-7细胞Bax、Bcl-2蛋白表达。结果与对照组比较,实验组苦参碱呈时间、剂量依赖性明显抑制MCF-7细胞生长,诱导MCF-7细胞凋亡,上调Bax蛋白表达,下调Bcl-2蛋白表达。结论苦参碱对MCF-7细胞具有明显的生长抑制和促凋亡作用,与上调Bax和下调Bcl-2表达有关。     

Different effects of adenylyl cyclase activators and phosphodiesterases inhibitors on cervical cancer (HeLa) and breast cancer (MCF-7) cells proliferation

Breast and cervical cancers are the most common cancers in Iran and worldwide. Hormonal stimulation of cyclic adenosine mono phosphate (cAMP) and the cAMP-dependent protein kinase PKA regulates cell growth by different mechanism. cAMP can stimulate cell growth in many cell types while inhibiting cell growth in others. In some cell lines have been shown that the proliferation of tumor cells is reduced by increasing cAMP in cells. In this study, we evaluate growth arrest of selective PDE3 and non-selective PDE inhibitors, which lead to increase level of cAMP in cervical (HeLa) and breast cancer (MCF7) cell lines have been studied. Cells were incubated with different concentrations of selective, non-selective PDE inhibitors, beta adrenergic receptor agonist and direct stimulator of adenylyl cyclase. Cell viability was quantitated by MTT assay. Apoptotic cells were determined using PI staining of DNA fragmentation by flow cytometry (sub-G1 peak). Result showed that selective PDE inhibitors decreased cell viability in HeLa and MCF-7 cells as a time-dependent manner. Non-selective inhibitor and beta-adrenergic receptor agonist also decrease cell viability but they are less powerful than selective PDE3 inhibitors. Forskolin had no effect in viability of cells. Analysis of DNA fragmentation by flow cytometry showed apoptosis involved in selective PDE3 inhibitors induced toxicity in HeLa cell. Thus, the growth inhibitory effects of selective PDE3 inhibitors are more effective than non-selective inhibitor. Further studies are needed to investigate the mechanism of action is on the field.     

Proteomic analysis of prodigiosin-induced apoptosis in a breast cancer mitoxantrone-resistant (MCF-7 MR) cell line

Summary Prodigiosin (PG) is a bacterial, red-pigmented antibiotic with immunosuppressive and apoptotic activities. To better understand its mechanisms of action, we tried to identify proteins associated with apoptosis induced by PG. For this purpose, the variation of protein expression on exposure to apoptotic concentrations of PG was examined, by high-resolution two-dimensional gel electrophoresis (2D-E), in the MCF-7 cancer cell line resistant to mitoxantrone (MCF-7-MR). Six PG apoptosis-associated protein spots were further characterized by complementary peptide mass fingerprinting and tandem mass spectrometry data obtained on a matrix-assisted laser desorption ionization-time-of-flight/time-of-flight (MALDI-TOF/TOF) mass spectrometer. The proteins identified were involved in various cellular functions, including cell defence, DNA repair and cellular organization. Our data provide novel information on cell response to PG, a new apoptotic drug with interesting anticancer activity.     

Evaluation of chemopreventive and cytotoxic effect of lemon seed extracts on human breast cancer (MCF-7) cells

Extracts from lemon seed were investigated for the radical scavenging activity and apoptotic effects in human breast adenocarcinoma (MCF-7) cells and non-malignant breast (MCF-12F) cells for the first time. Defatted seed powder was successively extracted with ethyl acetate (EtOAc), acetone, methanol (MeOH), and MeOH:water (80:20). The chemical constituents were identified and quantified by LC-MS and HPLC analysis, respectively. The highest radical scavenging activity of 62.2% and 91.3% was exhibited by MeOH:water (80:20) at 833μg/mL in 1,1-diphenyl-2-picryl hydrazyl (DPPH) and 2,2'-azino-di-(3-ethylbenzothiazoline)-6-sulfonic acid (ABTS(+)), respectively. In addition, the MeOH:water (80:20) extract showed the highest (29.1%, P<0.01) inhibition of MCF-7 cells in MTT assay. Treatment of the MeOH:water (80:20) extract induced DNA fragmentation and poly(ADP-ribose) polymerase (PARP) cleavage. Increased levels of Bax and cytosolic cytochrome C and decreased levels of Bcl2 were also observed in MeOH:water (80:20) treated MCF-7 cells. In conclusion, the MeOH:water (80:20) extract from lemon seed has potent antioxidant activity and induces apoptosis in MCF-7 cells, leading to the inhibition of proliferation. These results suggest that aglycones and glucosides of the limonoids and flavonoid present in MeOH:water (80:20) extract may potentially serve as a chemopreventive agent for breast cancer.     

Differential anti-proliferative actions of peroxisome proliferator-activated receptor-gamma agonists in MCF-7 breast cancer cells

Peroxisome proliferator-activated receptor-gamma (PPARgamma) activation has been a new approach to cancer therapy. In the present study, we investigated the effects of two structurally different PPARgamma agonists, rosiglitazone and KR-62980 on MCF-7 breast cancer cells. Both agonists inhibited the cell proliferation and colony formation via apoptosis. PTEN expression was increased with decreased Akt phosphorylation by the agonists, whereas agonists actions were abolished in PTEN knockdown cells, indicating the critical role of PTEN in the anti-proliferative effects of PPARgamma activation. Rosiglitazone induced the MCF-7 cell differentiation but KR-62980 did not alter the differentiation pattern with little effects on the lipid accumulation and the expression of lipogenesis markers. These results suggest that PPARgamma activation may result in the inhibition of cell proliferation and/or induction of cell differentiation depending on the type of PPARgamma agonists, and that KR-62980 may be useful in breast cancer therapy by inducing apoptosis.     

Chimaphilin induces apoptosis in human breast cancer MCF-7 cells through a ROS-mediated mitochondrial pathway

Chimaphilin, 2,7-dimethyl-1,4-naphthoquinone, is extracted from pyrola [Passiflora incarnata Fisch.]. In this study, the anticancer activity and underlying mechanisms of chimaphilin toward human breast cancer MCF-7 cells are firstly investigated. Chimaphilin could inhibit the viability of MCF-7 cells in a concentration-dependent manner, and the IC₅₀ value was 43.30 μM for 24 h. Chimaphilin markedly induced apoptosis through the investigation of characteristic apoptotic morphological changes, nuclear DNA fragmentation, annexin V-FITC/propidium iodide (PI) double staining. Flow cytometry assay revealed that chimaphilin triggered a significant generation of ROS and disruption of mitochondrial membrane potential. Additionally, western blotting assay showed that chimaphilin suppressed Bcl-2 level and enhanced Bad level, then activated caspase-9 and caspase-3, and further activated the poly ADP-ribose polymerase (PARP), finally induced cell apoptosis involving the mitochondrial pathway. Furthermore, free radical scavengers N-acetyl-L-cysteine (NAC) pretreatment test testified that chimaphilin could increase the generation of ROS, then induce cell apoptosis. In general, the present results demonstrated that chimaphilin induced apoptosis in human breast cancer MCF-7 cells via a ROS-mediated mitochondrial pathway.     

N-乙酰半胱氨酸预防亚硒酸钠诱导大鼠白内障的研究

目的 对亚硒酸钠诱导白内障大鼠采用N-乙酰半胱氨酸进行预防性治疗,探讨N-乙酰半胱氨酸对白内障大鼠模型氧化损伤和晶状体浑浊程度的影响。方法 30只SD乳鼠随机分为3组,模型组ip 3.46 mg/kg亚硒酸钠溶液造模后,ip 0.1 mL/10 g生理盐水;实验组ip N-乙酰半胱氨酸10 mg/g,30 min ip亚硒酸钠溶液造模;对照组ip 0.1 mL/10 g生理盐水。所有动物均为隔日注射,连续6次。比较大鼠晶状体浑浊程度和SOD活性、NOS活性和MDA水平。结果 在晶状体核区、后囊区、前囊区+后囊区、前囊区+皮质区+核区+后囊区中,实验组与模型组比较得分差异具有统计学意义（P<0.05）;实验组SOD活性和MDA活性与模型组比较差异均具有统计学意义（P<0.05）。结论 N-乙酰半胱氨酸能够预防后囊和核区白内障的发生,其作用是通过减少MDA水平、提高SOD活性、改善氧化应激实现的。     

SAHA-CTSV轴通过诱导过度自噬抑制乳腺癌MCF-7细胞的生长

目的研究SAHA-CTSV轴通过诱导过度自噬抑制乳腺癌MCF-7细胞生长的作用和机制。方法以人乳腺癌MCF-7细胞为研究目标,Muse细胞分析仪检测SAHA对细胞活力的作用;Real-time PCR和Western blot检测SAHA对CTSV的mRNA及蛋白表达的影响,同时检测利用siRNA干扰技术敲除CTSV后的细胞CTSV表达水平;细胞免疫荧光法检测SAHA对LC3II分子的作用;Real-time PCR和Western blot检测SAHA诱导自噬过程中相关ATG分子和LC3的mRNA和蛋白的变化;Bafilomycin A1抑制自噬,随后利用Western blot和Muse细胞分析仪分别检测p62蛋白水平和SAHA作用后的细胞活力。结果SAHA可以抑制MCF-7细胞的增殖,并促进CTSV的表达。CTSV-siRNA转染细胞后,CTSV的mRNA和蛋白水平均显著性降低。SAHA增加了LC3免疫荧光点的分布,与CTSV-siRNA共同作用可恢复由CTSV-siRNA敲减引起的细胞自噬减弱。SAHA在增强ATG4C和Beclin1表达的同时,LC3B也随之升高而mTOR则下降。抑制CTSV表达后也同时逆转了SAHA的效应。Bafilomycin A1阻断自噬后,被SAHA抑制的p62蛋白的表达和细胞活力显著增强。结论SAHA-CTSV轴通过诱导过度自噬抑制乳腺癌MCF-7细胞的生长。     

藻红蛋白诱导人乳腺癌 MCF-7 细胞衰老研究

目的：研究藻红蛋白对人乳腺癌 MCF-7 细胞的诱导衰老作用。方法：采用 MTT 法检测藻红蛋白对 MCF-7 细胞的生长抑制作用,采用倒置显微镜观察形态学变化,酸性 β-半乳糖苷酶染色法和衰老相关异染色质聚集检测藻红蛋白诱导人乳腺癌 MCF-7 细胞衰老作用。结果：MTT 法结果显示,藻红蛋白对体外培养的人乳腺癌细胞 MCF-7 具有明显的生长抑制作用,IC₅₀ 为 134.87 μg/mL。结果发现藻红蛋白作用 72 h 后,细胞呈现变大变扁平,细胞内形成空泡、颗粒增多等明显衰老形态。在倒置显微镜下观察,酸性 β-半乳糖苷酶染色呈阳性;在荧光显微镜下观察,DAPI 染色出现衰老相关异染色质聚集现象。结论：藻红蛋白可通过诱导 MCF-7 细胞衰老达到抗肿瘤作用。     

神经酰胺诱导人结肠癌细胞凋亡作用

目的探讨外源性神经酰胺诱导人结肠癌HT-29细胞的凋亡作用。方法采用光镜、电镜、荧光显微镜和琼脂糖凝胶电泳方法检测C2-神经酰胺诱导HT-29细胞凋亡。MTT法检测C2-神经酰胺对HT-29细胞线粒体功能的影响。结果C2-神经酰胺使HT-29细胞发生核染色质断裂、DNA Ladder、凋亡小体等典型凋亡表现,12和24 h凋亡细胞率分别为64.1%和81.3%,呈现时间-剂量依赖关系。同时C2-神经酰胺处理细胞6 h后,细胞线粒体功能即出现损伤。结论外源性神经酰胺能诱导人结肠癌HT-29细胞凋亡,参与其凋亡调控。     

Scorpion venom (Odontobuthus doriae) induces apoptosis by depolarization of mitochondria and reduces S-phase population in human breast cancer cells (MCF-7)

Venom of some species of scorpions induces apoptosis and arrests proliferation in cancer cells. This is an important property that can be harnessed and can lead to isolation of compounds of therapeutic importance in cancer research. Cytotoxicity was investigated using MTT reduction and confirmed with lactate dehydrogenase release following venom exposure. Apoptosis was evaluated with determination of mitochondrial membrane potential, reactive nitrogen species assay, measurement of Caspase-3 activity and DNA fragmentation analysis. To confirm that venom can inhibit DNA synthesis in proliferating breast cancer cells, immunocytochemical detection of BrdU incorporation was done. Our results demonstrated that venom of Odontobuthus doriae not only induced apoptosis but lead to the inhibition of DNA synthesis in human breast cancer cells (MCF-7). Cell viability decreased with parallel increment of LDH release in dose dependent manner after treatment with varying concentrations of venom. Moreover, venom depleted cellular antioxidants evidenced by depression of GSH and Catalases and concomitantly increased reactive nitrogen intermediates (RNI). These events were related to the depolarization of mitochondria and associated Caspase-3 activation following venom treatment in a concentration dependent manner. Finally, fragmentation of nuclear DNA following venom treatment confirmed the apoptotic property of the said venom. These results suggest that venom of O. doriae can be potential source for the isolation of effective anti-proliferative and apoptotic molecules.     

染料木素在乳腺癌MCF-7细胞内的代谢研究

目的研究染料木素在乳腺癌细胞中的代谢行为。方法将10μmol.L-1染料木素与乳腺癌MCF-7细胞在37℃下孵化一定时间后终止反应。采用UPLC-MS-MS法测定孵化液中染料木素及II相代谢产物的浓度。结果染料木素在乳腺癌MCF-7细胞中可迅速代谢为7或4′-葡萄糖醛酸结合物以及7或4′-硫酸结合物。孵化24、48和72 h后,每100μg孵化样品蛋白中,7-葡萄糖醛酸结合物的生成率分别为2.9、5.2和8.9μg.h-1,4′-葡萄糖醛酸结合物的生成率分别为1.1、1.9和3.0μg.h-1,7-硫酸结合物的生成率分别为20.3、32.9和47.3μg.h-1,4′-硫酸结合物的生成率分别为1.7、3.1和4.5μg.h-1。结论染料木素在乳腺癌MCF-7细胞中的主要代谢产物是7-硫酸结合物。     

Growth inhibition and induction of apoptosis in MCF-7 breast cancer cells by a new series of substituted-1,3,4-oxadiazole derivatives

The multiple pharmacological actions of unique synthetic compounds are a prerequisite for classifying a drug as highly efficacious, because the multiple pharmacological actions offer the possibility of treating various diseases like cancer. 1,3,4-Oxadiazoles are an important class of heterocyclic compounds with broad spectrum of biological activities. In this study we focused on the ability of these derivatives to induce apoptosis in cultured MCF-7 breast cancer cells. Treatment of MCF-7 cells with varying concentrations of the different derivatives resulted in dose and time dependent sequence of events marked by apoptosis, as shown by loss of cell viability, chromatin condensation, internucleosomal DNA fragmentation and sub G(0) phase accumulation. Furthermore, apoptosis in MCF-7 cell was induced by upregulation of proto-oncoprotein Bax and activation of Caspase-3 activated DNase. Although the derivatives induced apoptosis was associated with Bax protein levels, negligible Bcl-2 reduction was observed. Analysis of the data suggests that the substituted oxadiazole derivatives exert antiproliferative action and growth inhibition on MCF-7 cells through apoptosis induction and that it may have anticancer properties valuable for application in drug products.