Quercetin inhibits the growth of a multidrug-resistant estrogen-receptor-negative MCF-7 human breast-cancer cell line expressing type II estrogen-binding sites

Summary It has been demonstrated that the flavonoid quercetin (3,3,4,5,7-pentahydroxyflavone; Q) inhibits the growth of several cancer cell lines. There is evidence suggesting that the antiproliferative activity of this substance is mediated by the so-called type II estrogen-binding, site (type II EBS). We looked for the presence of type II EBS and the effect of Q on the proliferation of an Adriamycinresistant estrogen-receptor-negative human breast-cancer cell line (MCF-7 ADRr). By whole-cell assay using estradiol labelled with 6,7-tritium ([³H]-E2) as a tracer, we demonstrated that MCF-7 ADRr cells contain type II EBSs. Competition analysis revealed that diethylstilbestrol (DES) and Q competed with similar potency for [³H]-Es binding to type II EBSs. The antiestrogen tamoxifen (TAM) competed for type II EBSs, albeit to a lesser extent than either DES or Q. Growth experiments demonstrated that Q and DES exerted a dose-dependent inhibition of cell proliferation in the range of concentrations between 10 nM and 10 m, whereas TAM was less effective. Q could also inhibit colony formation in a clonogenic assay. Our results indicate that multidrug-resistant estrogen-receptor-negative MCF-7 cells express, type II EBSs and are sensitive to the inhibitory effect of Q. This substance could be the parent compound of a novel class of anticancer agents.     

黄芩素对卵巢癌耐药细胞株A2780/ADM逆转作用实验研究

目的探讨黄芩素对卵巢癌耐药细胞株A2780/ADM耐药逆转作用及机制.方法MTT法观察药物对细胞的生长抑制作用;免疫组化法检测细胞P-糖蛋白(P-Gp)和多药耐药相关蛋白(MRP)的表达;采用高效液相色谱法(HPLC)测定细胞内ADM的浓度.结果卵巢癌耐药细胞株A2780/ADM对多种化疗药物产生多药耐药性,耐药细胞P-Gp和MRP的阳性率显著高于亲本细胞(P＜0.01);黄芩素在非细胞毒剂量时与ADM合用后能逆转A2780/ADM对ADM的耐药性;ADM与黄芩素合用时,其细胞内ADM的含量较单独使用明显增高.结论卵巢癌耐药细胞株A2780/ADM对多种化疗药物具有多药耐药性,其多药耐药性与P-Gp和MRP过量表达有关;黄芩素在非细胞毒剂量时能逆转A2780/ADM对ADM的耐药性,其逆转作用可能与降低P-Gp药物外排功能、增加细胞内药物浓度有关.     

免疫效应细胞促进阿霉素诱导乳腺癌耐药细胞凋亡

目的 探讨免疫效应细胞促进阿霉素（ADR）诱导乳腺癌耐药细胞MCF7／ADR凋亡的作用机制。方法 采用干扰素-γ（IFN-γ）、CD3单抗、白介素（IL）-1和IL-2诱导并扩增免疫效应细胞。利用末端脱氧核苷酸转移酶介导的dUTP缺口末端标记（TUNEL）技术和P-糖蛋白（P-gP）免疫组织化学染色,观察P-gP表达与凋亡的关系。用流式细胞术检测乳腺癌相关抗原P185蛋白的表达,以及Annexin V-FITC／PI阳性率。荧光显微镜下观察ADR在靶细胞内的分布和Annexin V的表达。结果 免疫效应细胞使MCF7／ADR细胞P185和P-gP表达明显下降,CIK细胞作用后,MCF7／ADR细胞P185标记荧光几何均数下降了91．9％。免疫效应细胞增加了ADR在MCF7／ADR细胞内的积累及其在细胞核的分布。联合应用免疫效应细胞和ADR后,MCF7／ADR细胞凋亡率达78．9％,比单纯加入ADR组增高了10倍,较单纯加入免疫效应细胞组增高了13倍。结论 免疫效应细胞可同时下调P185蛋白和P-gp蛋白的表达,协同ADR可提高MCF7／ADR细胞的凋亡率。     

乳腺癌阿霉素耐药细胞株MCF-7/ADM的上皮间质化现象及机制

目的:建立乳腺癌阿霉素耐药细胞株,命名为MCF-7/ADM,探讨耐药细胞的上皮间质化(EMT)现象及机制,为耐药乳腺癌的治疗奠定基础.方法:中等浓度间歇法建立乳腺癌阿霉素耐药细胞株MCF-7/ADM.采用MTT和Transwell实验分别检测耐药对细胞增殖、迁移与转移能力的影响;荧光定量PCR检测MCF-7/ADM细胞EMT相关分子标志物E-钙黏蛋白及间质细胞分子标记N-钙黏蛋白、波形蛋白、纤维连接蛋白等的表达.ELISA检测TGF-β的表达,Western blot检测TGF-β及Smad2/3在蛋白水平的表达.结果:乳腺癌阿霉素耐药细胞MCF-7/ADM的耐药指数为32.2,对5-氟脲嘧啶、顺铂、环磷酰胺产生交叉耐药性.与MCF-7细胞相比,MCF-7/ADM细胞迁移和转移能力明显增强,E-cadherin表达降低而N-cadher-in表达显著增高,细胞上清中TGF-β1表达增加,细胞内TGF-β及磷酸化Smad2/3在蛋白水平表达升高.结论:乳腺癌阿霉素耐药细胞MCF-7/ADM发生EMT现象,可能与经典TGF-β通路激活有关.     

电化学治疗对具多药耐药表型的人乳腺癌细胞MCF-7adr的作用

目的：研究电化学治疗（ＥＣＴ）对具多药耐药表型的人乳腺癌细胞ＭＣＦ－７＾ａｄｒ的作用。方法：以敏感型人乳腺细胞ＭＣＦ－７为对照,采用细胞计数法观察ＥＣＴ对乳腺癌耐药细胞株ＭＣＦ－７＾ａｄｒ的抑制效应,以ＡｎｎｅｘｉｎＶ标记法观察ＥＣＴ的诱导ＭＣＦ－７＾ａｄｒ细胞凋亡的作用。结果：ＥＣＴ对耐药细胞ＭＣＦ－７＾ａｄｒ的诱导凋亡及抑制效应接近于其对敏感细胞ＭＣＦ－７的作用,无显著性差异（Ｐ〉０．０５）。     

In vitro andin vivo modulation of growth regulation in the human breast cancer cell line MCF-7 by estradiol metabolites

Background  The natural estrogen 17β-estradiol (E₂) functions as a potent tumor promoter during tumorigenic transformation of the mammary gland. From amongst the various pathways of E₂ metabolism upregulation of C16α-hydroxylation of E₂ has been associated with carcinogenesis. In the present studyin vitro andin vivo experiments were performed on estrogen receptor positive human breast cancer MCF-7 cells to examine whether the natural estrogen E₂ and its metabolites 16α-hydroxyestrone (16α-OHEi) and 2-hydroxyestrone (2-OHE₁) function as modulators of tumor cell growth. Methods  An anchorage-independent growth assay was used forin vitro study by counting the number of tri-dimensional colonies formed by MCF-7 cells suspended in 0.33% agar.In vivo experiments examined the effect of implanting metabolite material pellets into female nude mice. Results  In the anchorage-independent growth assay (AIG), continuous 14-day exposure to E₂ and to 16α-OHE₁ at 200 ng/ml induced a 59.4% and a 105.9% increase (P=0.001) respectively in the number of colonies of MCF-7 cells. Identical treatment with 2-OHE₁, however, failed to increase AIG relative to that seen in the solvent treated control cultures. In thein vivo tumorigenicity assay, treatment of nude mice with 1.5 mg E₂ or 16α-OHE₁ resulted in a 335.4% and a 384.1% increase (P<0.0002) in tumor growth, while identical treatment with 2-OHE₁ failed to exhibit any increase relative to the control group. Conclusions  These results suggest that the 16α- and 2-hydroxylated metabolites of E₂ may directly affectin vitro growth of MCF-7 cells via an autocrine mechanism andin vivo growth via paracrine mechanisms. Thus, E2-mediated growth regulation in MCF-7 cells may in part be due to distinct effects of specific E₂ metabolites on the breast cancer cells.     

Regulation of expression of P-glycoprotein and gst by multidrug-resistant reversors in adm-sensitive and adm-resistant human tumor cell lines

Threemultidrugresisors,verapamil(VER),dipyriamole(DPM),andcyclosporina(CsA),havebeenfoundtoovercomemultidrugresistanceinvarioustumorcellsmainlythroughcompetivecombinationwithanticancerdrugsforp-glycoprotein(PgP)fromoursandotherinvestigatorsfindings..d.li-3]Wereportedpreviouslytheregulativeeffectofthesereversorsonsevenkindsofoncoproteinsortumorsuppressoroncoproteinsexpressions.fWereportheretheregulationofPgPandGSTexpressionfromthesereversorsinADM-sensitiveandADMresistanthumanleukemicce…     

槲皮素对多药耐药细胞株KB-MRP的耐药逆转研究

目的:研究槲皮素(quercetin)对多药耐药细胞株KB-MRP的耐药逆转作用。方法:使用四甲基偶氮唑盐(MTT)法检测不同浓度槲皮素对细胞株KB-MRP的抑制率,确定其非细胞毒性浓度。用MTT法分别检测阿霉素(ADM)、依立替康(CPT-11)、长春新碱(VCR)、环磷酰胺(CTX)对细胞株KB-3-1、KB-MRP以及使用槲皮素处理后的KB-MRP的半数抑制浓度(IC50)。通过流式细胞术检测KB-3-1、加入槲皮素前后KB-MRP细胞内的化疗药物阿霉素(ADM)的荧光强度以及加入槲皮素前后多药耐药细胞株KB-MRP的MRP1蛋白表达。结果:终浓度为0~40μmol/L的槲皮素对KB-MRP无明显细胞毒作用。KB-MRP对ADM、VCR、CPT-11均有不同程度的耐药但对CTX不耐药。使用20μmol/L槲皮素处理后KB-MRP对上述ADM、CPT-11、VCR的敏感度均有不同程度的提高,胞内的化疗药物阿霉素(ADM)的荧光强度增强。槲皮素的浓度提高至40μmol/L时,KB-MRP对上述药物的敏感程度提高更明显。流式细胞仪结果显示经槲皮素处理后KB-MRP细胞MRP1表达明显下降。结论:槲皮素可以逆转KB-MRP的多药耐药,逆转效果与剂量相关,逆转机制可能与MRP1的表达下调有关。     

腺苷酸活化蛋白激酶增强乳腺癌对多柔比星化疗敏感性的机制

背景与目的：腺苷酸活化蛋白激酶(AMP-activated protein kinase,AMPK)在调控细胞代谢和能量平衡方面起着重要作用,并与细胞增殖、生存和多种信号通路密切相关。近年来发现AMPK参与肿瘤的抑制和耐药。该研究旨在探讨AMPK对多柔比星抑制乳腺癌作用的影响及其机制。方法：采用四甲基偶氮唑蓝(meth-ylthiazolyl tetrazolium,MTT)法检测多柔比星作用后对MCF-7/adr、MCF-7/adr-vector及MCF-7/adr-AMPKα细胞增殖的影响;Hoechst染色法观察各组细胞凋亡形态;流式细胞术(flow cytometry,FCM)检测各组细胞凋亡率;荧光酶标仪检测3组细胞多柔比星累积量;蛋白[质]印迹法(Western blot)检测各组细胞中耐药蛋白及凋亡相关蛋白的表达。结果：多柔比星对MCF-7/adr细胞增殖抑制作用呈剂量和时间依赖性,其作用24、48 h的IC50值分别为(36.8±2.1)和(28.8±1.3)μg/mL。过表达AMPKα可增加多柔比星对MCF-7/adr细胞的生长抑制作用,呈剂量和时间依赖性,其作用24、48 h的IC50值分别为(16.0±0.7)和(4.2±0.2)μg/mL。荧光形态分析发现多柔比星联合AMPKα能诱导MCF-7/adr细胞凋亡。1.0μg/mL多柔比星作用48 h后,MCF-7/adr、MCF-7/adr-vector及MCF-7/adr-AMPKα细胞的凋亡率分别为(12.0±1.4)%、(12.7±1.6)%和(32.0±4.2)%,MCF-7/adr细胞中AMPKα过表达明显提高MCF-7/adr细胞对多柔比星的敏感性。荧光酶标仪检测显示,过表达AMPKα能明显提高细胞内多柔比星的累积量,具有浓度依赖性。Western blot实验结果显示,与MCF-7/adr和MCF-7/adr-vector细胞比较,MCF-7/adr-AMPKα细胞中Bax、细胞色素c(Cyto c)的释放、caspase-3和多聚腺苷二磷酸-核糖聚合酶降解产物(cleaved PARP)蛋白表达明显增加,而细胞外排泵P-糖蛋白(P-gp)和Bcl-2蛋白表达降低。结论：AMPKα可通过抑制耐药细胞外排泵以及调控凋亡相关蛋白的表达,从而增强乳腺癌耐药细胞对多柔比星的化疗敏感性。     

Effect of tamoxifen on mitoxantrone cytotoxicity in drug-sensitive and multidrug-resistant MCF-7 cells

The influence of the antiestrogen tamoxifen (TAM) on the activity of mitoxantrone (MXN), was evaluated against wild-type MCF-7/WT and their multidrug-resistant variant MCF-7/ADR cells. Multidrug resistance (MDR) in this cell line which was selected for resistance to Adriamycin (ADR), is associated with increased expression of P-glycoprotein (P-gp). In a clonogenic assay it was observed that TAM (1–10 M) significantly enhanced the activity of MXN in the MCF-7/ADR but not in the drug-sensitive cell line. Isobologram analysis indicated that the effect of the combination was additive in the parental MCF-7/WT cells and strongly synergistic in the MDR MCF-7/ADR cells. Also, TAM (10 M) caused a three-fold increase in the steady-state levels (C_ss) of MXN in MCF-7/ADR cells but did not modulate MXN levels in MCF-7/WT cells. The observed synergism in MCF-7/ADR cells was perhaps due to the increase in C_ss of MXN that may involve interaction of TAM with P-gp. The combination of MXN and TAM may be useful in the treatment of drug-sensitive and drug-resistant breast cancer.     

外源MDR-1基因对小鼠肺腺癌细胞耐药性产生的影响及机理研究

目的 了解ＭＤＲ－１基因表达中细胞药性的关系。方法 用ＦＵＧＥＮ＾ＴＭ６转染试剂将ＭＤＲ－１基因导入小鼠Ｌｅｗｉｓ肺腺癌细胞株（３ＬＬ）,建立池耐药细胞株（３ＬＬ－ＭＤＲ－１）．用ＭＴＴ方法测定和划搏定对药的逆转效应,ＭＤＲ－１基因表达产物ｐ－ｇｐ采用免疫组化方法观察,应用流式细胞仪分析其对示踪剂罗丹明１２３的摄取与排泄。结果 建立了一株能稳定表达ｐ－ｇｐ和对长春生物碱类、鬼白纱类具有显著耐药性的     

米非司酮逆转人乳腺癌细胞MCF-7/ADR耐多柔比星机制的探讨

目的 探讨米非司酮对乳腺癌耐药细胞株MCF-7/ADR的逆转耐药作用.方法 以亲本乳腺癌细胞MCF-7和耐多柔比星(阿霉素)乳腺癌细胞MCF-7/ADR为研究对象,分别应用MIF进行干预后,流式细胞仪检测MIF作用前后瘤细胞P-gp的表达、细胞内阿霉素蓄积量的变化以及细胞周期的分布.结果 (1)10 μmol/L MIF作用72小时后,MCF-7/ADR细胞P-gp表达率[(23.21±1.80)％]明显高于MCF-7细胞[(19.37±2.37)％,P＜0.05].(2)5 μmol/L ADR处理后,MCF-7/ADR细胞内ADR蓄积量为(47.13±4.11)％,低于MCF-7细胞[(60.24±2.61)％,P＜0.05].(3) 10 μmol/L MIF联合5 μmol/L ADR处理细胞,MCF-7/ADR和MCF-7细胞内ADR的蓄积量分别为(82.72±2.42)％及(88.63±2.75)％ (P ＞0.05);但均较单用ADR时升高(均P＜0.01).(4) MIF作用前,MCF-7/ADR细胞G0/G1期比例[(77.21±3.10)％]高于MCF-7细胞G0/G1期比例[(59.05±2.16)％,P＜0.05];MCF-7/ADR细胞S期比例明显低于MCF-7细胞(P＜0.05).经10 μmol/L MIF作用后,MCF-7细胞G0/G1期比例(75.28±2.53)％较MIF作用前明显升高(P＜0.05);S期比例则较MIF作用前显著降低(P＜0.05);MCF-7/ADR细胞G0/G1期比例和S期比例分别为(80.13±2.72)％及(13.52±1.03)％,与MIF作用前比较差异均无统计学意义(均P＞0.05);两种瘤细胞的G2/M期比例与MIF作用无关(P＞0.05).结论 (1)MIF可以逆转MCF-7/ADR的耐药性,其作用机制与降低细胞P-gp含量、增加细胞内ADR蓄积量有关.(2) 10 μmol/L浓度的MIF对MCF-7/ADR细胞周期分布影响不大.     

Modulation of Adriamycin Resistance in Human Breast Carcinoma MCF-7 Cells in vitro and in vivo by Medroxyprogesterone Acetate

The combination effect of adriamycin (ADM) and medroxyprogesterone acetate (MPA) was examined in vitro against human breast carcinoma MCF-7 and its ADM-resistant line (MCF-7/ADM). MCF-7 cells, which are positive for estrogen receptors, progesterone receptors and high-affinity MPA-binding activity, were more susceptible to the growth-inhibitory activity of ADM or MPA than MCF-7/ADM cells. A combination effect of ADM and MPA was observed against MCF-7/ADM cells, which are negative for steroid receptors, and furthermore against human nasopharynx carcinoma KB and its ADM-resistant line KB-A1. This combination effect of ADM and MPA against MCF-7/ADM cells was demonstrated to be synergistic by using the median effect plot method. The activity of MPA was almost equivalent to that of chlormadinone acetate or tamoxifen, greater than that of progesterone, and less than that of verapamil. The accumulation of ADM in MCF-7/ADM cells was enhanced by treatment with 10 μ M MPA as well as 10 μ M verapamil. The efflux of accumulated ADM from MCF-7/ADM cells was also partially inhibited by treatment with MPA or verapamil. MPA augmented the growth-inhibitory activity of ADM against MCF-7/ADM tumors inoculated into nude mice, although statistical significance was not observed. It is suggested that the clinical advantage of the combination of MPA with ADM against advanced breast cancers may be partly explained by the modulation of ADM resistance by MPA.     

Modulatory effect of tamoxifen and ICI 182,780 on adriamycin resistance in MCF-7 human breast-cancer cells

In this study the ability of the new pure anti-estrogen ICI 182,780 to modulate the cytotoxic action of adriamycin (ADR) on parental and ADR-resistant MCF-7 (MCF-7 ADRr) human breast-cancer cells was investigated and compared with that of tamoxifen (TAM). TAM enhanced ADR cytotoxicity in MCF-7 ADRr cells in a dose-related manner, but this effect was slight or absent in MCF-7 WT. In contrast, ICI 182,780 was able to enhance ADR toxicity both in MCF-7 ADRr and in the parental cell line. ICI 182,780 was up to 2.5-fold more effective than TAM in reducing the IC₅₀ of ADR in MCF-7 ADRr cells. Analysis of the data by the isobole method showed that the combination ADR/TAM and ADR/ICI 182,780 produced synergistic anti-proliferative activity in MCF-7 ADRr cells. Because ADR resistance in these cells is associated with the expression of high levels of P-glycoprotein (Pgp), we evaluated the effect of anti-estrogens on Pgp expression and activity. Both ICI 182,780 and TAM failed to modulate Pgp expression as assessed by flow cytometry and Western-blot analysis, performed using the monoclonal antibodies MM4.17 and C219, which are specific for an external or an internal determinant respectively. Pgp activity was investigated by flow cytometry measuring the extrusion of ADR and the cationic dye Rhodamine 123 (Rh 123). ICI 182,780, but not TAM, reduced the activity of Pgp in MCF-7 ADRr cells. Flow cytometry was also used to investigate cell-cycle modifications induced by ADR in MCF-7 ADRr cells, both in the presence and in the absence of anti-estrogens. After 72 hr, higher doses induced an arrest of cells at the G₂/M phase. The same effect was visible when lower doses of ADR were combined with ICI 182,780 or TAM. In terms of cell-cycle-blocking activity ICI 182,780 was largely more effective than TAM. © 1996 Wiley-Liss, Inc.     

Estradiol stimulates synthesis of a major intracellular protein in a human breast cancer cell line (MCF-7)

Summary Previous studies have shown that synthesis of a 24,000 molecular weight (24k) protein (7) and the mRNA activity coding for a 24k protein (8) are selectively stimulated by estradiol in MCF-7 human breast cancer cells. Utilizing a double-label electrophoresis technique to measure the relative rates of specific protein synthesis, the present study relates further characterization of this estrogen response. The 24k protein was found to be stimulated by estradiol provided cells were preincubated with antiestrogen. Under these conditions, antiestrogens inhibit cell growth and addition of estradiol reverses growth inhibition (estrogen growth rescue). As a control experiment, specific protein synthesis was measured in another growth rescue model produced by withdrawing serum from the medium to inhibit growth and then stimulating cells by readdition of serum (serum growth rescue). This failed to stimulate synthesis of a 24k protein, implying that the effect observed following estrogen rescue was not a non-specific result of growth stimulation. Increased 24k synthesis was found to be both time and estrogen-dose dependent and required specifically those steroid hormones which interact with the estrogen receptor. Moreover, the dose response curve for 24k stimulation paralleled closely the dose response for estrogen stimulation of nuclear estrogen receptor processing, an event correlated with another estrogenic response in MCF-7 cells, induction of progesterone receptor. These findings suggest that estradiol itself directly stimulates synthesis of the 24k protein through interaction with the estrogen receptor system. Further separation of double-label cytosols by two-dimensional electrophoresis resolved the 24k protein into two isoelectric species with pI's in the range of 6.7–6.9 and 6.4–6.7, the latter being the most abundant or rapidly labeling protein. Based on the percentage of total cytosol incorporation contained in individual two-dimensional gel spots, the major species of 24k represented in the fully stimulated cell (³H-counts) about 1.6% of newly synthesized protein. It also represented about 0.7% of newly synthesized protein in the unstimulated (¹⁴C-counts) cell, which indicates that 24k is synthesized constitutively. Because of its relative abundance, the 24k protein may be a suitable end product for investigating the mechanisms of estrogen action in human breast cancer.Abbreviations used are PgR, progesterone receptor ER estrogen receptor - DHT dihydrotestosterone - Hepes N¹-2-hydroxyethylpiperazine-N¹-ethane sulfonic acid - MEM Eagle's minimum essential medium - TCA trichloroacetic acid - SDS sodium dodecylsulfate; nafoxidine, (1-(2-[p-(3,4-dihydro-6-methoxy-2phenyl-1-naphthyl) Phenoxy]ethyl) pyrrolidine hydrochloride; Upjohn); tamoxifen, (1-p--dimethylaminoethyoxyphenyl-trans-1,2-diphenylbut-1-ene; ICI-46474); CI-628,1-[2-[p[-(p-methoxyphenyl)--nitrostyryl]-phenoxy]ethyl]pyrrolidine monohydrate monocitrate Address for reprints: William L. McGuir, M.D., Department of Medicine, University of Texas Health Science Center 7703 Floyd Curl Drive. San Antonio, TX 78284.     

Enhancement of radiosensitivity of the MCF-7 breast cancer cell line with human chorionic gonadotropin

Secretion of human chorionic gonadotropin (hCG) during pregnancy induces differentiation of the mammary gland, thereby making breast tissue less susceptible to carcinogenesis. HCG binds to specific hCG receptors on mammary epithelial cells inducing changes in gene expression that can inhibit cell proliferation and, therefore, interfere with tumorigenesis. Since breast cancer cells also contain a relatively high level of the hCG receptor, hCG has potential as a therapeutic agent. We postulated that hCG might also enhance the radiosensitivity of breast cancer cells and, therefore, be useful as an adjunctive therapy. In the present study, MCF-7 breast cancer cells grown in cell culture were treated with hCG (0.2–5IU/ml) for 24h prior to exposing the cells to 0 Gy, 3Gy, 4Gy, or 5Gy of radiation. Following irradiation, the MCF-7 cells were incubated either in the presence or absence of hCG. Cell survival was monitored with an MTT assay 1 day, 4 days, and 7 days after irradiation. All of the concentrations of hCG tested enhanced radiosensitivity of MCF-7 cells. The maximum enhancement occurred with MCF-7 cells that had been exposed to 2IU/ml of hCG for at least 24h prior to irradiation with 4Gy. The use of higher concentrations of hCG or a higher dose of radiation did not increase the enhancement effect. Treatment of MCF-7 cells with hCG for only 24h was sufficient to achieve the maximum effect. However, maintaining the cells in hCG beyond 24h increased the effectiveness of the lowest hCG concentration. Using a linear-quadratic equation to analyze the data, we determined that the use of hCG would result in an 8–10% reduction in MCF-7 cell survival at a dose of 2Gy, a typical dose used in conventional cancer therapy.     

Histone acetylation decreased by estradiol in the MCF-7 human mammary cancer cell line

Summary The effect of estradiol (E₂) on the [³H]-acetylation of nuclear histones was studied in the MCF-7 human mammary cancer cell line in culture. Cells (~ 10⁸) were incubated with 8 × 10^–6M [³H]-acetate in the absence (control) or in the presence of estradiol (10^–5–10^–8M). After 20 min incubation, the nuclear histones were extracted and separated by electrophoresis, and each histone band was measured and calculated in DPM/mg protein. It was observed that only the H₂ + H₃ and H₄ histones were associated with the [³H]-acetate. Estradiol (10^–6–10^–8M) provoked a significant inhibition in the incorporation of the acetate. The negative effect, in percentage of the non-treated cell value, was as follows: in E₂ (10^–6M): – 25 ± 10 (SE) for H₂ + H₃ and – 26 ± 5 for H₄; in E₂ (10^–7M): – 35 ± 9 and – 39 ± 10; and in E₂ (10^–8M): – 56 ± 22 and – 30 ± 13 respectively. It is concluded that estradiol has a negative effect in the acetylation of H₂, H₃ and H₄ histones of this mammary cancer cell; no acetylation or effect is observed in H₁ histones. The relationship of this finding to the biological responses of the hormone is to be explored.     

三氧化二砷逆转人乳腺癌MCF-7/ADM细胞耐药的机制研究

目的　探讨三氧化二砷 (As2 O3 )对人乳腺癌MCF 7/ADM细胞耐药性的逆转作用及其相关机制。方法　采用MTT法检测As2 O3 的细胞毒性及非细胞毒性剂量 ,对MCF 7/ADM细胞药敏性的影响。以荧光分光光度计测定细胞内药物浓度的改变 ,明确As2 O3 对MCF 7/ADM的逆转作用。同时 ,于逆转前后分别采用生化方法测定谷胱甘肽S 转移酶 (GSTs)酶活性 ,RT PCR方法检测GST πmRNA表达水平的改变。结果　As2 O3 的非细胞毒性剂量为 0 .2 μmol/L ,低毒剂量为 0 .8μmol/L。0 .2 μmol/LAs2 O3 可增加MCF 7/ADM细胞内阿霉素 (ADM)浓度 ,使细胞MCF 7/ADM的IC50 由原来的5 3.74 μmol/L降低至 2 5 .0 μmol/L ,其逆转倍数为 2 .1倍。在 0 .2 μmol/L、0 .8μmol/LAs2 O3 作用前后 ,GSTs酶活性有显著性改变 ,由 2 9.6 8± 0 .2 9(U/ml)分别降低为 19.2 9± 2 .10 (U/ml)、12 .6 6± 2 .78(U/ml) ,P <0 .0 5 ,而GST πmRNA的表达水平也呈不同程度的下调。结论　As2 O3 可部分逆转人乳腺癌MCF 7/ADM细胞对ADM的耐药性 ,其逆转机制可能与细胞内GST π的改变有关。     

人肝癌细胞耐药模型-7721/Adm的建立及其部分生物学特性

目的：建立耐药肝癌细胞模型－７７２１／Ａｄｕ并研究其生物学特性。方法：应用人肝癌细胞株－７７２１（简称７７２１细胞株）,采用阿霉素（Ａｄｍ）高浓度间歇诱导法,建立耐药人肝癌细胞模型－７７２１／Ａｄｍ（简称７７２１／Ａｄｍ细胞株）。观察该细胞的生长规律;用ＭＴＴ法检测该耐药细胞模型的多药耐药性;以流式细胞术检测该耐药模型细胞周期中的分布,细胞表面多药耐药基因（ｍｄｒ）的表达产物Ｐ－糖蛋白（Ｐ－ｇｐ）     

甲基莲心碱逆转肝癌HepG2/thermotolerance细胞对阿霉素耐受性的作用

背景与目的:如何成功地逆转耐热癌细胞的多药耐药性(multidrug resistance,MDR)是当前肿瘤热疗的研究热点.本研究探讨耐热肝癌细胞能否对阿霉素(adriamycin,ADR)产生耐受性,以及甲基莲心碱(neferine,Nef)能否逆转耐热肝癌细胞的阿霉素耐药性.方法:MTT法检测肿瘤细胞存活率,PI染色流式细胞仪检测细胞凋亡率,间接免疫荧光流式细胞术检测bcl-2表达.结果:在43℃环境中培养24 h后,耐热肝癌细胞HepG2/thermotolerance的细胞存活率和细胞凋亡率分别为(89.6±5.4)%和(13.6±5.4)%,而非耐热肝癌细胞HepG2的细胞存活率和细胞凋亡率分别为(23.9±3.6)%和(68.9±7.3)%.正常培养环境情况下(37℃),ADR对HepG2/thermotolerance细胞的IC50为(113.7±12.7)μmol/L,而对HepG2细胞的IC50为(10.5±2.3)μmol/L,耐药倍数达10.8倍.1、10、100 μmol/L ADR分别作用24 h后,HepG2/thermotolerance细胞的凋亡率分别为(9.3±2.6)%、(17.8±7.3)%和(32.9±8.6)%,而HepG2细胞的凋亡率分别为(14.3±3.9)%、(38.9±6.8)%和(62.7±5.9)%.在37℃培养环境下,10、40 μmol/L Nef对HepG2细胞和HepG2/thermotolerance细胞无增殖抑制作用和凋亡诱导作用,但可使ADR对HepG2/thermotolerance细胞的IC50分别下降至(63.7±5.6)μmol/L和(16.8±2.8)μmol/L,逆转倍数分别为1.78和6.79,并可使10 μmol/L ADR对HepG2/thermotolerance细胞凋亡的诱导作用升高至(26.8±5.9)%和(34.9±8.7)%;HepG2/thermotolerance细胞较HepG2细胞高表达Bcl-2蛋白,而Nef能下调HepG2/thermotolerance细胞的Bcl-2表达.结论:HepG2/thermotolerance细胞对ADR可产生耐受性,Nef可逆转HepG2/thermotolerance细胞对ADR的耐受性,其机制可能与其下调HepG2/thermotolerance细胞Bcl-2蛋白表达有关.