维甲酸逆转人宫颈癌耐药细胞株耐药性的实验研究

目的：本研究旨在探讨全反式维甲酸(ATRA)逆转入宫颈癌Hela／MMc耐药株对MMc的耐药作用。方法：观察ATRA作用下Hela／MMC细胞的增殖，采用MTT法观察ATRA对Hela／MMC药物敏感性的影响，半定量RT-PCR检测mdr I基因的表达情况。结果：ATRA能提高MMC对Hela／MMC细胞的杀伤作用，上调细胞mdr l mRNA的表达。结论：ATRA能逆转Hela／MMc对MMc的耐药性，但这种作用与mdfl基因表达无关。     

维甲酸逆转K526／ADM耐药细胞株耐药性的研究

维甲酸逆转Ｋ５２６／ＡＤＭ耐药细胞株耐药性的研究沈世人王衡邹长木炎苏颖高频关键词维甲酸Ｋ５６２／ＡＤＭ细胞多药耐药逆转中图号Ｒ７３－３６肿瘤患者反复用药引起耐药性的产生是治疗失败的重要原因，研究如何克服耐药性是提高癌症化疗效果的关键之一。本实验结果提...     

诺美孕酮逆转乳腺癌耐药细胞株多药耐药性的研究

目的 研究诺美孕酮（NOM）对乳腺癌耐药细胞株（MCF7／ADR）多药耐药性（MDR）的影响,探讨其调节机制。方法 应用MTT法,研究NOM对MCF7／ADR药物敏感性的影响。采用半定量逆转录聚合酶联反应（RT－PCR）和免疫细胞化学染色,分析耐药基因MDR1、谷胱甘肽S转移酶Pi(GSTπ）、拓扑异构酶Ⅱα（Topo Ⅱα)和多药耐药相关蛋白（MRP）表达的变化。通过流式细胞技术观察NOM对MCF7／ADR细胞内药物积累和细胞周期的影响。结果 NOM对MCF7／ADR的MDR具有明显的逆转作用,在20,10和5μmol/L浓度时的逆转倍数分别为21,12和8倍,逆转强度明显高于前身化合物甲地孕酮,而与维拉帕米相当。5μmol/L NOM处理MCF7／ADR后,MDRI的mRNA表达水平明显降低,呈现时间依赖性变化;P糖蛋白（P－gp)和GSTπ蛋白的变化符合相应mRNA表达的变化;MRP和Topo Ⅱα基因表达未见明显变化。用NOM 20,10和5μmol/L分别处理2h后,ADR在细胞内的积累分别增加到2．7倍、2．3倍和1．5倍。同时,NOM可明显加强ADR对MCF7／ADR细胞在G2M期的阻滞作用。结论 NOM具有较强的逆转MCF7／ADR细胞MDR的作用,其逆转机制为多种途径,包括时间依赖性下调MDRI和GSTπ基因的表达,增加细胞内药物积累,加强ADR对MCF7／ADR在G2M期的阻滞作用等。     

丹参酮和维甲酸对人宫颈癌细胞株的体外诱导分化作用

在体外细胞培养的基础上，通过细胞形态学、细胞增殖动力学、裸鼠成瘤性的研究，观察丹参酮对人宫颈癌细胞株（ＭＥ＿（１８０））的体外诱导分化作用（并以全反式维甲酸作对照）。结果表明：经无毒剂量的丹参酮和维甲酸处理后，细胞形态趋向良性分化，生长减慢，集落形成率和￣３Ｈ－ＴｄＲ掺入率明显降低，在裸鼠身上的成瘤时间延长，成瘤能力明显降低；经统计学处理，丹参酮和维甲酸对ＭＥ＿（１８０）细胞均具有较好的诱导分化作用，两者差异无显著意义。     

帕博西尼逆转人食管鳞癌奥沙利铂耐药细胞株耐药性的研究

  目的  建立人食管鳞癌(esophageal squamous cell carcinoma,ESCC)奥沙利铂(oxaliplatin,L-OHP)耐药细胞株KYSE150-OXA,研究细胞周期蛋白依赖性激酶4/6(cyclin-dependent kinase 4/6,CDK4/6)抑制剂帕博西尼(palbociclib)对耐药细胞株耐药性的逆转及其机制。  方法  CCK-8检测耐药细胞对奥沙利铂、紫杉醇(paclitaxel,PTX)、5-氟尿嘧啶(5-fluorouracil,5-FU)以及CDK4/6抑制剂palbociclib的耐药性;流式细胞术检测化疗药物对耐药细胞凋亡和周期的影响;Western blot检测细胞中多药耐药基因1(multidrug resistance gene 1,MDR1)及细胞周期相关蛋白的表达量;CalcuSyn软件计算palbociclib与L-OHP联合用药对细胞增殖的作用。  结果  成功建立人体外ESCC L-OHP耐药细胞株KYSE150-OXA,耐药指数(resistance index,RI)为11.23±0.59,耐药细胞KYSE150-OXA对PTX和5-FU也表现出耐药性,Western blot结果显示耐药细胞中MDR1蛋白表达升高(4.46±0.22)倍;与亲本细胞KYSE150相比,耐药细胞株G1期细胞显著增多,且CDK4、CDK6及Cyclin D1表达显著升高;palbociclib增加KYSE150-OXA对L-OHP的敏感性;低浓度palbociclib与L-OHP表现出拮抗作用,高浓度时表现为协同作用;palbociclib处理组细胞中磷酸化视网膜母细胞瘤蛋白(phospho-retinoblastoma,pRb)降低(91±2)%及细胞周期蛋白A(Cyclin A)降低(89±6)%。  结论  成功建立人体外ESCC L-OHP耐药细胞株KYSE15 0-OXA;palbociclib可能通过抑制异常激活的Cyclin D-CDK4/6-pRb信号通路逆转耐药细胞对L-OHP的耐药性。      

潘生丁对KB细胞株对多药耐药逆转作用的实验研究

目的：观察了潘生丁对ＫＢ细胞株多药耐药的逆转作用并对其作用机制进行探讨。方法：使用ＭＴＴ法检测比较ＫＢ母细胞和耐药细胞株对长春新碱（ＶＣＲ）的药物敏感度并观察潘生丁对耐药的逆转程度;用＾３Ｈ标记的ＶＣＲ检测细胞内药物浓度的变化。结果：潘生丁作用下,ＫＢ母细胞和耐药细胞株的药物敏感度均有增高并呈剂量相关关系,有ＭＰＰ表达的耐药细胞株对ＭＲＰ表达的耐药细胞株对ＶＣＲ的敏感度比母细胞株高１７倍,母细胞株     

屈洛昔芬逆转人乳腺癌耐药细胞株MCF7／ADR多药耐药性的研究

目的：研究屈洛昔芬（DRO）对乳腺癌耐药细胞株（MCF7／ADR）多药耐药性（MDR）的影响。方法：应用MTT，半定量RT－PCR和免疫细胞化学染色，研究DRO对MCF7／ADR药物敏感性，耐药相关基因MDR1，谷胱甘肽S转移酶Pi(GST-π），拓扑异构酶Ⅱα（Topo Ⅱα)和多药耐药相关蛋白（MRP）表达的影响。流式细胞技术观察细胞内药物积累。结果：在20，10和5μmol/L浓度时DRO对MCF7／ADR耐药性的逆转倍数分别为16，8和3倍，逆转强度与同类化合物三苯氧胺相当。DRO处理后，MDR1和GSTπ基因表达水平明显降低。DRO使ADR在细胞内的积累分别增加到2.5(20μmol/L),2.0(10μmol/L)和1.5(5μmol/L)倍。结论：DRO具有较强的逆转MCF7／ADR耐药性的作用，其逆转机制是多途径的。     

卡维地洛逆转膀胱癌细胞株多药耐药性

目的：研究卡维地洛对人膀胱癌耐药细胞株BIU-87／ADM的多药耐药性逆转作用。方法：应用MTT法检测人膀胱癌耐药细胞株(BIU-87／ADM)的耐药性；将卡维地洛与多柔比星联合作用于B10-87和BIU-87／ADM细胞，检测细胞内多柔比星聚集量。结果：卡维地洛与多柔比星共同温育BIU-87／ADM细胞后．能使BIU-87／ADM细胞内多柔比星聚积量增加，与单用多柔比星比较，差异有显著性。结论：卡维地洛对多柔比星有增敏作用，卡维地洛可逆转人膀胱癌细胞株BIU-87／ADM的多药耐药性。     

肿瘤耐药基因表达的临床意义及其逆转研究

对人白细胞病多药耐药细胞株K562／A02和人肺癌耐PDD细胞株A549PDD系统研究表明，耐药表型是多基因的，它随诱导的药物不同，肿瘤肿类，分化程度以及宿主微环境的不同而表现为一种或同种耐药同时表达，临床研究肿瘤共524例包括白血病260例，实体瘤264但目前能在临床证实跟一些肿瘤耐药相关的基础只有mdrl/P-gp。用比较敏感细菌和对应的耐药细胞cDNA阵表达水平的判别新技术发现了除已知mdrl等等耐药基因以外新的与药相关基因，这也为基因组学在耐药基因研究开辟了新的途径。开发出新的耐药逆转剂PZT11、PND和抗耐药肿新药PHⅡ-7有望在临床发挥作用。抗P－gp／抗CD3微型双功能抗体也有望成为逆转肿瘤耐药新有力工具。     

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。     

Reversing Effect of Agosterol A, a Spongean Sterol Acetate, on Multidrug Resistance in Human Carcinoma Cells

The effect of agosterol A, a novel polyhydroxylated sterol acetate isolated from a marine sponge, on P-glycoprotein (P-gp)-mediated multidrug-resistant cells (KB-C2) and the multidrug resistance associated protein (MRPl)-mediated multidrug-resistant cells (KB-CV60) was examined. Agosterol A reversed the resistance to colchicine in KB-C2 cells and also the resistance to vincristine in KB-CV60 cells at 3 to 10 μM concentration. Agosterol A at 3 μM increased the vincristine concentration in both KB-C2 cells and KB-CV60 cells to the level in parental KB-3-1 cells. Agosterol A also decreased the efflux of vincristine from both KB-C2 cells and KB-CV60 cells to the level seen in KB-3-1 cells. Agosterol A inhibited the [³H]azidopine-photolabeling of P-gp and also inhibited the uptake of [³H]S-(2,4-dinitrophenyl)glutathione (DNP-SG) in inside-out membrane vesicles prepared from KB-CV60 cells. We conclude that agosterol A directly inhibited drug efflux through P-gp and/or MRP1.     

二甲亚砜对K562/ADM耐药细胞株耐药性的逆转作用

目的：研究二甲亚砜对K562／ADM耐药细胞的逆转作用。方法：通过加入二甲亚砜后耐药细胞的生长情况，逆转倍数的计算，细胞内过氧化物染色的阳性程度以及电镜下观察耐药细胞诱导前后的变化。结果：二甲亚砜诱导K562／ADM耐药细胞前后生长情况有显著差异且逆转借故为4．0，Pox染色阳性程度明显递增，电镜下可观察诱导后K562／ADM耐药细胞有向成熟分化趋势。结论：二甲亚砜对逆转K562／ADM的耐药性是有益的。     

体内诱导艾氏腹水瘤细胞的耐药性研究

肿瘤细胞耐药性的存在是临床化疗失败的主要原因之一。本实验在小鼠体内用阿霉素（ADR）诱导艾氏腹水瘤细胞（EHR）的耐药性，探讨细胞产生耐药性的机理。HPLC法测定细胞内药物浓度．结果表明耐药细胞─—EHR/ADR细胞内ADR积聚低于EHR细胞，而对ADR外排快于EHR细胞；异博定（VER）增加EHR／ADR细胞对ADR的摄取并阻滞其外排．而对EHR影响不大，揭示EHR／ADR细胞具有MDR特性。     

RNAi-based Knockdown of Multidrug Resistance-associated Protein 1 is Sufficient to Reverse Multidrug Resistance of Human Lung Cells

Up-regulation of multidrug resistance-associated protein 1 (MRP1) is regarded as one of the main causes formultidrug resistance (MDR) of tumor cells, leading to failure of chemotherapy-based treatment for a multitude ofcancers. However, whether silencing the overexpressed MRP1 is sufficient to reverse MDR has yet to be validated.This study demonstrated that RNAi-based knockdown of MRP1 reversed the increased efflux ability and MDRefficiently. Two different short haipin RNAs (shRNAs) targeting MRP1 were designed and inserted into pSilence-2.1-neo. The shRNA recombinant plasmids were transfected into cis-dichlorodiamineplatinum-resistant A549lung (A549/DDP) cells, and then shRNA expressing cell clones were collected and maintained. Real time PCRand immunofluorescence staining for MRP1 revealed a high silent efficiency of these two shRNAs. Functionally,shRNA-expressing cells showed increased rhodamine 123 retention in A549/DDP cells, indicating reduced effluxability of tumor cells in the absence of MRP1. Consistently, MRP1-silent cells exhibited decreased resistance to3- (4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) and DDP, suggesting reversal of MDRin these tumor cells. Specifically, MRP1 knockdown increased the DDP-induced apoptosis of A549/DDP cellsby increased trapping of their cell cycling in the G2 stage. Taken together, this study demonstrated that RNAibasedsilencing of MRP1 is sufficient to reverse MDR in tumor cells, shedding light on possible novel clinicaltreatment of cancers.     

氧化苦参碱逆转多药耐药细胞系K562/A02耐药性的研究

目的: 观察非细胞毒性浓度(生长率>95%)氧化苦参碱对耐阿霉素的人白血病细胞系K562/A02多药耐药性的逆转作用,并探讨其逆转机制。 方法: 采用MTT法测定氧化苦参碱的细胞毒性及其对K562/A02细胞敏感性的影响,用流式细胞仪检测非细胞毒性浓度的氧化苦参碱处理后K562/A02细胞膜表面糖蛋白P170表达的变化与细胞内柔红霉素的浓度,应用SPSS13.0软件包对实验结果进行统计学处理。 结果: 氧化苦参碱对K562/A02细胞有一定的细胞毒作用,其非细胞毒性浓度为50μg/ml,低细胞毒性浓度(生长率为85%～90%)为135μg/ml,把非细胞毒性剂量的氧化苦参碱作为最佳逆转浓度,非细胞毒性浓度氧化苦参碱可显著降低阿霉素对K562/A02细胞的IC₅₀,使K562/A02细胞的IC₅₀由原来的(34.9±0.21)μg/ml降低至(13.3±0.21)μg/ml,其逆转倍数为2.62倍;氧化苦参碱作用于K562/A02细胞后,细胞膜糖蛋白P170的表达从90.22%下调至44.24%(P<0.01);柔红霉素外渗试验显示,氧化苦参碱作用于K562/A02细胞后,细胞内化疗药物的浓度明显增加。 结论: 氧化苦参碱可部分逆转人白血病K562/A02细胞对阿霉素的耐药性,氧化苦参碱通过下调K562/A02细胞膜糖蛋白P170的表达,使其将化疗药物泵出细胞外的功能受到了抑制,使化疗药物在白血病细胞内能达到有效浓度,从而可以杀灭耐药的白血病细胞,达到逆转白血病细胞耐药性的目的。     

SiRNA逆转T24/ADM细胞耐药性的研究

目的探讨siRNA（small interfering RNA）对膀胱癌多药耐药性的逆转作用。方法设计针对MDR1基因的siRNA,转染膀胱癌T24/ADM细胞,应用RT-PCR分析MDR1 mRNA的表达,免疫印迹检测MDR1蛋白表达,流式细胞仪检测细胞内阿霉素积累量。MTT法检测阿霉素对T24/ADM细胞的半数抑制浓度（IC50）。结果 3条siRNA均能不同程度地逆转T24/ADM细胞的多药耐药性,使MDR1基因的mRNA及蛋白表达水平下调,细胞内阿霉素积累量显著增加（P〈0.05）。结论 siRNA可逆转膀胱癌的多药耐药性。     

Establishment by Adriamycin Exposure of Multidrug-resistant Rat Ascites Hepatoma AH130 Cells Showing Low DT-diaphorase Activity and High Cross Resistance to Mitomycins

A resistant subline (AH130/5A) selected from rat hepatoma AH130 cells after exposure to adriamycin (ADM) showed remarkable resistance to multiple antitumor drugs, including mitomycin C (MMC) and porflromycin (PFM). PFM, vinblastine (VLB), and ADM accumulated in AH130/5A far less than in the parent AH130 (AH130/P) cells. AH130/5A cells showed overexpression of P-glycoprotein (PGP), an increase in glutathione S-transferase activity, and a decrease in DT-diaphorase and glutathione peroxidase activity. The resistance to MMC and VLB of AH130/5A cells was partly reversed by H-87, an inhibitor of PGP. Buthionine sulfoximine, an inhibitor of glutathione synthase, did not affect the action of MMC. tert -Butylhydroquinone induced DT-diaphorase activity, increased PFM uptake, and enhanced the growth-inhibitory action of MMC in AH130/5A cells. Dicumarol, an inhibitor of DT-diaphorase, decreased PFM uptake and reduced the growth-inhibitory action of MMC in AH130/P cells. These results indicated that the adriamycin treatment of hepatoma cells caused multifactorial multidrug resistance involving a decrease in DT-diaphorase activity.     

Mechanistic Analysis of Taxol-induced Multidrug Resistance in an Ovarian Cancer Cell Line

Objectives: To establish a taxol-resistant cell line of human ovarian carcinoma (A2780/Taxol) and investigateits biological features. Methods: The drug-resistant cell line (A2780/Taxol) was established by continuous stepwiseselection with increasing concentrations of Taxol. Cell morphology was assessed by microscopy and growth curveswere generated with in vitro and in vivo tumor xenograft models. With rhodamine123 (Rh123) assays, cell cycledistribution and the apoptotic rate were analyzed by flow cytometry (FCM). Drug resistance-related and signalassociated proteins, including P-gp, MRPs, caveolin-1, PKC-α, Akt, ERK1/2, were detected by Western blotting.Results: A2780/Taxol cells were established with stable resistance to taxol. The drug resistance index (RI) was430.7. Cross-resistance to other drugs was also shown, but there was no significant change to radioresistance.Compared with parental cells, A2780/Taxol cells were significantly heteromorphous, with a significant delay inpopulation doubling time and reduced uptake of Rh123 (p < 0.01). In vivo, tumor take by A2780 cells was 80%,and tumor volume increased gradually. In contrast, with A2780/Taxol cells in xenograft models there was notumor development. FCM analysis revealed that A2780/Taxol cells had a higher percentage of G0/G1 and lowerS phase, but no changes of G2 phase and the apoptosis rate. Expression of P-gp, MRP1, MRP2, BCRP, LRP,caveolin-1, PKC-α, Phospho-ERK1/2 and Phospho-JNK protein was significantly up-regulated, while Akt andp38 MARK protein expression was not changed in A2780/Taxol cells. Conclusion: The A2780/Taxol cell line isan ideal model to investigate the mechanism of muti-drug resistance related to overexpression of drug-resistanceassociated proteins and activation of the PKC-α/ERK (JNK) signaling pathway.     

异汉防己碱增强多药耐药肿瘤细胞对阿霉素的敏感性及其机制

目的:以K562/DOX和MCF-7/DOX细胞为对象,探讨异汉防己碱对化疗药物阿霉素（DOX）的增敏作用及其作用机制。方法:采用MTT法检测异汉防己碱的内在细胞毒性及其对阿霉素的增敏作用,并以RF值评价其增敏效果。应用流式细胞术（FCM）检测细胞膜上P-gp的表达以及细胞内DOX和罗丹明123（Rh123）的蓄积量。结果:异汉防己碱在10μg/ml的无毒剂量可明显增强DOX的细胞毒性。K562/DOX和MCF-7/DOX细胞膜上P-gp均呈强阳性表达,但异汉防己碱对该P-gp表达水平无明显影响。异汉防己碱可使K562/DOX和MCF-7/DOX细胞内DOX和123的荧光密度（FI）均明显增加,由此证明异汉防己碱可有效抑制P-gp的功能。结论:异汉防己碱可通过抑制P-gp的功能而增强阿霉素的敏感性,从而有效逆转肿瘤细胞的多药耐药性（MDR）,它可能成为有效多药耐药逆转剂的候选药物。