欧白芷素对K562/A02细胞阿霉素耐药的逆转作用(英文)

目的:探讨欧白芷素对耐药细胞株K562/A02中P-糖蛋白(P-glycoprotein,P-gp)的影响,为抗白血病多药耐药(multi-drug resistance,MDR)提供新方法。方法:采用MTT法观察阿霉素对细胞活力的影响。应用流式细胞术检测欧白芷素对K562和K562/A02细胞内阿霉素累积和细胞中P-gp功能的影响。采用实时定量RT-PCR技术检测MDR1基因在mRNA表达水平的变化。结果:欧白芷素对耐药细胞株K562/A02有显著的逆转耐药活性,最大逆转倍数为7.36。在K562/A02细胞中,欧白芷素明显增加阿霉素的累积,增加了罗丹明123(rhodaminel123,Rh123)蓄积,抑制了Rh123的外排,同时欧白芷素还在mRNA水平抑制了K562/A02细胞中P-gp的表达。结论:欧白芷素能够抑制K562/A02耐药细胞株中MDR1基因表达和P-gp的功能。     

六神丸逆转K562/DOX细胞阿霉素耐药的研究

目的:探讨六神丸对阿霉素耐药细胞株K562/DOX的耐药逆转作用。方法:MTT法检测单独应用阿霉素或阿霉素联合六神丸含药血清对K562/DOX细胞的生存率的影响;RT-PCR检测MDR1 mRNA表达;Western blot检测MDR1蛋白的表达。结果:阿霉素与六神丸含药血清联合作用:①细胞抑制率最高,最高值出现在72h(49.4%),且与对照组和单独应用阿霉素组相比有显著性差异;②48h细胞MDR1 mRNA表达明显降低,与单独应用阿霉素组相比有显著性差异;③48h细胞MDR1蛋白表达下降,且与对照组和单独应用阿霉素组相比有显著性差异。结论:六神丸可能通过下调MDR1表达水平进而逆转K562/DOX细胞对阿霉素的耐药性,增加肿瘤耐药细胞对化疗药物的敏感性。     

六神丸及其成分蟾酥对K562/DOX细胞的增殖和MDR1基因表达的影响

目的:探讨传统中成药六神丸及其成分蟾酥对K562/DOX细胞的增殖和MDR1基因表达的影响。方法:采用中药血清药理学方法制备六神丸和蟾酥的含药血清,以MTT法检测六神丸以及蟾酥含药血清、阿霉素处理后K562/DOX细胞的抑制率;RT-PCR检测MDR1 mRNA表达;Western blot检测MDR1蛋白的表达。结果:与对照组相比,蟾酥和六神丸对K562/DOX细胞均有抑制作用;六神丸组MDR1 mRNA表达降低,MDR1蛋白表达下降;而蟾酥组、阿霉素组未有明显变化。结论:六神丸可能通过下调MDR1表达水平进而逆转K562/DOX细胞的耐药性,而其成分蟾酥作用不明显。     

硒化壳聚糖逆转K562/ADM细胞多药耐药的实验研究

目的:研究硒化壳聚糖对人慢性粒细胞白血病耐阿霉素细胞株( K562/ADM) mdr-1基因/P-糖蛋白(P-gp)表达及功能的影响,为逆转肿瘤多药耐药提供新的途径.方法:硒化壳聚糖100,200 mg·L-1作用K562/ADM细胞24 h,应用RTPCR法和免疫印迹法检测mdr-1/P-gp表达的改变;应用高效液相色谱法检测细胞内阿霉素积聚浓度;应用MTT法检测阿霉素对K562/ADM细胞增殖的影响.结果:硒化壳聚糖可增强K562/ADM细胞对阿霉素的敏感性,增加细胞内阿霉素集聚浓度,200 mg·L-1硒化壳聚糖作用效果显著强于100 mg·L-1硒化壳聚糖(P＜0.01);硒化壳聚糖能明显抑制K562/ADM细胞mdr-1/P-gp的表达(P＜0.01),100 mg·L-1硒化壳聚糖可使mdr-1/P-gp表达分别下降(40.87 -3.19)％和(35.08±0.09)％,200 mg·L-1硒化壳聚糖可使mdr-1/P-gp表达分别下降(78.24±3.42)％和(79.61±0.23)％.结论:硒化壳聚糖可明显抑制K562/ADM细胞mdr-1/P-gp表达,增加细胞内阿霉素含量,恢复细胞对化疗药物敏感性,逆转mdr-1编码蛋白P-gp介导的多药耐药.     

三七总皂甙体外逆转K562/VCR细胞多药耐药的实验研究

目的:从中药中寻找安全有效的多药耐药逆转剂,并初步研究三七总皂甙(PNS)对人白血病细胞K562(敏感细胞)及K562/VCR(耐药细胞)的影响.方法:以MTT法测定三七总皂甙注射液对K562及K562/VCR的直接细胞毒作用;测定阿霉素(DOX)对两种细胞的毒性作用并计算出耐药倍数;用流式细胞仪测定PNS作用后敏感细胞和耐药细胞内阿霉素的浓度;以流式细胞仪测定敏感细胞和三七总皂甙作用的耐药细胞表达多药耐药糖蛋白P-gp(P-170)的阳性率.以上实验均用维拉帕米作为阳性对照.结果:①三七总皂甙在300μg/ml浓度以下时对两细胞基本无毒性,为安全有效的逆转剂量.②阿霉素对敏感细胞和耐药细胞的半数抑制浓度(IC50)分别为28.7ng/ml和1141.8ng/ml,耐药倍数为39.7倍.③三七总皂甙能够增强阿霉素(DOX)对K562/VCR的细胞毒作用.④三七总皂甙能够增加耐药细胞K562/VCR细胞内的阿霉素的蓄积浓度.⑤三七总皂甙(200μg/ml)可下调多药耐药基因mdr-1表达的P-gp糖蛋白的量,而未发现维拉帕米的类似作用.结论:三七总皂甙能够部分逆转耐药细胞K562/VCR的多药耐药性.     

小檗碱增强K562/DOX细胞对多柔比星敏感性的研究

 目的 探讨小檗碱增强耐药K562/DOX细胞对化疗药多柔比星的敏感性的作用。方法 四甲基偶氮唑蓝法检测小檗碱的细胞毒性及其对多柔比星抗肿瘤活性的增强作用;高内涵活细胞成像系统检测无毒剂量小檗碱作用后,多柔比星在K562/DOX细胞内的蓄积量;PI/Hoechst33342双染法检测小檗碱对多柔比星诱导的K562/DOX细胞凋亡的影响;罗丹明123蓄积实验检测小檗碱对P-糖蛋白外排功能的影响。结果 1 μmol·L^-1为小檗碱的无毒剂量,在此无毒剂量下,小檗碱使多柔比星对K562/DOX细胞的IC₅₀降低了1.5倍;1 μmol·L^-1小檗碱可使多柔比星在K562/DOX细胞内的蓄积量增加,增强多柔比星诱导的K562/DOX细胞凋亡, 增加K562/DOX细胞内罗丹明123的蓄积量,从而抑制P-糖蛋白的外排功能。结论 小檗碱可通过抑制K562/DOX细胞膜上P-糖蛋白的外排功能,增加K562/DOX细胞内多柔比星浓度,促进多柔比星对耐药细胞的诱导凋亡作用,逆转K562/DOX细胞的多药耐药性。     

蟾酥逆转人白血病K562/DOX细胞多药耐药的实验研究

目的 探讨传统中药蟾酥逆转人白血病K562/DOX细胞多药耐药性（MDR）的作用。方法 采用中药血清药理学方法制备蟾酥含药血清,以MTT法分别检测蟾酥含药血清联合柔红霉素,或单用蟾酥含药血清处理后,不同时段作用于K562/DOX细胞的抑制率;实时荧光定量PCR法检测蟾酥含药血清对K562/DOX细胞的Bcl-2 mRNA表达。结果 与空白对照组比较,蟾酥对K562/DOX细胞均有抑制作用;蟾酥组Bcl-2 mRNA表达降低。结论 蟾酥可能通过下调MDR1表达水平进而逆转K562/DOX细胞的耐药性。     

蛇葡萄素对K562/ADR细胞耐药性的逆转作用及机制研究

目的:研究蛇葡萄素与阿霉素合用对人白血病多药耐药细胞株K562/ADR增殖的抑制作用及其机制.方法:MTT法测定蛇葡萄素与阿霉素合用对K562/ADR细胞的细胞毒作用,应用金正均公式进行联合用药效果分析;藻红蛋白标记抗体检测蛇葡萄素对K562/ADR细胞膜表面P-糖蛋白表达的影响;流式细胞术测定蛇葡萄素对K562/ADR细胞内阿霉素累积的影响.结果:1.25～5 mg·L~(-1)的蛇葡萄素能够明显逆转K562/ADR细胞对阿霉素的耐药性.1.25 mg·L~(-1)蛇葡萄素与低浓度阿霉素合用可表现出拮抗效应,而浓度在2.5 mg·L~(-1)以上的蛇葡萄素与阿霉素合用可表现出相加至协同效应.蛇葡萄素能够浓度依赖性减少K562/ADR细胞膜P-糖蛋白表达,增加细胞内阿霉素的浓度.结论:蛇葡萄素能通过抑制P-糖蛋白外排药物,促进抗癌药物在细胞内的累积,增强抗癌药物对耐药细胞的细胞毒作用,逆转肿瘤多药耐药.     

白藜芦醇四甲氧基衍生物提高耐药人乳腺癌细胞MCF-7/DOX对阿霉素的敏感性

目的:研究白藜芦醇四甲氧基衍生物(TMS)对人乳腺癌细胞MCF-7和耐阿霉素(doxorubicin,DOX)的人乳腺癌细胞MCF-7/DOX这两株细胞增殖的影响,探讨白藜芦醇四甲氧基衍生物能否提高耐药细胞MCF-7/DOX对阿霉素的敏感性。方法:用SRB法检测白藜芦醇四甲氧基衍生物的细胞毒性,同时检测白藜芦醇四甲氧基衍生物和DOX联用后对细胞株MCF-7/DOX药物敏感性的影响,用Western-blot方法检测不同浓度的白藜芦醇四甲氧基衍生物对P-gp蛋白表达的影响,罗丹明转运实验考察白藜芦醇四甲氧基衍生物对P-gp转运活性的影响。结果:白藜芦醇四甲氧基衍生物(7.5、15、30μg/ml)能够有效抑制耐药细胞MCF-7/DOX的增殖,然而与此相应的浓度范围内白藜芦醇的细胞增殖抑制作用较差,使用非细胞毒性剂量(3、6μg/ml)的白藜芦醇四甲氧基衍生物能有效提高MCF-7/DOX对阿霉素的敏感性,Western-blot结果显示6μg/ml的白藜芦醇四甲氧基衍生物能够明显下调P-gp的表达,此外,罗丹明转运实验显示白藜芦醇四甲氧基衍生物组(3、6μg/ml)细胞内罗丹明123积累量均有上升,表明白藜芦醇四甲氧基衍生物可以抑制P-gp的转运活性。结论:白藜芦醇四甲氧基衍生物可通过抑制P-gp表达和活性的方式增加MCF-7/DOX对阿霉素的敏感性,从而逆转耐药。     

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??OBJECTIVE To investigate the effect of TiO₂ nanoparticles on overcoming cancer multidrug resistance(MDR). METHODS Doxorubicin-TiO₂ nanoparticles(DTN) were prepared, the K562/DOX cells were chosen as the model cells. And doxorubicin solution(F-DOX) and doxorubicin liposomes(DOX-L) were also prepared as the control. The MTT assay were measured, and the amount of doxorubicin in the K562/DOX cells at different time were determined by HPLC. The P-gp expression were detected by flow cytometry.RESULTS The MTT assay shows that IC₅₀ of group DTN were lower than that of group F-DOX. The uptake test shows that amount of doxorubicin in K562/DOX cells of group DTN was 1.23 times of group DOX-L when in 4 h, and the efflux test shows that amount of doxorubicin in K562/DOX cells was 1.18 times of group DOX-L. The flow cytometry result revealed that the effect of TiO₂ nanoparticles on overcoming MDR maybe through down-regulating the expression of P-gp in K562/DOX cells.CONCLUSION The TiO₂ nanoparticles are a new inorganic materials-based nanoparticles which promising approach to overcome MDR.     

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??OBJECTIVE To investigate the synergetic effect of schisandrin B and liposomes on overcoming multidrug resistance(MDR). METHODS Co-delivery of doxorubicin and schisandrin B by liposome(DS-L) were prepared, and doxorubicin solution(F-DOX), schisandrin B and doxorubicin mixture(Sch B+DOX), doxorubicin liposomes(D-L), doxorubicin liposomes and schisandrin B mixture(D-L+Sch B), verapamil and doxorubicin mixture(Ver+DOX) were also prepared as the control. The MTT test were measured, and the amount of doxorubicin in the K562/DOX cells at different time were determined, and time course of uptake and efflux were drawn. RESULTS The MTT test shows that the resistance factor(RF) of group DS-L were 1.68, 14.52 and 1.42 times of group Sch B+DOX, group D-L and group D-L+Sch B respectively. The uptake test shows that amount of doxorubicin of K562/DOX cells in group DS-L was 1.30 and 1.21 times of that in group D-L and group Sch B+DOX respectively. And the efflux test shows DS-L could delay doxorubicin efflux from K562/DOX cells. CONCLUSION The co-delivery of chemotherapeutics and P-gp inhibitors schisandrin B by liposome is a promising approach to overcome MDR. And there is a synergistic effect between liposome and schisandrin B to overcome MDR.     

长期用药在血脑屏障上诱导P-糖蛋白介导的耐药性

的：考察长期用药在血脑屏障上是否引起耐药性及P-糖蛋白(P-gp)表达增强。方法：原代牛脑微血管内皮细胞(BCEC)加入环孢素A(CsA)，长春新碱(VCR)，阿霉素(Dox)，或粉防己碱(Tet)，初始剂量分别为0．0083，0．091，0．34，或0．32μmol/L，培养至传代剂量翻倍。连续作用21，37，51或69d后，用罗丹明123(Rh123)检测P-gp功能。将各药物诱导69天的BCEC制膜，用酶联免疫吸附法(ELISA)测定P-gp表达结果：Dox用药37d使胞内Rh123浓度降低38％连续给药51和69d后，Dox分别使胞内Rh123浓度降低47％和57％，VCR组分别降低36％和40％。而CsA和Tet组一直未见明显变化。维拉帕米(10μmol/L)分别使CsA、Tet、Dox和VCR诱导组BCEC内Rh123的摄取增加92％、85％、143％和186％Dox和VCR连续用药69d使P-gp的表达增强45％和32％结论：长期使用Dox或VCR可在血脑屏障上诱导P-gp介导的耐药性以及P-gp表达增强。     

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??OBJECTIVE To prepare a redox and pH dual sensitive nano-carrier based on PAMAM in order to co-loading chemotherapeutics doxorubicin and breast cancer multidrug resistance reversal agent elacridar, and study their in vitro reversal effect. METHODS The infrared spectrum FTIR was used to characterize the carrier. Confocal was used to investigate the intracellular triggered drug release. The reversal effect of breast cancer multidrug resistance and the in vitro anti-tumor activity of doxorubicin and elacridar co-loaded nanoparticles were investigated using flow cytometry and cell toxicity tests, respectively. RESULTS The doxorubicin and elacridar co-loaded nanoparticles (PSSP/DOX/ELC) were successfully prepared, and pH-redox dual sensitive of carrier was proved by cell experiments.And the carrier was uptaken into cells and delivery to lysosome, and drug release was triggered in the lysosome acid condition, then the released drug diffused to the nucleus. The trial of rhodamine 123 accumulation and efflux assay revealed that the accumulation of rhodamine 123 was notably increased after incubation of elacridar in MCF-7/ADR cells. The cytotoxicity of PSSP/DOX/ELC nanoparticles against MCF-7/ADR cell line was significantly stronger than that of either free doxorubicin or only doxorubicin loaded nanoparticles (PSSP/DOX). CONCLUSION The reversal effect of multidrug resistance and the cytotoxicity of cancer cells were significantly enhanced by PSSP/DOX/ELC nanoparticles. PSSP/DOX/ELC nanoparticles is a promising delivery system.     

复方中药紫龙金克服肿瘤细胞多药耐药性的机制

目的探讨中药紫龙金(Zilongjin,ZLJ)对多药耐药肿瘤细胞的作用机制。方法采用MTT法检测ZLJ对细胞增殖的影响;流式细胞术检测细胞周期以及罗丹明123的荧光强度变化;Western blot方法检测相关蛋白的表达变化。结果 ZLJ分别处理人乳腺癌MCF-7和MCF-7/DOX耐药细胞,以及人口腔上皮癌KB和KBV200耐药细胞。MTT法测定表明:ZLJ作用耐药和敏感细胞的IC50值相近,耐药细胞对ZLJ没有交叉耐药性;无论对敏感和耐药细胞,流式细胞术分析发现ZLJ阻断细胞于S期;ZLJ单独处理MCF-7/DOX和KBV200耐药细胞,可以微弱地降低其耐药性,分别与多柔比星(doxorubicin,DOX)和长春新碱(vincris-tine,VCR)合用,可以明显地增加DOX和VCR的活性;ZLJ处理耐药细胞MCF-7/DOX后,检测到细胞内耐药蛋白P-糖蛋白(P-glyco protein,P-gp)呈时间依赖性降低。Western blot检测表明,ZLJ的抑制作用是通过使凋亡标志蛋白PARP出现切割,启动凋亡通路实现的。结论中药ZLJ抑制耐药细胞增殖,没有交叉耐药性;其抑制作用与诱导细胞凋亡以及降低P-gp表达有关。     

Reversal of multidrug resistance by morning glory resin glycosides in human breast cancer cells

Reversal of multidrug resistance (MDR) by thirty resin glycosides from the morning glory family (Convolvulaceae) was evaluated in vinblastine-resistant human breast carcinoma cells (MCF-7/Vin). The effects of these amphipathic compounds on the cytotoxicity and P-glycoprotein (P-gp)-mediated MDR were estimated with the sulforhodamine B colorimetric assay. Active noncytotoxic compounds exerted a potentiation effect of vinblastine susceptibility by 1- to over 1906-fold at tested concentrations of 5 and 25 μg/mL. Murucoidin V (1) enhanced vinblastine activity 255-fold when incorporated at 25 μg/mL and also, based on flow cytometry, significantly increased the intracellular accumulation of rhodamine 123 with the use of reserpine as a positive control for a MDR reversal agent. Incubation of MCF-7/Vin cells with 1 caused an increase in uptake and notably lowered the efflux rate of rhodamine 123. Decreased expression of P-glycoprotein by compound 1 was detected by immunofluorescence flow cytometry after incubation with an anti-P-gp monoclonal antibody. These results suggest that resin glycosides represent potential efflux pump inhibitors for overcoming MDR in cancer therapy.     

隐丹参酮对白血病细胞伊马替尼敏感性和P-糖蛋白表达作用的研究

隐丹参酮是丹参中主要的脂溶性活性化合物之一,对白血病等多种恶性肿瘤有显著的抑制作用,并可有效提高伊马替尼的化疗效果,但其发挥功能的分子机制仍未阐明.该文利用白血病细胞K562及其伊马替尼耐药株K562-R,系统研究了隐丹参酮对细胞伊马替尼的敏感性和P-糖蛋白表达的影响.利用MTT法检测隐丹参酮对伊马替尼增殖抑制功能的影响,Annexin V-FITC/PI双标记流式细胞术检测细胞凋亡率的变化,Western blot检测细胞凋亡调控蛋白Caspase-3,Caspase-9,PARP的活性变化情况;梯度浓度的隐丹参酮处理细胞后,Western blot和流式细胞仪检测细胞中P-糖蛋白的表达水平变化,高效液相色谱法检测伊马替尼的胞内药物浓度的变化.结果隐丹参酮预处理后,显著增强了伊马替尼对K562和K562-R细胞的增殖抑制和凋亡诱导作用,细胞的促凋亡蛋白Caspase-3,Caspase-9,PARP的降解产物表达水平显著提高,与对照组相比有显著性差异(P＜0.01).而隐丹参酮对白血病细胞中P-糖蛋白表达和伊马替尼胞内药物积累没有影响.结果表明隐丹参酮可提高CML细胞对伊马替尼的化疗敏感性,其作用机制不依赖于对P-糖蛋白表达的抑制和胞内药物浓度的提高.