硼替佐米对伊马替尼耐药细胞株K562/G01增殖抑制和诱导凋亡作用的研究

 目的　探讨蛋白酶体抑制剂硼替佐米（BOR）对慢性粒细胞白血病（CML）伊马替尼耐药细胞株K562／G01的增殖抑制和诱导凋亡作用。方法　采用MTT法观察细胞的生长抑制效应;流式细胞术（FCM）检测细胞周期与凋亡。结果　K562／G01细胞对伊马替尼不敏感,伊马替尼对K562／G01和K562细胞的IC50分别是（20.09±1.38）、（0.54±0.13）μmol／L;BOR对K562／G01细胞具有增殖抑制作用,并于48 h时作用效果达高峰,IC50（23.10±2.71）nmol／L,随着BOR浓度和作用时间的增加,FCM检测可见G2／M 期细胞周期阻滞及明显的凋亡峰。结论　BOR对CML伊马替尼耐药细胞株具有增殖抑制和诱导凋亡的作用,其机制可能与细胞周期G2／M 期的阻滞有关。     

Experimental Study on the Mechanism of Reversal of Leukemia Multidrug Resistance by Proteasome Inhibitor Bortezomib

OBJECTIVE In this study, we applied multidrug resistant leukemia cell line expressing mdr1-mRNA to observe changes in mdr1-mRNA, the P-gp, cell cycle and apoptosis before and after bortezomib was used, in order to explore the mechanism of reversal of leukemia multidrug resistance by the proteasome inhibitor bortezomib.METHODS Flow cytometry (FCM) was used to detect the intracellular drug concentration, expression of P-gp, cell apoptosis and cell cycle status of K562/DNR cells before and a er treatment with different concentrations of bortezomib. Fluorescence quantitative PCR was applied to detect the mdr1-mRNA expression in K562/DNR and K562/S cells.RESULTS Bortezomib could increase the intracellular DNR content in K562/DNR cells, but showed no e. ect in K562/S cells.5-100 nmol/L bortezomib could significantly reduce the P-gp/mdr1-mRNA expression in K562/DNR cells in vitro, and showed a dose-dependent effect. There was a statistically significant di. erence (P < 0.05) between di. erent concentration groups and the control group. P-gp/mdr1-mRNA expression was negatively correlated with cell apoptosis (r = -0.912 and P < 0.01). After treatment with different concentrations of bortezomib for 24 h,K562/DNR cells in G2 + M phases were significantly increased,while cells in G0 + G1 phases and S phase were significantly decreased, accompanied by an increased apoptotic rate.CONCLUSION Bortezomib can induce G0 + G1 phase to G2 + M phase, and thereby enhance the chemosensitivity of leukemia, and may also reverse the multidrug resistance in leukemia mediated by P-gp overexpression encoded by mdr1 gene. This confi rms that bortezomib can reverse leukemia multidrug resistance at the levels of nucleic acid and protein molecules.     

三氧化二砷联合甲磺酸伊马替尼、咖啡因对K562白血病细胞的协同作用

 目的 探讨三氧化二砷（Arsenic trioxide,As2O3）与酪氨酸激酶抑制剂甲磺酸伊马替尼（实验药物代号STI571）以及细胞周期调节剂咖啡因联合诱导K562细胞凋亡的作用及机制,为寻找克服K562细胞对As2O3抵抗的有效手段提供实验依据。方法 以As2O3与STI571、咖啡因联合作用于K562细胞,采用MTT方法检测细胞增生活性,PI染色流式细胞仪检测细胞周期,PI和Annexin V双染色流式检测细胞凋亡,Western blot法检测细胞周期相关调节蛋白的表达。结果 5.0 μmol／L浓度的咖啡因对K562细胞增生无抑制作用,亦不能增强As2O3的抑制作用,单独以STI571 1.0 μmol／L处理,即可有效抑制K562细胞的生长,与As2O3联用能明显增加其抑制增生的效应;咖啡因与As2O3联合作用,不增加As2O3诱导的K562细胞凋亡率[（14.7±3.6）%vs（15.3±3.3）%,P＞0.05]; STI571具有轻度诱导K562细胞凋亡的作用[（18.3±4.5）%],与As2O3合用可显著增加诱导凋亡率[（14.7±3.6）%vs（42.8±4.2）%,P＜0.01],并明显降低As2O3诱导的G2／M期细胞比例。与单用As2O3比较,As2O3 + STI571明显抑制cdc2、cdc2-p及survivin蛋白表达,而As2O3与咖啡因合用不能诱导survivin蛋白表达下调,对cdc2、cdc2-p蛋白的表达无明显影响。结论 周期调节药物咖啡因对As2O3诱导K562细胞凋亡无增敏作用;酪氨酸激酶抑制剂STI571能协同As2O3诱导K562细胞凋亡,下调抗凋亡蛋白survivin的表达可能是其机制之一,值得深入研究其临床应用效果。     

反义寡核苷酸对K562细胞增殖和端粒酶活性的影响

 目的　研究靶向封闭端粒酶反转录酶（hTERT）mRNA的反义硫代寡核苷酸（ASPSODN）对K562细胞目的基因的抑制及其对端粒酶活性及细胞增殖周期和凋亡的影响 。方法　ASPSODN转染到人类红白血病细胞株K562,采用MTT法、酶联免疫吸附（ELISA）法和流式细胞术（FCM）检测K562细胞的增殖、端粒酶活性、细胞凋亡和细胞周期的改变,RT-PCR检测hTERT mRNA的表达。结果　0.6 μmol／L的ASPSODN（0.42±0.16）能明显下调hTERT表达（P＜0.05）,端粒酶相对活性降至52 %;MTT法检测显示明显抑制K562细胞增殖活性;FCM显示细胞凋亡率为10.31 %,PI染色显示细胞被阻止在G1／G0期,S期及G2／M期的细胞减少,但无特征性的凋亡峰。结论　ASPSODN靶向 hTERT 能特异性抑制K562细胞hTERT mRNA的表达,明显下调端粒酶活性,抑制K562细胞增殖,并通过降低细胞的端粒酶活性而诱发细胞凋亡。     

Anti-Proliferative Effects of Dendrophthoe pentandra Methanol Extract on BCR/ABL-Positive and Imatinib-Resistant Leukemia Cell Lines

Background: Imatinib mesylate, a tyrosine kinase inhibitor specifically targeting the BCR/ABL fusion protein, induces hematological remission in patients with chronic myeloid leukemia (CML). However, the majority of CML patients treated with imatinib develop resistance with prolonged therapy. Dendrophthoe pentandra (L.) Miq. is a Malaysian mistletoe species that has been used as a traditional treatment for several ailments such as smallpox, ulcers, and cancers. Methods: We developed a resistant cell line (designated as K562R) by long-term co-culture of a BCR/ ABL positive CML cell line, K562, with imatinib mesylate. We then investigated the anti-proliferative effects of D. pentandra methanol extract on parental K562 and resistant K562R cells. Trypan blue exclusion assays were performed to determine the IC50 concentration; apoptosis and cell cycle analysis were conducted by flow cytometry. Results: D. pentandra extract had greater anti-proliferative effects towards K562R (IC50= 192 μg/mL) compared to K562 (500 μg/ mL) cells. Upon treatment with D. pentandra extract at the IC50. concentration: K562 but not K562R demonstrated increase in apoptosis and cell cycle arrest in the G2/M phase. Conclusion: D. pentandra methanol extract exerts potent anti-proliferative effect on BCR/ABL positive K562 cells.     

土木香内酯通过调节细胞周期相关蛋白的表达抑制慢性粒细胞白血病耐药细胞株K562/ADR增殖

目的 研究土木香内酯对慢性粒细胞白血病(CML)耐药细胞株K562/ADR增殖、周期分布以及周期相关蛋白的影响.方法 选用0、1.0、2.0、4.0、6.0、8.0、10.0μmol/L土木香内酯作用K562/ADR细胞12、24、48 h,四甲基偶氮唑蓝(MTT)比色法检测土木香内酯对K562/ADR细胞的增殖抑制率,流式细胞术检测土木香内酯对K562/ADR细胞周期分布的影响,Western blot检测周期相关蛋白的表达.结果 土木香内酯能够显著抑制K562/ADR细胞增殖,半数抑制浓度为5.0 μmol/L.流式细胞术检测结果显示,不同浓度(0、2.5、5.0、7.5 μmol/L)的土木香内酯能够使K562/ADR细胞周期阻滞在G2/M期,G2/M期细胞比例由(15.8±1.7)％分别提高到(21.0±2.4)％、(26.4±2.7)％、(30.1±3.9)％(P＜0.05).土木香内酯能够明显降低CDK1、CyclinB1的表达,上调周期抑制蛋白p21的表达.此外,土木香内酯能够有效地抑制bcr-abl融合蛋白的表达水平.结论 土木香内酯可能通过调节细胞周期相关蛋白的表达介导K562/ADR细胞G2/M期阻滞,抑制细胞的增殖.     

The synergistic reversal effect of multidrug resistance by quercetin and hyperthermia in doxorubicin-resistant human myelogenous leukemia cells

PURPOSE: This study aimed to evaluate the multidrug resistance (MDR) reversal activity of quercetin (Que) in combination with hyperthermia (HT) in human myelogenous leukemia cells K562/A. METHODS: The cytotoxicity of Que alone and the effect of Que and HT to doxorubicin (Dox) cytotoxicity were determined using MTT assay in K562 and K562/A cells. K562/A cells was heated with or without Que pretreatment, and the protein and mRNA levels of heat shock protein 70 (HSP70) and P-glycoprotein (P-gp) were determined by flow cytometry (FCM) and RT-PCR, respectively. Intracellular accumulation of Dox, cell cycle and apoptosis were monitored with FCM. RESULTS: Que alone inhibited cell growth in a dose-dependent manner in K562 and K562/A cells. Either Que or HT alone had a weak reversal effect on Dox resistance, however, combination HT and Que showed a much more significant reversal effect on Dox resistance (reverse fold 9.49). The elevated protein expression and mRNA level of HSP70 and P-gp in response to HT were inhibited by Que. Pretreatment with Que caused the cells to accumulate Dox 8.3-fold higher than in control cells. In addition, Que induced apoptosis and G2/M arrest in a dose-dependent manner, and the combination of Que and HT was found to have a synergistic effect on apoptosis. CONCLUSIONS: Que pretreatment could significantly enhance the MDR reversal activity of HT in resistant cell line, by sensitizing the cell to reversing MDR activity of HT.     

柚皮素对人类白血病K562细胞生长的作用及其机制探讨

 目的 研究柚皮素（naringenin）对人类慢性髓系白血病K562细胞株的作用及其机制。方法 体外培养K562细胞,MTT法检测柚皮素在不同浓度、不同时间点对K562细胞生长的影响;用免疫组织化学的方法检测细胞增殖核抗原（PCNA）的表达,并计算PCNA标记指数（LI）;显微镜和透射电镜下观察细胞形态变化;流式细胞仪分析细胞周期分布和细胞凋亡率;用RT-PCR和Western blot的方法检测K562细胞中p53、p21WAF1 mRNA和蛋白表达的变化。结果 柚皮素对K562细胞增生有明显抑制作用,作用24、48和72 h后的半数抑制浓度（IC50）分别为455 μmol／L、225 μmol／L和175 μmol／L;PCNA在柚皮素实验组的表达显著低于对照组;普通光镜以及电镜观察均见实验组细胞显著的增生抑制和典型的细胞凋亡形态学改变;流式细胞仪分析证实柚皮素能使K562细胞聚积在G0／G1期、S期细胞减少,凋亡率增加;RT-PCR和Western-blot检测表明,实验组细胞表达p21 mRNA和蛋白质呈浓度和时间依赖性升高,而p53变化不明显。结论 柚皮素对K562细胞具有明显的增生抑制作用,细胞周期阻滞和诱导凋亡可能是其重要机制,而非p53依赖性的p21上调可能是其发挥作用的重要途径。     

重楼皂苷D对人慢性粒细胞白血病K562细胞增殖的抑制作用及其机制

目的 探讨重楼皂苷D对人慢性粒细胞白血病细胞株K562增殖的抑制作用及其机制.方法 选用浓度为0、0.1、0.2、0.4、0.8、1.2、2.4μmol/L重楼皂苷D作用于K562细胞24 h,用CCK-8法检测重楼皂苷D对K562细胞增殖的抑制作用;流式细胞术检测重楼皂苷D对K562细胞凋亡以及细胞周期分布的影响;Western blot方法检测相关蛋白的表达.结果 重楼皂苷D可显著抑制K562细胞增殖,24 h的半数有效抑制浓度(IC50)为(0.9±0.1)μmol/L.流式细胞术检测结果显示,浓度为0.9μmol/L重楼皂苷D作用于K562细胞12、24 h后,细胞早期凋亡率较对照组的(2.05±0.45)%分别提高到(11.46±1.51)%、(28.87±2.35)%,差异有统计学意义(F=38.637,P<0.05).重楼皂苷D能够显著下调bcl-2、CDK1、cyclin B1、bcr-abl融合蛋白的表达,上调Bax、细胞色素C、活化的caspase-3以及p21的表达(均P<0.05).此外,重楼皂苷D能够将细胞周期阻滞在G2/M期(F=42.355,P<0.05).结论 重楼皂苷D能够明显抑制慢性粒细胞白血病K562细胞增殖,其作用机制可能是诱导细胞凋亡及促进细胞周期阻滞.     

科罗索酸对白血病K562细胞增殖和凋亡的影响

目的:探究科罗索酸（corosolic acid）对白血病K562细胞增殖和凋亡的影响。方法:使用不同浓度的科罗索酸处理K562细胞,CCK-8法检测对细胞增殖的影响,流式细胞术检测对细胞周期和凋亡的影响,分光光度法检测对Caspase-3/7,-8,-9活性的影响。结果:科罗索酸能显著抑制白血病K562细胞的增殖,并且这种抑制作用随着科罗索酸浓度升高和培养时间延长而显著增强。科罗索酸处理后的K562细胞中,处于G0/G1期的细胞比率明显升高（P＜0.05）,S期和G2/M期的细胞明显减少（P＜0.05）,凋亡细胞明显增多（P＜0.05）,Caspase-3/7,-8和-9的活性明显增强。结论:科罗索酸能抑制白血病K562细胞的增殖,促进其凋亡,这一效应可能与Caspase活性的增强有关。     

丹参酮ⅡA抑制髓系白血病细胞株增殖及诱导凋亡的研究

目的 探讨丹参酮ⅡA（TanⅡA）对髓系白血病细胞株NB4、K562、THP-1增殖抑制作用及促凋亡作用.方法 将不同浓度TanⅡA与NB4、K562、THP-1细胞共培养24、48、72 h,以柔红霉素为阳性对照,采用四甲基偶氮唑蓝（MTT）法检测TanⅡA对白血病细胞株的增殖抑制作用,AnnexinV-FITC/PI流式细胞术检测细胞凋亡和细胞周期,紫外分光光度法检测Caspase-3蛋白表达.结果 TanⅡA作用24、48、72 h对NB4细胞的IC50值分别为24.11、9.60、7.28 μmol/L,对K562细胞IC50值分别为31.75、11.88、6.81 μmol/L,对THP-1细胞IC50值分别为111.10、32.82、11.82 μmol/L.流式细胞术检测结果显示：与空白对照组相比,TanⅡA与各白血病细胞株作用48 h后,细胞凋亡明显增加,G1期细胞数增加,Caspase-3蛋白表达显著升高（P＜0.05）.结论 TanⅡA对白血病细胞具有增殖抑制与促凋亡作用,其作用强度从高到低依次为NB4、K562、THP-1细胞.其抗癌作用机制可能与阻滞细胞于G1期,上调Caspase-3表达有关.     

FGFR抑制剂BGJ398对白血病细胞生物学行为及耐药的影响

背景与目的:成纤维细胞生长因子(fibroblast growth factor,FGF)家族成员能够调节细胞增殖、迁移和分化,在肿瘤性疾病的发生和转移等过程中发挥重要作用,FGF受体(FGF receptor,FGFR)抑制剂具有抗肿瘤活性.探讨FGFR抑制剂BGJ398对白血病耐药细胞株K562/ADM增殖、迁移、...     

RNA干扰survivin基因对卵巢癌细胞生长凋亡及药物敏感性的影响

目的 探讨下调survivin基因对卵巢癌顺铂(DDP)耐药细胞株SKOV3/DDP细胞生长、凋亡及药物敏感性的影响.方法 构建survivin基因的小干扰RNA(siRNA)表达载体pSilencer-survivin,用脂质体方法转染SKOV3/DDP细胞株,另设未转染组和转染pSilencer-control组作为对照.显微镜下观察转染前后细胞的变化,逆转录聚合酶链反应(RT-PCR)和Western blot分别检测survivinmRNA及蛋白的表达,二苯基溴化四氮唑蓝(MTT)法检测细胞生长情况,流式细胞仪检测细胞凋亡及周期的变化.各组细胞加入DDP,检测其对DDP药物敏感性的影响.结果 survivin siRNA可明显下调SKOV3/DDP细胞中survivin mRNA及蛋白表达水平.SKOV3/DDP细胞生长受到明显抑制,生长曲线低平.转染48 h后,转染pSilencer-survivin组细胞凋亡率为19.1%,明显高于末转染组(2.6%)和转染pSilencer-control组(3.5%),G1/G0期细胞所占的比例增高,而G2/M期细胞所占的比例降低.转染pSilencer-survivin组细胞的IC50明显降低,为1.16 μg/mi.结论 survivin siRNA可下调SKOV3/DDP细胞中survivin的表达,使细胞生长减慢,凋亡增加,对DDP的药物敏感性增强.     

The synergistic reversal effect of multidrug resistance by quercetin and hyperthermia in doxorubicin-resistant human myelogenous leukemia cells

Purpose: This study aimed to evaluate the multidrug resistance (MDR) reversal activity of quercetin (Que) in combination with hyperthermia (HT) in human myelogenous leukemia cells K562/A.

Methods: The cytotoxicity of Que alone and the effect of Que and HT to doxorubicin (Dox) cytotoxicity were determined using MTT assay in K562 and K562/A cells. K562/A cells was heated with or without Que pretreatment, and the protein and mRNA levels of heat shock protein 70 (HSP70) and P-glycoprotein (P-gp) were determined by flow cytometry (FCM) and RT-PCR, respectively. Intracellular accumulation of Dox, cell cycle and apoptosis were monitored with FCM.

Results: Que alone inhibited cell growth in a dose-dependent manner in K562 and K562/A cells. Either Que or HT alone had a weak reversal effect on Dox resistance, however, combination HT and Que showed a much more significant reversal effect on Dox resistance (reverse fold 9.49). The elevated protein expression and mRNA level of HSP70 and P-gp in response to HT were inhibited by Que. Pretreatment with Que caused the cells to accumulate Dox 8.3-fold higher than in control cells. In addition, Que induced apoptosis and G2/M arrest in a dose-dependent manner, and the combination of Que and HT was found to have a synergistic efeect on apoptosis.

Conclusions: Que pretreatment could significantly inhance the MDR reversal activity of HT in resistant cell line, by sensitizing the cell to reversing MDR activity of HT.     

葛根总黄酮诱导白血病细胞株凋亡及其分子机制的研究

 目的　探讨葛根总黄酮（PR）对慢性粒细胞白血病（CML）细胞株K562和急性早幼粒细胞白血病（APL）细胞株NB4细胞增殖及凋亡的影响。方法　采用MTT法检测PR对K562细胞、NB4细胞的增殖抑制率;光学显微镜及荧光显微镜观察细胞形态改变;Hoechest33258荧光染色AnnexinV／PI双染法检测细胞凋亡率;DNA PI染色法分析细胞周期及亚二倍体峰。Western blot分别检测NB4细胞JNK、PARP、bcl-2、Caspase3,K562细胞bcr-abl、p53、bcl-2、Fas／FasL蛋白表达的变化。结果　12.5～200 μg／ml PR均能抑制K562、NB4细胞增殖。光学显微镜及荧光显微镜下观察到核固缩、凋亡小体等典型的细胞凋亡改变;Annexin V+／PI-细胞呈时间-剂量依赖性增加;DNA PI染色法发现细胞亚二倍体比例增加,G1期比例下降、S期比例增加。PR呈时间-剂量依赖性抑制K562细胞、NB4细胞增殖,诱导细胞凋亡。不同浓度PR干预后K562细胞bcr-abl蛋白水平呈浓度依赖性下调（F＝18.74,P＜0.05）,而bcl-2则无明显变化;p53 表达呈浓度依赖性上调;Fas／FasL 表达无明显变化。NB4细胞JNK、PARP及Caspase 3蛋白表达与PR浓度呈正相关,与凋亡抑制蛋白bcl-2则呈负相关（F＝42.32,P＜0.05）。结论　PR能有效抑制K562、NB4细胞增殖,阻滞细胞周期进程,诱导细胞凋亡,但分子机制不同。提示一定浓度PR具有较广谱的抗白血病效应。     

去甲斑蝥素对白血病细胞系K562的增殖、凋亡、细胞周期及NF-κB活性的影响

目的 探讨去甲斑蝥素（NCTD）对白血病细胞系K562的增殖、凋亡、细胞周期及核因子（NF）-κB活性的影响。方法 采用CCK-8试剂盒检测不同浓度NCTD（0、10、25、50、100 μmol／L）处理K562细胞24、48、72、96 h后的增殖抑制率,采用流式细胞术检测不同浓度NCTD处理24、48 h后的细胞凋亡率和处理48 h后的细胞周期,采用原位细胞凋亡检测试剂盒（Tunnel）检测不同浓度NCTD处理24、48 h后的细胞凋亡指数,采用Western blotting检测不同浓度NCTD处理48 h后的NF-κB p65及IκB α水平以评价NF-κB活性的变化。结果 在10～100 μmol／L范围内,NCTD可呈时间和浓度依赖的方式升高K562细胞的增殖抑制率,各浓度间的差异有统计学意义（P＜0.05）;与0 μmol／L相比,其余各浓度处理24、48 h后的凋亡率和凋亡指数均升高,处理48 h后的G0／G1期细胞比例升高,S、G2／M期细胞比例均降低,NF-κB p65水平降低,IκB-α水平升高,以上差异均有统计学意义（P＜0.05）。结论 NCTD对白血病细胞系K562有细胞毒性,可抑制该细胞的增殖、诱导其凋亡及细胞周期阻滞并下调NF-κB表达。     

Overexpression of glucosylceramide synthase in associated with multidrug resistance of leukemia cells

Ceramide, as a second messenger, initiates one of the major signal transduction pathways in tumor apoptosis. Glucosylceramide synthase (GCS) catalyzes glycosylation of ceramide and produces glucosylceramide. Through GCS, ceramide glycosylation allows cellular escape from ceramide-induced programmed cell death. Here we investigated the expression of GCS in human leukemia cells and an association between GCS and multidrug resistance of leukemia cells. Using RT-PCR technique the level of GCS gene was detected in 65 clinical multidrug resistance/non-resistance cases with leukemia, and in K562 and K562/A02 cell lines. AlamarBlue Assay was applied to confirm the multidrug resistant of K562/A02 cells. PPMP, which is a chemical inhibitor for GCS, was used to determine the relationship between GCS and drug-resistance in K562/A02 cells. In addition, multidrug resistance gene (mdr1), Bcl-2 and Bax mRNA was also analyzed by RT-PCR. The expression of GCS and mdr1 mRNA in clinic multidrug resistance samples exhibited significantly increased compared with clinic drug sensitive group (P<0.05). There was the positive correlation both the expression of GCS and mdr1 genes in leukemia samples (P<0.01, gamma=0.7). AlamarBlue Assay showed that the K562/A02 cell line was 115-fold more resistant to adriamycin and 36-fold more resistant to vincristine compared with drug-sensitive K562 cell line. There also was significant expression difference of GCS and mdr1 genes between K562 and K562/A02 cells. Bcl-2 gene exhibited higher expressions whatever in clinic drug-resistance samples or K562/A02 cells, whereas the expressions of Bax gene were higher in drug-sensitive samples and K562 cells. PPMP increased sensitivity to adriamycin toxicity by inhibiting GCS in K562/A02 cells. Therefore, it is suggested that a high level of GCS in leukemia is possible contributed to multidrug resistance of leukemia cells. Abnormally expressions of the genes in associated with cell apoptosis might be one of the main molecular pathology mechanisms of multidrug resistance caused by GCS gene.     

过表达DACT1对白血病K562细胞生物学行为的影响及机制

目的:研究过表达DACT1对白血病K562细胞增殖、凋亡及细胞周期的影响,并探讨其作用机制。方法:用Attractene Transfection试剂在K562细胞中转染DACT1质粒,使其在细胞中过表达,应用CCK-8法、流式细胞术检测过表达DACT1对K562细胞增殖、凋亡及细胞周期的影响。Western blot法检测过表达DACT1后凋亡相关蛋白的变化。结果:过表达DACT1能够抑制K562细胞集落形成,抑制细胞增殖,促进细胞凋亡,使细胞发生G0/G1期阻滞。同时可使促凋亡蛋白Bax、caspase-3及caspase-9表达增加,抗凋亡蛋白Bcl-2表达减少。结论:过表达DACT1可以抑制白血病K562细胞增殖并诱导其凋亡,干扰其增殖周期,可能具有潜在的抑癌作用,其机制可能与调节凋亡相关蛋白有关。     

Berberine-induced apoptosis via decreasing the survivin protein in K562 cell line

Berberine is an isoquinoline alkaloid with multiple pharmacological activities, including anti-inflammatory and anti-diarrhea effect, the induction of apoptosis and anti-cancer effect. It has been reported that berberine exerts its anti-inflammatory effect via suppressing nuclear factor-kappa B (NF-κB) expression. Survivin and inducible nitric oxide synthase (iNOS) proteins may contribute to the causal relationship between anti-inflammatory and anti-apoptotic function. To investigate the mechanism of berberine-induced apoptotic activities, the human erythro-myeloblastoid leukemia cell line (K562 cell line) was treated with different concentrations of berberine (25–100 μM). The most significant cellular growth arrest and apoptotic effects were observed in the cells treated with 75 μM of berberine for 72 h. The results indicate that survivin and iNOS protein levels were decreased in berberine-treated cells. However, decrease in the iNOS activity did not affect the cell growth and apoptosis. Moreover, the addition of NO donor, sodium nitroprusside, to culture medium decreased the cell growth in the present cell line, but it seemed that its concentration was too low to induce apoptosis. So despite its production by iNOS in untreated cells, NO does not play a significant role in carcinogenesis in this cell line. These results indicate that the apoptotic activity of berberine may be mediated through the reduction of survivin in K562 cells, but iNOS level and its activity does not play a significant role in berberine-induced apoptosis.