雌马酚对映异构体影响前列腺癌PC3、DU145细胞侵袭性研究

目的比较雌马酚对映异构体R-(+)雌马酚与S-(-)雌马酚对人前列腺癌PC3、DU145细胞侵袭性的影响,并分析相关机制。方法分别采用细胞划痕实验、侵袭实验检测癌细胞的运动能力与侵袭性的变化,采用明胶酶谱法、RT-PCR法检测相关蛋白活性和基因表达。结果 R-(+)雌马酚与S-(-)雌马酚均可显著抑制PC3、DU145细胞的运动能力及侵袭性,其中抑制PC3细胞运动能力及侵袭能力的作用较强,两种化合物比较,R-(+)雌马酚的作用更强。R-(+)雌马酚与S-(-)雌马酚还可显著抑制侵袭所必需的蛋白水解酶基质金属蛋白酶家族中MMP-2、MMP-9的活性。R-(+)雌马酚可显著增加PC3细胞ER-α的表达,对ER-β则无明显作用。结论 R-(+)雌马酚与S-(-)雌马酚可显著抑制PC3、DU145细胞运动能力及侵袭性。其中R-(+)雌马酚对PC3细胞的抗侵袭作用最强,可能由ER-α介导。     

沉默信息调节因子1小干扰RNA诱导前列腺癌细胞PC3细胞凋亡

摘要:目的　观察沉默信息调节因子1（SIRT1）小干扰RNA（siRNA）对前列腺癌细胞PC3细胞生长增殖、DNA合成、细胞凋亡和Bcl-2和Bax蛋白的表达变化,探讨SIRT1在前列腺癌发生中的可能机制。方法　体外培养PC3细胞,分空白对照组（mock组）,转染阴性对照组（scramble siRNA组）和SIRT1 siRNA转染组;Western blot检测PC3细胞中SIRT1的干涉效能;MTT法测定PC3细胞的增殖率;BrdU掺入法测定DNA合成;流式细胞术检测细胞凋亡;Western blot检测PC3细胞中细胞凋亡关键调控因子Bcl-2和Bax的蛋白表达。结果　与对照组比较,SIRT1 siRNA组SIRT1蛋白表达降低（P＜0.01）,PC3细胞的增殖和DNA合成明显受抑制（P＜0.01）,细胞凋亡比例增加（P＜0.01）,Bcl-2蛋白表达减少,Bax的表达增加。结论　下调SIRT1的表达抑制细胞增殖和DNA合成,诱导前列腺癌PC3细胞发生凋亡,其机制可能与改变细胞凋亡关键调控因子Bcl-2和Bax的蛋白表达相关。     

染料木素对前列腺癌细胞恶性增殖的抑制作用

目的研究染料木素（Gen）对人前列腺癌细胞（PC-3）的影响。方法将PC-3细胞在DMEF-12培养液（含10％胎牛血清）中采用开放式单层贴壁培养，采用四甲基偶氮噻唑蓝（MTT）比色法、细胞核分裂指数、集落形成和^3H-TdR掺入试验方法进行检测。染料木素剂量设为10，20，40，80μmol／L。以二甲基亚砜（DMSO）为溶剂对照。结果不同剂量的染料木素对PC-3细胞的生长增殖有不同的抑制作用。在MTT试验中，不同浓度染料木素对PC-3细胞的7d抑制率分别为27．7％，58．8％，72．3％，84．4％。在核分裂指数试验中，随着染料木素剂量的增高，其核分裂指数明显降低；遮集落形成试验中，随着染料木素剂量的增加，集落形成抑制作用增加；在^3H-TdR掺入试验中，可见随着染料木素剂量的增高，^3H-TdR掺入到PC-3细胞中明显的减少，与阴性对照组比较差异有统计学意义。结论染料木素对PC-3细胞的生长增殖有明显抑制作用，可能是染料木素抑制前列腺癌的机制之一。     

Cell killing and radiosensitization by caffeic acid phenethyl ester (CAPE) in lung cancer cells

Caffeic acid phenethyl ester (CAPE) is a biologically active ingredient of honeybee propoplis. The cytotoxicity and radiation sensitization effects of CAPE were evaluated in human lung cancer A549 cells and normal lung fibroblast WI-38 cells. A549 cells treated with 6 microg/ml CAPE showed marked growth inhibition (60%) at 48 hr after treatments. During the same time, the number of viable cells decreased to 46% of the control value. In contrast, WI-38 cells showed 20% growth inhibition with no change in the number of viable cells under the same treatment conditions. At 72 hr after CAPE treatment (6 microg/ml), the percentage of apoptotic cells in A549 cultures increased significantly to 67% and an S/G2 arrest was also detected in the culture. Furthermore, there was a significant decrease in the level of intracellular glutathione and hydrogen peroxide contents within one hr after CAPE treatment, and the expression of cyclin B1 was reduced 6 hr after treatment. The radiation sensitization effect of CAPE on A549 cells was determined from the clonogenic survival curves, and the results showed a small but significant difference in radiation survival between cells treated with or without CAPE. Taken together, our results suggest that the effects of CAPE on differential cytotoxicity, apoptosis, and radiosensitization are associated with glutathione depletion that occurred shortly after treatments.     

蒿甲醚对人结直肠癌细胞的毒性和放疗增敏作用的 研究

摘要:目的　探讨蒿甲醚对人结直肠癌细胞株HCT116及HT29的细胞毒性和放射增敏作用。方法　采用MTT法检测蒿甲醚对人结直肠癌细胞株HCT116及HT29的细胞毒性。克隆形成实验检测蒿甲醚对人结直肠癌细胞株HCT116及HT29的放射增敏作用,多靶单击模型拟合HCT116及HT29细胞的剂量存活曲线,计算蒿甲醚对HCT116及HT29细胞的放射增敏比,评价其放射增敏效果。结果　蒿甲醚对HCT116及HT29细胞作用24 h、48 h、72 h的IC50分别为:HCT116:300 μg/ml、270 μg/ml、170 μg/ml;HT29:280 μg/ml、250 μg/ml、200 μg/ml。蒿甲醚对HCT116及HT29细胞在平均致死剂量的放射增敏比（SERD0）分别为1.31和1.25。结论　蒿甲醚对人结直肠癌细胞株HCT116及HT29的细胞毒性作用呈剂量和时间依赖性,并可对人结直肠癌细胞系HCT116及HT29有放射增敏作用。     

Cell cycle arrest and induction of apoptosis by lycopene in LNCaP human prostate cancer cells

Lycopene is one of the major carotenoids and is found almost exclusively in tomatoes and tomato products. Since tomato consumption is associated with decreased risk of prostate cancer, characterizing the effects of lycopene on cell growth or survival, cell cycle progression, and apoptosis in LCNaP human prostate cancer cells might elucidate the mechanisms of actions of lycopene. To discover the possible anti-cancer mechanism of lycopene, water-soluble lycopene was used, and cell cycle arrest and apoptosis were measured. Placebo formulation at each lycopene dose at 0.1, 1, and 5 microM was used as a control. After 6, 24, and 48 hours of incubation, cells were harvested and measured for cell viability. Lycopene at 1 microM inhibited cell growth by 31%, compared with its placebo formulation after a 48-hour incubation. Lycopene at 5 microM increased the number of cells in the G(2)/M phase of the cell cycle from 13% to 28% and decreased S-phase cells from 45% to 29%, while no shifts in cell cycle were detected in placebo-treated groups. Apoptosis was observed at the 5 microM lycopene formulation at the late stages during the 24- and 48-hour treatments. Lycopene, therefore, deserves further study as a potential chemopreventive/chemotherapeutic agent.     

胡桃醌对人前列腺癌PC-3细胞抑制作用

目的探讨胡桃醌对前列腺癌PC-3细胞的抑制作用。方法溴化四氮唑蓝(MTT)法检测不同剂量胡桃醌对PC-3细胞生长抑制影响;采用Annexin V-FITC/PI染色实验,流式细胞仪检测PC-3细胞凋亡情况;蛋白印迹法检测B细胞淋巴瘤2(Bcl-2)蛋白和Bcl-2相关 X 蛋白(Bax)的表达量。结果与对照组比较,12.5、25、50和100 μmol/L胡桃醌对前列腺癌PC-3细胞具有抑制作用且呈剂量依赖性,抑制率分别为20.6%、39.7%、57.3%和66.2%;流式细胞仪检测胡桃醌(≥ 25 μmol/L)作用细胞后诱导PC-3细胞发生早期和晚期凋亡,当胡桃醌浓度为25、50和100 μmol/L时其早、晚期凋亡率分别是7.37%和2.07%、11.03%和5.8%及18.62%和8.54%,均高于对照组诱导的细胞凋亡率(P<0.05);随着胡桃醌浓度(≥ 50 μmol/L)不断增加,可上调Bax并下调Bcl-2蛋白的表达(P<0.05)。结论胡桃醌对前列腺癌PC-3细胞具有细胞毒作用,抑制细胞生长。     

Apoptotic effect of IP6 was not enhanced by co-treatment with myo-inositol in prostate carcinoma PC3 cells

Inositol hexaphosphate (IP₆) is a major constituent of most cereals, legumes, nuts, oil seeds and soybean. Previous studies reported the anticancer effect of IP₆ and suggested that co-treatment of IP₆ with inositol may enhance anticancer effect of IP₆. Although the anticancer effect of IP₆ has been intensively studied, the combinational effect of IP₆ and inositol and involved mechanisms are not well understood so far. In the present study, we investigated the effect of IP₆ and myo-inositol (MI) on cell cycle regulation and apoptosis using PC3 prostate cancer cell lines. When cells were co-treated with IP₆ and MI, the extent of cell growth inhibition was significantly increased than that by IP₆ alone. To identify the effect of IP₆ and MI on apoptosis, the activity of caspase-3 was measured. The caspase-3 activity was significantly increased when cells were treated with either IP₆ alone or both IP₆ and MI, with no significant enhancement by co-treatment. To investigate the effect of IP₆ and MI of cell cycle arrest, we measured p21 mRNA expression in PC3 cells and observed significant increase in p21 mRNA by IP₆. But synergistic regulation by co-treatment with IP₆ and MI was not observed. In addition, there was no significant effect by co-treatment compared to IP₆ treatment on the regulation of cell cycle progression although IP₆ significantly changed cell cycle distribution in the presence of MI or not. Therefore, these findings support that IP₆ has anticancer function by induction of apoptosis and regulation of cell cycle. However, synergistic effect by MI on cell cycle regulation and apoptosis was not observed in PC3 prostate cancer cells.     

Red raspberries have antioxidant effects that play a minor role in the killing of stomach and colon cancer cells

Berries and berry extracts possess properties that make them important in the prevention of cancer. The high antioxidant levels of these extracts play a role, but components of the berries can have other effects on cell replication and survival. We chose to test the hypothesis that (i) although the antioxidant capacity of raspberry extracts is important for inhibiting the proliferation of tumor cells, other characteristics of the berry extracts are responsible for a major part of their antiproliferative activity, and that (ii) the relative importance of the antioxidant effect can depend on the cell type being studied. The aim of this study was to assess the relative roles of low pH and high antioxidant levels in the killing of 3 cell types by an aqueous extract from Meeker red raspberries. Stomach, colon, and breast cancer cells were treated with berry extract and with HCl and ascorbic acid solutions of the same pH. A dilution of 7.5% ascorbic acid solution, of the same pH and slightly higher antioxidant concentration than the berry extract, killed less than 10% of the stomach and colon cancer cells. In contrast, the berry extract at this same dilution killed more than 90% of these cells. Antioxidants played a more significant role in the killing of breast cancer cells, however. For these cells, approximately 50% of the killing could be attributed to antioxidant effects. We conclude that the antioxidant effect plays a minor role in the killing of 2 gastrointestinal cell types, but its role in inactivating a breast cancer cell line is much more significant. No evidence of apoptosis was observed, and caspase activation did not contribute to cell killing by the extract.     

锰对多巴胺能神经细胞PC12的毒性及其机制研究

为探讨锰 (Mn)抑制神经细胞增殖的机制 ,体外培养神经细胞株PC12 ,培养基分别含有 10 0、2 0 0及5 0 0mmol LMnCl2 ,作用 2 4、48、72h后 ,用四唑盐比色实验 (MTT)和平板克隆形成实验检测细胞的存活和生长 ;台盼蓝拒染法绘制生长曲线 ;DTNB法测定谷胱甘肽过氧化物酶 (GSH -Px)的活性、丙二醛比色法测定丙二醛 (MDA)的含量 ;DNA凝胶电泳检测神经细胞凋亡。结果显示 ,MTT和平板克隆形成实验检测结果显示10 0～ 5 0 0mmol L的Mn均对PC12细胞株有显著的抑制作用 ,且呈剂量 效应关系。GSH Px活性降低 ;MDA的活性升高。DNA凝胶电泳结果显示锰能够诱导PC12细胞凋亡。提示锰对神经细胞PC12的增殖具有显著的抑制作用 ,其机制是通过降低过氧化物酶的活性、干扰神经细胞的DNA代谢和诱导神经细胞凋亡     

曲古抑菌素A阻断Stat3信号通路诱导前列腺癌细胞凋亡的实验研究

目的本实验研究组蛋白去乙酰化酶抑制剂曲古抑菌素A(TSA)对前列腺癌细胞DU145的抑制作用及其对信号传导与转录激活因子3(Stat3)信号通路的影响。方法采用不同浓度的TSA处理DU145不同时间后,四氮甲基唑蓝比色法测定TSA对细胞活力的抑制效应;流式细胞仪分析细胞周期的改变;蛋白印迹实验检测细胞凋亡相关蛋白半胱氨酸蛋白酶(Caspase)家族的Caspase-8、Caspase-9、Caspase-3、二磷酸腺苷核糖多聚酶(PARP)及Stat3信号蛋白活性(phospho-Stat3)的变化。结果 TSA时间和剂量依赖性地抑制DU145细胞的增殖,TSA处理细胞24 h和48 h后,细胞生存率分别是85.7%和68.7%;细胞经TSA处理后,细胞形态和细胞周期均发生明显的变化,细胞周期被阻断在G0/G1期,其细胞百分比从55.6%增加到68.5%;Western blot检测结果显示,TSA作用DU145细胞后,Stat3信号蛋白的磷酸化水平下降,同时IL-6对Stat3的刺激诱导作用也被TSA所阻断;Caspase-8、Caspase-9、Caspase-3、PARP等凋亡蛋白被TSA诱导活化,并发生显著剪切。结论 TSA能够通过抑制Stat3信号通路的活性来诱导DU145细胞凋亡。     

STAT3 as a target for sensitizing prostate cancer cells to irradiation

Radioresistance of prostate cancer (PCa) is a major factor leading to local failure of radiotherapy. STAT3 is an oncogenic protein that was recently found to be activated in PCa tumors. This study aimed to investigate the radiosensitization effect of targeting STAT3 in PCa tumors. Here, the radiosensitization effect of STAT3 blockade was investigated by clonogenic assay, flow cytometry and western blot analysis in human PCa cells in vitro and in vivo. We demonstrated that STAT3 blockade with a STAT3 inhibitor or siRNA increased the radiosensitivity of PCa cells and that radiation together with STAT3 blockade induced more apoptosis and double-strand breaks (DSBs) than radiation alone in LNCaP cells. In addition, radiation induced STAT3 activation and survivin expression in PCa cells, which was inhibited by STAT3 blockade. Transfection with survivin cDNA attenuated the radiosensitization effect of STAT3 blockade. These effects were further confirmed by in vivo studies, which showed that the STAT3 inhibitor enhanced the treatment efficacy of radiation on LNCaP xenografts with decreased STAT3 activation and survivin expression.These findings suggest that STAT3 blockade radiosensitizes PCa cells through regulation of survivin. Thus, our study has revealed STAT3 as a potential sensitizer for irradiation in PCa cells. Its clinical application as an adjuvant in radiotherapy of PCa should be explored in the future.     

Reevaluation of the radiosensitizing effects of sanazole and nimorazole in vitro and in vivo

Sanazole (AK-2123, 3-nitrotriazole derivative, N1-(3-methoxypropyl)-2-(3-nitro-1 H-1,2,4-triazol-1-yl)acetamide) and nimorazole (5-nitroimidazole derivative, 4-(2-(5-nitro-1H-1-imidazolyl)ethyl)morpholine) have been tested clinically as hypoxic cell radiosensitizers, mainly outside Japan. To determine if these sensitizers deserve clinical investigation in Japan, we reevaluated the radiosensitizing effects of these compounds in vitro and in vivo, in comparison with a fluorinated 2-nitroimidazole derivative KU-2285 (N1-(2-hydroxyethyl)-1,2-difluoro-3-(2-nitro-1 H-1-midazolyl)propanamide). KU-2285 is a known and established radiosensitizer, but is not suitable for clinical studies because of the high cost of synthesis. In vitro, the radiosensitizing effects of the three compounds on SCCVII (squamous cell carcinoma line in C3H mice) tumor cells were examined at 0.5 and 1 mM under aerobic or hypoxic conditions, using a colony assay. In vivo, SCCVII tumors grown subcutaneously in the hind legs of C3H/HeN mice were irradiated with or without prior intraperitoneal administration of 100, 200 or 400 mg/kg of the drugs. Thereafter, tumor growth delay was measured. In vitro, no sensitizing effect was observed under aerobic conditions at 1 mM. Under hypoxic conditions, the sensitizer enhancement ratio (SER) determined at 1% cell survival level for sanazole, nimorazole and KU-2285 was 1.55, 1.45 and 1.95, respectively, at 1 mM, and 1.40, 1.40 and 1.75, respectively, at 0.5 mM. In vivo, all three compounds had significant radiosensitizing effects; their effects appeared to decrease in the order of KU-2285, sanazole, and nimorazole. It was suggested that sanazole may be more suitable for clinical trials than nimorazole.     

Milk stimulates growth of prostate cancer cells in culture

Concern has been expressed about the fact that cows' milk contains estrogens and could stimulate the growth of hormone-sensitive tumors. In this study, organic cows' milk and two commercial substitutes were digested in vitro and tested for their effects on the growth of cultures of prostate and breast cancer cells. Cows' milk stimulated the growth of LNCaP prostate cancer cells in each of 14 separate experiments, producing an average increase in growth rate of over 30%. In contrast, almond milk suppressed the growth of these cells by over 30%. Neither cows' milk nor almond milk affected the growth of MCF-7 breast cancer cells or AsPC-1 pancreatic cancer cells significantly. Soy milk increased the growth rate of the breast cancer cells. These data indicate that prostate and breast cancer patients should be cautioned about the possible promotional effects of commercial dairy products and their substitutes.     

叶黄素对人前列腺癌PC-3M细胞增殖抑制作用

目的探讨叶黄素对体外培养人前列腺癌PC-3M细胞增殖的抑制作用及机制。方法选择人前列腺癌PC-3M细胞为研究对象,随机分为5组:0(对照组)、5、10、20、40μmol/L叶黄素组,采用噻唑蓝法检测PC-3M细胞增殖抑制率,采用流式细胞术检测PC-3M细胞周期,采用荧光显色法观察细胞形态,紫外分光光度法检测细胞上清液中caspase-3的含量。结果与对照组比较,各剂量叶黄素组PC-3M细胞增殖抑制率明显升高,具有剂量和时间依赖性(P <0.01);与对照组比较,叶黄素10、20、40μmol/L剂量组PC-3M细胞G1期细胞比例明显增加,G2M期细胞比例明显减少(P <0.01);与对照组比较,叶黄素各剂量组PC-3M细胞形状发生明显变化,细胞变圆,核碎裂且核染色质凝聚,凋亡小体形成,且随剂量增加凋亡数显著升高;与对照组比较,各剂量叶黄素组PC-3M细胞上清液中caspase-3含量明显升高,呈剂量效应关系(P <0.05)。结论叶黄素可抑制体外培养的人前列腺癌PC-3M细胞的增殖,诱导其凋亡,其机制可能与叶黄素增加PC-3M细胞caspase-3表达有关。     

雷公藤内酯醇诱导PC3细胞凋亡的机制研究

目的观察雷公藤内酯醇（triptolide,TPL）对去势后抵抗性前列腺癌（Castration—Resistant Prostate Cancer,CRPC）PC3细胞的生长增殖抑制作用,对其诱导凋亡相关基因进行分析。方法MTY比色法观察TPL对PC3细胞的生长增殖抑制作用;Annexin—V／PI标记分析细胞凋亡;RT-PCR方法检测0、6、12、24h BCL-2、BAX、PIG3、P21、FAS、CASPASE3等与凋亡相关基因表达变化。结果什L抑制PC3细胞生长呈时间和剂量依赖性,24h和48h半数抑制浓度分别为18．3ng／ml和13．5ng／ml,与24h组相比,48h组低浓度（5ng／ml,t=1．47,P〉0．05）和高浓度（160ng／ml,t=0．91,P〉0．05）差异无统计学意义。而10ng／ml（t=3．26,P〈0．05）、20ng／ml（t=4．21,P〈0．05）、40．g／ml（t=4．09,P〈0．05）、80ng／ml（t=2．91,P〈0．05）差异有统计学意义。Annexin-v／PI检测经18．3ng／ml,I’PL处理后的PC3细胞凋亡随着作用时间的延长而增多,相对于对照组,6、12、24h组Anexin—V阳性／PI阴性表达率分别为（5．42±2．21）％（t=3．52,P〈0．05）、（13．51±3．37）％（t=6．53,P〈0．01）、（29．3±4．53）％（t=8．74,P〈0．01）差异有统计学意义,并且各组间差异有统计学意义（P〈0．05）;RT—PCR显示经18．3ng／ml TPL处理后,BAX、HG3表达递增,P21、BCL-2表达递降,FAS、CASPASE3表达无明显改变。结论TPL可以抑制PC3生长增殖并诱导细胞凋亡,而在凋亡的过程中,BAX、BCL-2、PIG3、P21等基因的改变可能扮演着重要的角色。