大蒜素抑制卵巢癌SKOV-3／DDP细胞生长作用浓度的实验研究

目的:筛选大蒜素有效作用于人上皮性卵巢癌耐顺铂细胞株SKOV-3/DDP的合适的浓度范围,为进一步探讨大蒜素抑制卵巢癌SKOV-3/DDP细胞生长的作用机制奠定基础.方法:首先培养人上皮性卵巢癌耐顺铂细胞株SKOV-3/DDP,细胞生长至时数生长期后选取0.625～200μg/ml共9个浓度的大蒜素分别作用SKOV-3/DDP细胞株24、48小时后,用MTT法测定细胞活力.结果:0.625～200μg/ml大蒜素均能抑制SKOV-3/DDP细胞的生长,且呈时间--剂量依赖性,但高浓度(100～200μg/ml)时细胞毒性作用明显,作用24小时细胞抑制率分别为75.12%、82.79%,作用48小时细胞几乎全部死亡.结论:以上MTT结果可作为后期药物实验的大蒜素浓度及观察时间筛选的实验依据,可选择中低浓度(0.625～50μg/ml)作为进一步探讨大蒜素抑制卵巢癌SKOV-3/DDP细胞生长的作用机制(如观察大蒜素对SKOV-3/DDP细胞周期的影响等)的浓度范围.     

黄芪成分F_3新制剂对胃癌细胞株的抑制作用

[目的]观察黄芪成分F3新制剂对胃癌(BGC-823)细胞株体外细胞生长、抑制作用.[方法]利用基因组DNA电泳,流式细胞仪检测细胞凋亡.[结果]基因组DNA电泳出现“梯形”条带;流式细胞仪分析出现低于G1期DNA含量的亚二倍体凋亡率.[结论]黄芪成分F3新制剂对胃癌细胞有明显的抑制作用.     

RP-HPLC法同时检测石榴皮3种多酚类提取物含量研究

采用RP-HPLC法,Symmetry C18(250 mm×4.6 mm, 5μm)色谱柱,以甲醇(A)-0.2%磷酸溶液(B)为流动相梯度洗脱,流速1.0 mL/min,柱温30℃,检测波长272 nm同时检测安石榴苷、鞣花酸和没食子酸含量。采用SPSS 17.0对测定结果进行分析,比较不同品种石榴皮中各成分的含量差异。结果表明,安石榴苷、鞣花酸、没食子酸分别在13.120～209.92μg/mL,13.325～213.20μg/mL和13.275～212.40μg/mL与峰面积呈线性关系,平均加样回收率为97.58%(RSD:1.02%,n=6),99.31%(RSD:0.94%,n=6)和98.63%(RSD:0.70%,n=6)。白玉石榴皮中安石榴苷、鞣花酸的含量分别为7.13%和1.00%,比红酸石榴皮中的含量高。安徽地区石榴皮中安石榴苷、鞣花酸、没食子酸3种成分含量同时检测方法的灵敏度高、准确度好、重现性一致,可用于安徽本地石榴皮部位中安石榴苷、鞣花酸、没食子酸成分含量同时检测。     

大蒜素对人卵巢癌耐顺铂细胞株SKOV-3/DDP超微结构的影响

目的:通过观察大蒜素对体外培养的人上皮性卵巢癌耐顺铂细胞株(SKOV-3/DDP)的超微结构的影响,以期寻找对耐顺铂治疗的上皮性卵巢癌患者的有效治疗药物。方法:细胞培养:体外培养人上皮性卵巢癌耐顺铂细胞株SKOV-3/DDP,细胞生长至对数生长期后,调整细胞密度为5×l04/ml置于200ml的培养瓶中培养。用透射电子显微镜观察大蒜素作用后SKOV-3/DDP细胞的超微结构:大蒜素组用含25μg/ml大蒜素的培养液培养SKOV-3/DDP细胞,空白对照组用不含药的培养液培养SKOV-3/DDP细胞,两组作用24小时后收集细胞,通过透射电子显微镜观察SKOV-3/DDP的超微结构。结果:大蒜素对SKOV-3/DDP细胞超微结构的影响:透射电子显微镜观察未加药物培养的SKOV-3/DDP细胞呈椭圆形,细胞核为卵圆形,未出现凋亡细胞,经过大蒜素作用后的SKOV-3/DDP细胞则出现明显的超微结构改变并具有典型凋亡细胞特征:细胞核固缩,核膜皱缩,胞浆浓缩并出现大小不等的空泡,染色质浓聚,形成凋亡小体。结论:大蒜素可诱导SKOV-3/DDP细胞呈凋亡改变。大蒜素可诱导SKOV-3/DDP细胞凋亡可能是其对人耐顺铂上皮性卵巢癌的抗癌机制之一,为使大蒜素成为治疗耐顺铂上皮性卵巢癌患者的有效药物提供了可靠的体外实验依据。     

鞣花酸对肿瘤细胞增殖抑制和诱导凋亡作用的初步研究

目的：探讨鞣花酸（ellagic acid）对肿瘤细胞的生长抑制作用及其机制.方法：用鞣花酸处理体外培养的肿瘤细胞,用细胞增殖试验（MTT法）、克隆形成试验研究ELA对肿瘤细胞的增殖抑制作用,Brdu掺入试验、流式细胞术检测ELA对肿瘤细胞DNA合成、凋亡和细胞周期的影响.结果：用ELA处理后,肿瘤细胞增殖活性降低（p〈0.05）,克隆形成下降（p〈0.005）,Brdu标记指数降低（p〈0.05）,细胞周期分布改变,凋亡指数升高（p〈0.01）,G0/G1期细胞数增加（p〈0.01）,S期细胞数减少（p〈0.01）,在一定剂量内对正常细胞无明显影响.结论：ELA对所试肿瘤细胞的增殖有显著地抑制作用,其作用机理可能与抑制肿瘤细胞DNA合成,诱导肿瘤细胞凋亡和细胞周期阻滞有关.     

虾青素对铜离子诱导的前列腺细胞氧化损伤的影响（英文）

目的:研究虾青素对铜离子诱导的前列腺细胞氧化损伤的影响,并探索其作用机制。创新点:首次研究虾青素对铜离子诱导的前列腺细胞及前列腺癌细胞氧化损伤的影响,并比较其对两种细胞作用的差异。方法:MTT法测定铜离子与虾青素对前列腺细胞(RWPE-1)和前列腺癌细胞(PC-3)生长的影响;采用细胞流式仪测定虾青素对铜离子诱导的RWPE-1和PC-3细胞凋亡的影响;荧光分光光度法测定了虾青素对铜离子诱导的活性氧自由基(ROS)产生的影响;采用罗丹明123(Rh123)染色检测虾青素对铜离子诱导的细胞线粒体膜电位(MMP)变化的影响;采用试剂盒测定了虾青素对铜离子存在下丙二醛(MDA)含量、超氧化物歧化酶(SOD)、过氧化氢酶(CAT)及谷胱甘肽过氧化物酶(GSH-Px)活性变化的影响。结论:结果表明,铜离子能诱导RWPE-1和PC-3细胞凋亡,并伴随细胞内ROS和MDA含量升高;虾青素处理可显著降低RWPE-1细胞中MDA含量,升高线粒体膜电位,并保持ROS含量稳定;虾青素处理可降低PC-3细胞中SOD、GSH-Px和CAT的活性,而对RWPE-1细胞则作用相反。因此,虾青素处理能有效降低铜离子对RWPE-1细胞引起的损伤,而通过降低抗氧化酶活性加剧铜离子对PC-3细胞的损伤。     

RP-HPLC法同时测定石榴皮和石榴汁中鞣花酸的含量

目的:建立同时检测石榴皮、汁中鞣花酸的含量测定方法.方法:采用反相高效液相色谱法,Hyperil C18(250 mm ×4.6 mm,5μm)色谱柱,以乙腈-0.3％ TFA(20∶80)为流动相,梯度洗脱,流速1.0mL/min,柱温30℃,检测波长254nm.结果:鞣花酸0.1048 ～ 20.96 μg/mL之间线性关系良好(r=0.9999),平均加样回收率为99.55％(RSD=1.44％,n=9).结论:此方法灵敏快速、准确易行,可用于石榴皮、汁中鞣花酸含量的同时检测.     

As2O3对人体外宫颈癌细胞的影响

采用光镜及荧光显微镜观察、溴化二甲噻唑二苯四氮唑(MTT)还原法、流式细胞仪检测法检测As2O3对人HeLa细胞的影响.HeLa细胞经As2O3作用后,出现凋亡的形态学改变,细胞增殖受到浓度依赖性抑制,流式细胞仪检测结果显示细胞凋亡率呈浓度依赖性增高.As2O3具有诱导HeLa细胞发生凋亡的生物活性.     

异乌药内酯通过ROS介导的线粒体凋亡途径诱导A549细胞凋亡的研究

为探究异乌药内酯诱导细胞内活性氧的产生在介导线粒体凋亡途径中的作用及对肺癌A549细胞凋亡的影响,将A549细胞分为对照组（DMSO）、异乌药内酯处理组（15μmol/L）、抗氧化剂NAC组（3 mmol/L）和异乌药内酯联合NAC处理组（Isolinderalactone 15μmol/L+NAC 3 mmol/L）进行实验。采用CCK-8方法检测细胞活性,通过流式细胞术检测细胞凋亡情况,DCFH-DA法检测A549细胞内活性氧水平变化,JC-1染色后流式细胞术检测细胞内线粒体膜电位,Western blot方法检测细胞内线粒体凋亡通路相关关键蛋白表达的变化。结果表明：异乌药内酯能抑制肺癌A549细胞生长并诱导细胞凋亡,促进ROS积累,线粒体膜显著去极化,上调促凋亡蛋白Bax表达水平而下调抗凋亡蛋白Bcl-2表达水平。抗氧化剂NAC与异乌药内酯联合作用后,与异乌药内酯单独处理组相比,以上各方面的变化水平都呈现出不同程度的下降。异乌药内酯能通过诱导细胞内活性氧积累,激活线粒体凋亡途径从而促进A549细胞发生凋亡。     

瓜子金皂苷己对鱼藤酮损伤PC-12细胞的保护作用及其对CREB的影响

目的：观察瓜子金皂苷己（polygalasaponin F,PS-F）对鱼藤酮损伤的PC-12细胞的保护作用及其对cAMP反应元件结合蛋白（cAMP response element binding protein,CREB）表达的影响。方法：以鱼藤酮损伤的PC-12细胞为模型,通过倒置显微镜观察细胞形态,caspase 3活性检测试剂盒测定caspase 3活性,荧光素酶报告基因技术检测CREB表达的变化。结果：PC-12细胞在4μmol·L-1鱼藤酮损伤后,细胞出现萎缩变圆,折光率降低,caspase 3活性显著升高,呈现凋亡的现象。不同浓度（0.1,1.0,10.0μmol·L-1）瓜子金皂苷己能够剂量依赖性的减轻对细胞形态的影响,降低caspase 3活性。荧光素酶报告基因标记显示,瓜子金皂苷己能够增加报告基因CREB的表达。结论：瓜子金皂苷己能够抑制鱼藤酮诱导的PC-12细胞凋亡,其机制可能与增加CREB的表达有关。     

球毛壳菌素F对Poly（I：C）诱导的RAW264．7细胞TNF-α水平的影响

该实验主要研究球毛壳菌素F对聚肌苷酸胞苷酸Poly（I：C）诱导的小鼠巨噬细胞系RAW264．7 TNF-α表达水平的影响．采用不同浓度的Poly（I：C）（50μg·mL-1、100μg·mL-1和200μg·mL-1）和球毛壳菌素F（2μg·mL-1）对RAW264．7进行处理,运用流式细胞术检测细胞内TNF-α的表达水平．结果表明,各浓度的Poly（I：C）与对照组相比均能极显著提高RAW264．7的TNF-α水平（P〈0．001）,并且具有浓度依赖性;球毛壳菌素F能够极显著地降低Poly（I：C）诱导的细胞TNF．O／水平（P〈0．01）．结论,球毛壳菌素F可能是通过降低炎症因子TNF-α的分泌而达到抗炎作用的．     

球毛壳菌素F对Poly（I：C）诱导的RAW264．7细胞TNF-α水平的影响

该实验主要研究球毛壳菌素F对聚肌苷酸胞苷酸Poly（I：C）诱导的小鼠巨噬细胞系RAW264．7 TNF-α表达水平的影响．采用不同浓度的Poly（I：C）（50μg·mL-1、100μg·mL-1和200μg·mL-1）和球毛壳菌素F（2μg·mL-1）对RAW264．7进行处理,运用流式细胞术检测细胞内TNF-α的表达水平．结果表明,各浓度的Poly（I：C）与对照组相比均能极显著提高RAW264．7的TNF-α水平（P〈0．001）,并且具有浓度依赖性;球毛壳菌素F能够极显著地降低Poly（I：C）诱导的细胞TNF．O／水平（P〈0．01）．结论,球毛壳菌素F可能是通过降低炎症因子TNF-α的分泌而达到抗炎作用的．     

Effects of astaxanthin on oxidative stress induced by Cu<Superscript>2+</Superscript> in prostate cells

Astaxanthin (AST), a carotenoid molecule extensively found in marine organisms and increasingly used as a dietary supplement, has been reported to have beneficial effects against oxidative stress. In the current paper, the effects of AST on viability of prostate cells were investigated by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay; cell apoptosis and intracellular reactive oxygen species (ROS) levels were determined by flow cytometry; the mitochondrial membrane potential (MMP) was measured by fluorospectrophotometer; and activities of malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) were evaluated by a detection kit. The results show that copper ion (Cu²⁺) induced apoptosis, along with the accumulation of intracellular ROS and MDA, in both prostate cell lines (RWPE-1 and PC-3). AST treatments could decrease the MDA levels, increase MMP, and keep ROS stable in RWPE-1 cell line. An addition of AST decreased the SOD, GSH-Px, and CAT activities in PC-3 cell line treated with Cu²⁺, but had a contrary reaction in RWPE-1 cell lines. In conclusion, AST could contribute to protecting RWPE-1 cells against Cu²⁺-induced injuries but could cause damage to the antioxidant enzyme system in PC-3 cells.     

Investigation on apoptosis of neuronal cells induced by Amyloid beta-Protein

Objective: To construct a PC12 cell strain with neuronal differentiation, and observe the apoptosis and pro-liferation activity effects induced these cells by Amyloid beta-Protein (Aβ-43). Methods: 1) PC12 cells in logarithmicgrowth phase were subcultured for 24 h. After the culture fluid was changed, the cells were treated with Rat-β-NGF andcultured for 9 days. 2) Neuronal differentiation of PC12 cells in logarithmic growth phase were divided into four groups:control group (0), experimental group (1), experimental group (2) and experimental group (3). The concentrations of Aβ inthe four groups were 0 μmol/L, 1.25 μ mol/L, 2.5 μ mol/L and 5 μmol/L, respectively. The cells were harvested at 24, 48 and72 h later and stained with AnnexinV-FITC/PI after centrifugation and washing. Then flow cytometry was conducted toexamine the apoptosis percentage. 3) NGF-induced PC12 cells were selected and Aβ with different concentrations wasadded. The final concentrations of Aβ were 0 μmol/L, 1.25 μmol/L, 2.5 μmol/L and 5 μ mol/L, respectively. After the cellswere incubated in an atmosphere of 5% CO2 at 37 ℃ in an incubator for 72 h, the OD values were examined. Results: 1)Neuronal differentiated PC 12 cell lines were successfully established. 2) Flow cytometric examination indicated that Aβ(1.25, 2.5, and 5.0 μmol/L) could effectively induce apoptosis of neuronal-differented cells at the 24 h, 48 h and 72 h timepoints. 3) Aβ (0-5.00 μ mol/L) had no obvious effect on proliferation or restraining of the neuronal differentiation of thePC 12 cells after a 72 h interacting process. Conclusion: This investigation revealed successful neuronal differentiation of thePC12 cell strain. The induction of apoptosis of the neurocytes by various concentrations of Aβ was observed and the in-fluence of Aβ on induced proliferation of PC 12 cells by Rat-β-NGF was revealed. This study -05 provide basis for futureresearch on the molecular cure of AD and interdiction of AD evolution.     

Green tea polyphenols induce cell death in breast cancer MCF-7 cells through induction of cell cycle arrest and mitochondrial-mediated apoptosis

In order to study the molecular mechanisms of green tea polyphenols (GTPs) in treatment or prevention of breast cancer, the cytotoxic effects of GTPs on five human cell lines (MCF-7, A549, Hela, PC3, and HepG2 cells) were determined and the antitumor mechanisms of GTPs in MCF-7 cells were analyzed. The results showed that GTPs exhibited a broad spectrum of inhibition against the detected cancer cell lines, particularly the MCF-7 cells. Studies on the mechanisms revealed that the main modes of cell death induced by GTPs were cell cycle arrest and mitochondrialmediated apoptosis. Flow cytometric analysis showed that GTPs mediated cell cycle arrest at both G1/M and G2/M transitions. GTP dose dependently led to apoptosis of MCF-7 cells via the mitochondrial pathways, as evidenced by induction of chromatin condensation, reduction of mitochondrial membrane potential (ΔΨm), improvement in the generation of reactive oxygen species (ROS), induction of DNA fragmentation, and activations of caspase-3 and caspase-9 in the present paper.     

Edaravone protects PC12 cells from ischemic-like injury via attenuating the damage to mitochondria

Background： Edaravone had been validated to effectively protect against ischemic injuries. In this study, we investigated the protective effect of edaravone by observing the effects on anti-apoptosis, regulation of Bcl-2/Bax protein expression and recovering from damage to mitochondria after OGD （oxygen-glucose deprivation）-reperfusion. Methods： Viability of PC 12 cells which were injured at different time of OGD injury, was quantified by measuring MTT （2-（4,5-dimethylthia-zol-2-yl）-2,5-diphenyltetrazolium bromide） staining. In addition, PC 12 cells＇ viability was also quantified after their preincubation in different concentration of edaravone for 30 min followed by （OGD）. Furthermore, apoptotic population of PC 12 cells that reinsulted from OGD-reperfusion with or without preincubation with edaravone was determined by flow cytometer analysis, electron microscope and Hoechst/Pl staining. Finally, change of Bcl-2/Bax protein expression was detected by Western blot. Results： （1） The viability of PC12 cells decreased with time （1-12 h） after OGD. We regarded the model of OGD 2 h, then replacing DMEM （Dulbecco＇s Modified Eagle＇s Medium） for another 24 h as an OGD-reperfusion in this research. Furthermore, most PC 12 cells were in the state of apoptosis after OGD-reperfusion. （2） The viability of PC 12 cells preincubated with edaravone at high concentrations （1, 0.1, 0.01 μmol/L） increased significantly with edaravone protecting PC 12 cells from apoptosis after OGD-reperfusion injury. （3） Furthermore, edaravone attenuates the damage of OGD-reperfusion on mitochondria and regulated Bcl-2/Bax protein imbalance expression after OGD-reperfusion. Conclusion： Neuroprotective effects of edaravone on ischemic or other brain injuries may be partly mediated through inhibition of Bcl-2/Bax apoptotic pathways by recovering from the damage of mitochondria.