姜黄素对肝癌细胞HepG2的抗癌作用及P21WAF1/CIP1表达的影响

目的：探讨姜黄素对肝癌HepG2细胞抗癌作用及相关周期蛋白依赖激酶抑制因子P21WAF1/CIP1表达的影响。方法：体外培养肝癌HepG2细胞,用MTT法检测姜黄素对HepG2细胞的抑制作用,以RT-PCR方法检测HepG2细胞中P21WAF1/CIP1mRNA的表达,用免疫细胞化学检测其P21WAF1/CIP1蛋白的表达。结果：姜黄素呈时间剂量性抑制HepG2细胞的生长,并显著上调HepG2细胞中P21WAF1/CIP1mRNA和蛋白的表达。结论：姜黄素能抑制HepG2细胞的生长,并上调其中P21WAF1/CIP1的表达。     

姜黄素对肝癌细胞HepG2 的抗癌作用及P21WAF1/CIP1 表达的影响

目的:探讨姜黄素对肝癌HepG2细胞抗癌作用及相关周期蛋白依赖激酶抑制因子P21WAF1/CIP1表达的影响.方法:体外培养肝癌HepG2细胞,用MTT法检测姜黄素对HepG2细胞的抑制作用,以RT-PCR方法检测HepG2细胞中P21WAF1/CIP1mRNA的表达,用免疫细胞化学检测其P21WAF1/CIP1蛋白的表达.结果:姜黄素呈时间剂量性抑制HepG2细胞的生长,并显著上调HepG2细胞中P21WAF1/CIP1mRNA和蛋白的表达.结论:姜黄素能抑制HepG2细胞的生长,并上调其中P21WAF1/CIP1的表达.     

姜黄素通过下调P-糖蛋白和热休克蛋白70逆转耐热肝癌细胞的阿霉素耐受性

探讨姜黄素对耐热肝癌细胞 (HepG2/TT) 阿霉素耐受性的逆转作用及其机制．用MTT检测细胞活力,PI染色流式细胞术检测细胞凋亡,高效液相色谱法检测细胞内阿霉素的积累,Western blot检测细胞P-糖蛋白 (P-glycoprotein,P-gp)、热休克蛋白70 (heat shock protein 70, Hsp70) 和caspase-3 的表达．耐热肝癌细胞HepG2/TT能耐受阿霉素引起的细胞毒性和 凋亡;姜黄素在5、10和20 μmol/L时,能浓度依赖性地降低阿霉素对HepG2/TT 细胞的IC50,增强阿霉素对HepG2/TT 细胞的凋亡诱导作用．耐热肝癌细胞HepG2/TT 与非耐热肝癌细胞HepG2比较,其P-gp和Hsp70 的表达水平明显增高; 10 μmol/L姜黄素处理24 h 后,HepG2/TT细胞P-gp和Hsp70的表达水平显著下降．HepG2/TT 细胞内阿霉素的积累低于HepG2细胞;10 μmol/L姜黄素处理 3 h后,HepG2/TT 细胞内阿霉素的积累明显增加．HepG2/TT细胞能抑制阿霉素激活 caspase-3;10 μmol/L姜黄素处理24 h后,阿霉素对 HepG2/TT细胞caspase-3的激活作用增强．上述结果表明,姜黄素能逆转耐热肝癌细胞HepG2/TT的阿霉素耐受性,其机制可能与其下调P-gp和Hsp70的表达,进而促进阿霉素激活caspase-3 有关．     

姜黄素类似物EF25-(GSH)_2抗肝癌作用的研究

在细胞及动物个体水平探讨姜黄素类似物EF25-(GSH)2的抗肝癌作用。用不同浓度的EF25-(GSH)2处理体外培养的肝癌细胞、正常肝细胞以及HepG2荷瘤裸鼠,MTT法检测细胞存活率,电镜及激光共聚焦显微镜观察细胞形态,蛋白质免疫印迹法(Western blot)检测AMPK/Akt/mTOR相关通路蛋白磷酸化水平的变化。结果显示,EF25-(GSH)2对HepG2作用48 h的IC50为7.2μmol/L,对其生长抑制作用明显优于姜黄素及顺铂,且对正常细胞的杀伤作用较小。形态学观察到细胞中有自噬现象的发生。免疫印迹法结果提示,EF25-(GSH)2可能通过AMPK/Akt/mTOR的相关通路抑制肿瘤细胞生长。对肝癌模型裸鼠的实验显示,给药后的肿瘤体积明显缩小。该实验结果证明,EF25-(GSH)2具有良好的作为肝癌治疗药物的开发前景。     

血管内皮生长因子受体信号转导通路与肿瘤血管生成

血管内皮生长因子是促进血管生成的重要调节因子.它能促进内皮细胞增殖、迁移,阻止内皮细胞凋亡、管腔网状结构退化,增加血管渗透性.所有这些作用都是通过血管内皮生长因子受体信号转导通路实现的.它们在肿瘤血管生成、肿瘤生长中起着重要的作用.以血管内皮生长因子受体信号转导通路为靶点是开发肿瘤血管生成抑制剂的理想策略.     

肿瘤组织缺氧诱导VEGF表达的分子机制

血管新生是恶性肿瘤生长、浸润和转移的前提条件,血管内皮生长因子是体内最重要的血管生长因子之一,而缺氧又是诱导肿瘤血管内皮生长因子表达的主要因素,本文对肿瘤组织缺氧诱导血管内皮生长因子表达的分子机制进行综述。     

肿瘤组织缺氧诱导VEGF表达的分子机制

血管新生是恶性肿瘤生长、浸润和转移的前提条件,血管内皮生长因子是体内最重要的血管生长因子之一,而缺氧又是诱导肿瘤血管内皮生长因子表达的主要因素,本文对肿瘤组织缺氧诱导血管内皮生长因子表达的分子机制进行综述。     

血管内皮生长因子的神经保护作用

血管内皮生长因子（vascular endothelial growth factor,VEGF）是内皮细胞特异性的生长因子,大多数关于VEGF的研究都是致力于其在血管生长方面的作用,而近年来有大量文献报道VEGF具有神经营养和促神经发生作用,它能够直接作用于神经元细胞和神经胶质细胞甚至是神经干细胞,促进其生长及存活。VEGF的多种功能使其和多种神经退行性疾病相关,如阿茨海默病,肌萎缩侧索硬化症,帕金森病等。导入VEGF基因能够改善肌萎缩侧索硬化症、帕金森病动物模型的病情。     

中药促缺血心肌血管新生机制研究进展

中药在缺血性心脏病的治疗方面具有重要作用,能够缓解症状并改善预后,促血管新生可能是其发挥作用的机制之一。已证实通过促进血管新生,可使心肌梗死面积缩小,减少细胞坏死和凋亡,改善心功能等,但其机制尚需进一步明确。目前中医药在促进梗死心肌血管新生研究方面已经取得了一定的成果,研究发现,大量中药单体、方剂及中成药具有促进血管新生作用。随着对中药促血管新生研究的不断深入,其机制的研究亦愈趋广泛,已深入到细胞及分子水平:涉及骨髓干细胞,促进血管生长的因子及其受体(血管内皮细胞生长因子、碱性成纤维细胞生长因子、胰岛素样生长因子等),抑制血管生长的因子(血管抑素、内皮抑素)及Akt/NOS/NO等通路。本文就相关研究回顾中药对缺血心肌血管新生影响并对机制进行探讨。     

乙肝病毒x基因致肝细胞癌机理探讨

构建HBx基因真核表达载体, 导入HepG2细胞中, 无血清培养同步化后, Western 杂交检测HepG2-X细胞IGF-ⅠR, PCNA和VEGF表达增强, 但无p21CIP1/WAF1表达. 流式细胞检测细胞周期, HepG-X0细胞70%进入G0～G1期, 而HepG2-X细胞仅56%进入G0～G1期, 与血清培养亲本HepG2细胞几乎相同. HBx基因增强IGF-ⅠR表达, 通过信号通路到细胞核, 同时使PCNA表达增强, p21CIP1/WAF1表达抑制, 推进细胞周期, 使G0～G1细胞明显减少. HBx基因增强VEGF表达, 通过旁分泌作用使血管内皮增生; 以上可能是HBx蛋白致HCC 的机理之一.     

Preventive Efficacy of Bulk and Nanocurcumin Against Lead-Induced Oxidative Stress in Mice

Chronic lead exposure is associated with several health disorders in humans and animals. Lead exposure leads to the generation of reactive oxygen species and depletes body antioxidant enzymes causing damage to various macromolecules and ultimately cell death. Curcumin has been widely recognized to protect against metal toxicity but has major limitations of reduced bioavailability. Nanoencapsulation of curcumin could be an effective strategy to combat lead induced toxic manifestations. The present study investigates the protective efficacy of bulk and nanocurcumin against lead-induced toxicity. Swiss albino mice were daily exposed to lead acetate (25 mg/kg, i.p.) alone and after 1 h treated either with curcumin (15 mg/kg, orally) or nanocurcumin (15 mg/kg, orally) for two consecutive weeks. The preventive efficacy of nanocurcumin was evaluated against various altered biochemical variables suggestive of oxidative stress and lead accumulation in blood and soft tissues. Coadministration of nanocurcumin with lead restored the altered δ-aminolevulinic acid dehydratase activity, glutathione (reduced and oxidized) levels, and also decreased reactive oxygen species, and thiobarbituric acid reactive substances levels. Nanocurcumin due to its possible chelating property and enhanced bioavailability efficiently removed lead from blood and soft tissues compared to bulk curcumin. Results demonstrate the enhanced preventive efficacy of nanocurcumin and suggest an interesting and novel approach for better treatment of lead toxicity.     

Efficacy of Biodegradable Curcumin Nanoparticles in Delaying Cataract in Diabetic Rat Model

Curcumin, the active principle present in the yellow spice turmeric, has been shown to exhibit various pharmacological actions such as antioxidant, anti-inflammatory, antimicrobial, and anti-carcinogenic activities. Previously we have reported that dietary curcumin delays diabetes-induced cataract in rats. However, low peroral bioavailability is a major limiting factor for the success of clinical utilization of curcumin. In this study, we have administered curcumin encapsulated nanoparticles in streptozotocin (STZ) induced diabetic cataract model. Oral administration of 2 mg/day nanocurcumin was significantly more effective than curcumin in delaying diabetic cataracts in rats. The significant delay in progression of diabetic cataract by nanocurcumin is attributed to its ability to intervene the biochemical pathways of disease progression such as protein insolubilization, polyol pathway, protein glycation, crystallin distribution and oxidative stress. The enhanced performance of nanocurcumin can be attributed probably to its improved oral bioavailability. Together, the results of the present study demonstrate the potential of nanocurcumin in managing diabetic cataract.     

Curcumin induces apoptosis through mitochondrial hyperpolarization and mtDNA damage in human hepatoma G2 cells

Curcumin, a major pigment of turmeric, is a natural antioxidant possessing a variety of pharmacological activities and therapeutic properties. But its mechanisms are unknown. In our previous study, we found that a 2-h exposure to curcumin induced DNA damage to both the mitochondrial DNA (mtDNA) and the nuclear DNA (nDNA) in HepG2 cells and that mtDNA damage was more extensive than nDNA damage. Therefore, experiments were initiated to evaluate the role of mtDNA damage in curcumin-induced apoptosis. The results demonstrated that HepG2 cells challenged with curcumin for 1 h showed a transient elevation of the mitochondrial membrane potential (DeltaPsim), followed by cytochrome c release into the cytosol and disruption of DeltaPsim after 6 h exposure to curcumin. Apoptosis was detected by Hoechst 33342 and annexin V/PI assay after 10 h treatment. Interestingly, the expression of Bcl-2 remained unchanged. A resistance to apoptosis for the corresponding rho0 counterparts confirmed a critical dependency for mitochondria during the induction of apoptosis in HepG2 cells mediated by curcumin. The effects of PEG-SOD in protecting against curcumin-induced cytotoxicity suggest that curcumin-induced cytotoxicity is directly dependent on superoxide anion O2- production. These data suggest that mitochondrial hyperpolarization is a prerequisite for curcumin-induced apoptosis and that mtDNA damage is the initial event triggering a chain of events leading to apoptosis in HepG2 cells.     

重组七鳃鳗富含半胱氨酸RGD蛋白rLj-RGD1对人肝癌HepG2细胞的抑制作用

rLj-RGD1为源于日本七鳃鳗口腔腺的基因重组蛋白,其富含半胱氨酸并具有一个RGD(Arg-Gly-Asp)模体.前期工作表明,rLj-RGD1具有抑制血小板聚集及血管新生的RGD毒素蛋白典型功能.为了研究rLj RGD1是否具有RGD毒素蛋白的另一典型抗肿瘤功能,以人肝癌HepG2细胞为模型,对rLj RGD1进行了活性研究.MTT法结果显示,rLj-RGD1呈剂量依赖方式抑制HepG2细胞的增殖,其半抑制浓度(IC50)为36 μmol/L;细胞迁移与浸润实验结果显示,rLj-RGD1能以剂量依赖方式抑制HepG2细胞碱性成纤维生长因子（bFGF）诱导的迁移与浸润.Hoechst染色和DNA Ladder等细胞凋亡实验表明,rLj-RGD1能够以剂量依赖方式诱导HepG2细胞发生凋亡.细胞黏附实验表明,rLj-RGD1以剂量依赖方式抑制HepG2细胞与玻连蛋白(vitronectin, VN)的黏附.上述结果表明,rLj-RGD1具有抑制人肝癌HepG2细胞增殖、迁移和浸润的功能,并可诱导其发生失巢凋亡. 本研究结果提示,rLj-RGD1具有典型的RGD毒素蛋白抗肿瘤功能,其未来具有成为抗肿瘤药物的潜力.     

四逆散对人肝癌HepG2细胞增殖、凋亡的影响及其机制*

目的: 探讨含四逆散药液血清对人肝癌HepG2细胞增殖、凋亡的影响及机制。方法: 将人肝癌HepG2细胞分为5组,每组3个复孔。实验组细胞用五氟尿嘧啶(5-FU)或不同浓度的含四逆散药液血清处理48 h后,用倒置显微镜观察含四逆散药液血清处理后人肝癌HepG2细胞形态的变化;MTT法检测含四逆散药液血清对HepG2细胞生长的抑制作用;荧光染色和流式细胞术分别分析含四逆散药液血清对HepG2细胞凋亡的影响。Rho123染色法检测线粒体膜电位变化,Western blot检测细胞凋亡相关蛋白的表达。结果: 与对照组比较,含四逆散药液血清处理人肝癌HepG2细胞后,细胞数量显著减少(P＜0.01),形态发生改变,呈现典型的凋亡细胞形态;G1期细胞数明显增加,而G2 期细胞数量显著减少(P＜0.05);Bax、Caspase-3、-9和Cyt-c的表达显著升高,而Bcl-2的表达显著降低(P＜0.05);随着含四逆散药液血清浓度增大,HepG2细胞线粒体膜电位显著下降(P＜0.05)。结论: 四逆散可以抑制HepG2细胞增殖,并通过线粒体途径诱导细胞凋亡。     

Nanocurcumin Prevents Hypoxia Induced Stress in Primary Human Ventricular Cardiomyocytes by Maintaining Mitochondrial Homeostasis

Hypoxia induced oxidative stress incurs pathophysiological changes in hypertrophied cardiomyocytes by promoting translocation of p53 to mitochondria. Here, we investigate the cardio-protective efficacy of nanocurcumin in protecting primary human ventricular cardiomyocytes (HVCM) from hypoxia induced damages. Hypoxia induced hypertrophy was confirmed by FITC-phenylalanine uptake assay, atrial natriuretic factor (ANF) levels and cell size measurements. Hypoxia induced translocation of p53 was investigated by using mitochondrial membrane permeability transition pore blocker cyclosporin A (blocks entry of p53 to mitochondria) and confirmed by western blot and immunofluorescence. Mitochondrial damage in hypertrophied HVCM cells was evaluated by analysing bio-energetic, anti-oxidant and metabolic function and substrate switching form lipids to glucose. Nanocurcumin prevented translocation of p53 to mitochondria by stabilizing mitochondrial membrane potential and de-stressed hypertrophied HVCM cells by significant restoration in lactate, acetyl-coenzyme A, pyruvate and glucose content along with lactate dehydrogenase (LDH) and 5'' adenosine monophosphate-activated protein kinase (AMPKα) activity. Significant restoration in glucose and modulation of GLUT-1 and GLUT-4 levels confirmed that nanocurcumin mediated prevention of substrate switching. Nanocurcumin prevented of mitochondrial stress as confirmed by c-fos/c-jun/p53 signalling. The data indicates decrease in p-300 histone acetyl transferase (HAT) mediated histone acetylation and GATA-4 activation as pharmacological targets of nanocurcumin in preventing hypoxia induced hypertrophy. The study provides an insight into propitious therapeutic effects of nanocurcumin in cardio-protection and usability in clinical applications.     

紫花牡荆素诱导人肝癌HepG2细胞凋亡的研究

目的：研究紫花牡荆素（Casticin）对肝癌HepG2细胞增殖抑制和凋亡诱导的作用,并探讨其作用机制。方法：用终浓度为0、0.5、1.0、2.0umol/L的Casticin作用于HepG2细胞,于12、24、48h后采用MTT法检测细胞增殖抑制率;Hoechst33342核染色,观察细胞形态学变化;24h后收集各组肝癌HepG2细胞,流式细胞术检测细胞周期及凋亡率;RT-PCR检测survivin mRNA表达。结果：MTT法检测显示,Casticin对肝癌HepG2细胞有增殖抑制作用,并存在浓度和时间依赖关系;Hoechst33342染色后,可见核染色质凝集,凋亡细胞呈致密浓染,与对照组相比,Casticin处理后凋亡细胞比例增加;Casticin作用24h后,细胞被阻滞于G2/M期,随药物质量浓度的增加,细胞凋亡率逐渐增加;RT-PCR结果显示,Casticin下调肝癌HepG2细胞survivin mRNA表达。结论：Casticin在体外对肝癌HepG2细胞有明显的增殖抑制和凋亡诱导作用,初步推断Casticin诱发肝癌细胞凋亡与其对survivin基因表达的抑制有关。     

维甲酸对肝癌细胞HepG2糖基化磷脂酰肌醇特异性磷酯酶D表达及细胞生物学特性的影响

应用MTT、流式细胞仪、免疫印迹法检测全反式维甲酸(ATRA)单独或联合糖基化磷脂酰肌醇特异性磷酯酶D(GPI-PLD)特异性抑制剂1,10-二氮杂菲对肝癌细胞HepG2生物学特性的改变.ATRA使肝癌细胞HepG2 GPI-PLD基因表达及酶活性上调,并呈现剂量和时间依赖性.ATRA可抑制肝癌细胞HepG2增殖,使肝癌细胞Caspase-3表达水平显著增加,Bcl-2表达水平下调,促进肝癌细胞凋亡(P<0.05).ATRA联合1,10-二氮杂菲诱导组细胞,Bcl-2、细胞增殖活性较ATRA单独诱导组显著增强,Caspase-3、凋亡率显著下降.维甲酸可促进肝癌细胞HepG2 GPI-PLD基因表达上调,高活性的GPI-PLD有助于维甲酸抑制肝癌细胞增殖,促进肝癌细胞凋亡.     

肝细胞癌组织中OATP1B3的表达及其作用的初步研究

摘要 目的：探讨有机阴离子转运多肽1B3（OATP1B3）在肝细胞癌（肝癌）组织中的表达及作用。方法：通过免疫组化实验和免疫印迹试验（Western blot）检测肝癌组织及其癌旁组织中OATP1B3情况,并分析其与患者临床病理特征的相关性。采用实时荧光定量聚合酶链反应（qRT-PCR）检测OATP1B3在多株肝癌细胞中的表达情况,选择表达量相对较低的人肝癌细胞（HepG2和Huh7）细胞进行过表达实验,细胞毒实验（MTT）和流式细胞术分别检测OATP1B3对细胞增殖及凋亡的影响。结果：肝癌组织中OATP1B3表达水平明显低于癌旁组织（P<0.05）,且与患者恶性肿瘤国际临床病期分类（TNM分期）、肿瘤分化程度、有无肿瘤复发显著相关（P<0.05）。过表达OATP1B3可抑制HepG2和Huh7细胞增殖,促进细胞凋亡。结论：OATP1B3在肝癌组织中低表达,上调其表达可抑制肝癌细胞增殖和促进细胞凋亡。OATP1B3可能是肝癌的抑癌基因,对肝癌的发生、发展具有重要作用。