维生素E同系化合物诱导人肝癌细胞凋亡的作用

为探讨α 生育酚 (α T)、γ 生育酚 (γ T)、δ 生育酚 (δ T)及维生素E琥珀酸酯 (VES) 4种维生素E同系化合物对肝癌细胞凋亡影响 ,在体外培养的人肝癌细胞 (HepG2 )培养液中分别加入 1 2 5、2 5 0、50 0、1 0 0 0及 2 0 0 0mg L的 4种维生素E同系化合物培养 48h后 ,采用流式细胞仪技术 (FCM)和DNA梯度电泳 (DNALadder)定量分析HepG2细胞凋亡及DNA降解片段情况。结果发现 ,FCM凋亡检测发现δ T和VES培养的HepG2细胞凋亡率增加 ,并呈现剂量依赖关系 ,即随着δ T、VES剂量增加HepG2细胞凋亡亦增加 ,其中δ T诱导HepG2细胞凋亡作用强于VES ;γ T处理的HepG2细胞只在 2 0 0mg L剂量时出现轻度的凋亡增加 ;未见α T有诱导HepG2细胞凋亡的作用。DNA梯度电泳检测发现δ T和VES处理HepG2细胞的DNA出现明显的排列成梯状的分子条带 ,α T、γ T处理组则未见明显的条带。维生素E同系化合物诱导肝癌细胞凋亡作用不同 ,效应排序为δ T >VES >γ T >α T ,δ T和VES具有潜在抗肝癌临床应用前景     

维生素E同系化合物诱导人肝癌细胞凋亡的作用

为探讨α 生育酚 (α T)、γ 生育酚 (γ T)、δ 生育酚 (δ T)及维生素E琥珀酸酯 (VES) 4种维生素E同系化合物对肝癌细胞凋亡影响 ,在体外培养的人肝癌细胞 (HepG2 )培养液中分别加入 12 5、2 5 0、5 0 0、10 0 0及 2 0 0mg L的 4种维生素E同系化合物培养 48h后 ,采用流式细胞仪技术 (FCM)和DNA梯度电泳 (DNALad der)定量分析HepG2细胞凋亡及DNA降解片段情况。结果显示 ,FCM凋亡检测发现δ T和VES培养的HepG2细胞凋亡率增加 ,并呈现剂量依赖关系 ,即随着δ T、VES剂量增加HepG2细胞凋亡亦增加 ,其中δ T诱导HepG2细胞凋亡作用强于VES ;δ T处理的HepG2细胞只在 2 0 0mg L剂量时出现轻度的凋亡增加 ;未见α T有诱导HepG2细胞凋亡的作用。DNA梯度电泳检测发现δ T和VES处理HepG2细胞的DNA出现明显的排列成梯状的分子条带 ,α T、γ T处理组则未见明显的条带。结果提示 ,维生素E同系化合物诱导肝癌细胞凋亡作用不同 ,效应排序为δ T >VES >γ T >α T ,δ T和VES具有潜在抗肝癌临床应用前景     

VES抑制Raji细胞增殖中对CDK2和Bcl-2/Bax蛋白表达的影响

目的比较观察维生素E琥珀酸酯(VES)和维生素E(VE)对人单核细胞白血病Raji细胞的抗增殖作用及对细胞周期素依赖性激酶2(CDK2),细胞凋亡相关蛋白Bcl-2、Bax的影响.方法应用荧光显微镜、流式细胞仪、免疫细胞化学染色等技术方法,体外观察VES和VE对Raji细胞的作用.结果 VES和VE对Raji细胞均有程度不一的生长抑制作用,20μg/mlVES对Raji细胞的生长抑制在72h即达100%,而20μg/mlVE的抑制率仅与5μg/mlVES的接近.VES较强的生长抑制作用与诱发细胞凋亡的明显增加同步发生,具有剂量效应和时间效应关系,同时VES作用后,细胞内CDK2蛋白,Bcl-2蛋白表达下降,Bax蛋白表达增加.结论 VES通过影响细胞周期调控因子CDK2,细胞凋亡相关蛋白Bcl-2、Bax的表达可能是其抗肿瘤的途径之一.     

吲哚胺2,3双加氧酶对肝癌细胞的作用

目的:用IDO以及色氨酸经IDO催化后分解生成的代谢产物(L-犬尿素、邻氨基苯甲酸、喹啉酸)作用于肝癌细胞,观察肝癌细胞的生长及凋亡的情况.方法:将IDO作用于肝癌细胞.用MTT法测定细胞的活力,并用IDO的抑制剂1-MT进行干预.观察各个代谢产物对肝癌细胞生长的影响,用流式细胞仪检测细胞周期、细胞凋亡率以及细胞坏死率的变化.结果:不同浓度的IDO作用于HepG2细胞具有明显的时间和剂量依赖性,IDO浓度在0.5 mmol/L和5 mmol/L时可以促进HepG2细胞的生长(P<0.05),加入1～MT后IDO的这一作用受到抑制.只有喹啉酸能发挥提高细胞坏死率和凋亡率,抑制肝癌细胞的作用.结论:IDO表现为促进肝癌细胞生长的作用,1-MT可以抑制这一作用.L-犬尿素、邻氨基苯甲酸能够促进HepG2细胞的生长.喹啉酸对HepG2细胞有抑制生长作用.     

测定食品中维生素E荧光分光光度法的探讨

192 2年加利福尼亚大学的EVANS和BISHOP从莴苣和胚胎脂溶性膳食因子首先发现维生素E。 192 4年阿肯色大学的休尔 (SURE)命名该因子为维生素E ;1936年EVANGS极其同事麦胚油中分离出结晶维生素E ,命名为生育酚 (tocopherol) ;1938瑞士化学家KARRER首次人工合成这种维生素。1　定义及功能维生素E是指含苯并二氢吡喃结构 ,具有α -生育酚生物活性的一类物质的总称 ,目前已知有四种生育酚 ,即α -T ,β -T ,γ -T ,δ -T ,和四种生育三烯酚 (TOCOTRienols,即α -TT ,β -TTγ -TTδ-TT) ,其中α -生育酚的活性最高 ,故通常以…     

脉冲磁场对肝癌细胞株HepG2细胞增殖和分化的影响

目的　研究低频脉冲磁场对肝癌细胞株HepG2细胞增殖和分化的影响。方法　 2种频率不同场强相同的低频脉冲磁场处理HepG2细胞 ,以 2 对磺酸基苯基 3 ( 4 ,5 二甲基噻唑 ) 5 ( 3 羧甲氧基苯基 ) 二氢四唑嗡盐 (MTS)方法检测细胞的增殖情况 ,酶联免疫法测定HepG2细胞甲胎蛋白的分泌量。结果　 80Hz、1.5 5mT磁场分别处理HepG2细胞 1、4、8、12和 2 4h ,对HepG2细胞的增殖均无明显影响 ,差异无显著性 (P >0 .0 5 )。 16Hz脉冲磁场分别处理HepG2细胞 1、4、8和 2 4h ,对HepG2细胞的增殖和甲胎蛋白 (AFP)的分泌量均无明显影响 ,差异无显著性 (P >0 .0 5 )。结论　本研究没有观察到 16、80Hz脉冲磁场对肝癌细胞株HepG2的增殖和特征蛋白 (AFP)分泌存在“窗口”效应     

吡柔比星联合二烯丙三硫抑制膀胱癌细胞生长的协同作用研究

目的探讨吡柔比星(THP)联合二烯丙三硫(DATS)对膀胱癌细胞T24的协同杀伤效应及其机制。方法 MTT法、TUNEL法、流式细胞仪分别检测THP、DATS及二者联合对T24细胞生长、细胞周期及凋亡的影响;western-blot测定Bcl-2、Bax的表达变化;比色法测定caspase-3活性。结果 THP、DATS均能抑制T24细胞生长,DATS可明显提高THP的杀伤效应;THP(0.25mg/L)联合DATS(3mg/L)作用48h的细胞生存率为(32.3±2.6)%,低于单独应用THP(63.2±1.4)%和DATS(61.9±4.2)%,二者具有协同抑瘤作用;联合药物组可明显提高细胞凋亡率及G2/M期细胞比率,并更能显著抑制Bcl-2的表达,促进caspase-3的活性增加。结论 THP联合DATS可通过共同诱导肿瘤细胞凋亡发挥协同作用,抑制膀胱癌细胞生长,该机制可能与细胞周期阻滞、Bcl-2的表达及caspase-3的活性变化有关。     

维生素E琥珀酸酯诱导人胃癌细胞分化的作用

目的研究维生素E琥珀酸酯(VES)对胃癌细胞的诱导分化作用。方法5，10μg／ml维生素E琥珀酸酯处理人胃癌SGE-7901细胞，分别采用噻唑蓝(MTT)法检测细胞的生长；^3H-TdR掺入法测定DNA合成速率；流式细胞术检测细胞周期的分布；分光光度法检测分化标志酶的活性。结果VES可使细胞形态学改变；抑制SGC-7901细胞的生长。降低DNA的合成速率；使细胞出现G0／G1期停滞；分化标志酶碱性磷酸酶(ALP)、乳酸脱氢酶(LDH)活性降低。结论VES可诱导SGC-7901细胞分化并抑制其恶性生长。     

维生素D类似物EB1089对肝癌细胞增殖和凋亡的影响

目的探讨维生素D类似物EB1089对肝癌细胞增殖和凋亡的影响。方法体外培养维生素D3受体(VDR)阳性和阴性的肝癌细胞株HepG2和HCCT细胞,培养时添加100、10和1nmol/LEB1089,分别作用2、4和6d后,用四唑蓝比色实验(MTT)、平板克隆形成实验检测细胞的存活和生长;用反转录PCR(RTPCR)检测VDRmRNA的表达;用流式细胞仪和电子显微镜检测细胞凋亡。结果EB1089对VDRmRNA表达阳性的HepG2细胞的抑制率为175%～721%,对VDRmRNA表达阴性的HCCT细胞没有抑制作用;电镜结果显示EB1089可诱导HepG2细胞凋亡,流式细胞仪结果显示凋亡细胞的百分比为214%,并可阻滞细胞周期于G1期。结论EB1089对人肝癌细胞株HepG2的增殖具有抑制作用,可通过VDR受体抑制人肝癌细胞的增殖,并可诱导HepG2细胞凋亡。     

EGCG对人肝癌HepG2细胞增殖和侵袭的影响

目的探讨表没食子儿茶素没食子酸酯(EGCG)对肝癌HepG2细胞增殖和侵袭的影响及可能的分子机制。方法不同浓度EGCG对HepG2细胞进行干预后,CCK-8法检测EGCG对肝癌HepG2细胞增殖的影响;荧光显微镜及流式细胞术(FCM)Annexin V-FITC/PI双染法检测EGCG对HepG2细胞的凋亡诱导作用;Transwell侵袭实验检测EGCG对HepG2细胞侵袭的影响;ELISA法检测HepG2细胞中基质金属蛋白酶-2(MMP-2)和血管内皮生长因子(VEGF)的表达水平。结果EGCG各浓度组干预HepG2细胞24、48h后,细胞的生长增殖均明显受到抑制,其24h IC25值和IC50值分别为58.19和133.90mg/L。以60、135mg/L EGCG干预肝癌HepG2细胞24h后,FCM结果显示药物组细胞凋亡率明显高于对照组(P<0.05),荧光显微镜观察显示随着药物浓度的增加,细胞凋亡程度加重。Transwell侵袭实验显示,60、135mg/L EGCG对HepG2细胞侵袭力抑制率分别为69.47%和100%(P<0.05)。ELISA检测结果显示,EGCG干预后HepG2细胞上清液中MMP-2和VEGF表达水平下降(P<0.05)。结论 EGCG对人肝癌HepG2细胞具有抑制增殖和侵袭作用,其机制可能与EGCG诱导HepG2细胞凋亡和抑制肿瘤细胞中MMP-2及VEGF表达水平有关。     

Dietary (+)-catechin and BHT markedly increase alpha-tocopherol concentrations in rats by a tocopherol-omega-hydroxylase-independent mechanism

The effects of dietary (+)-catechin (CAT) and BHT on plasma and tissue concentrations of alpha-tocopherol (alpha-T), gamma-tocopherol (gamma-T) and cholesterol (C) were studied in male Sprague-Dawley rats. The rats were fed the compounds during a 4-wk period at concentrations of 2 g/kg in standardized diets, low but adequate in vitamin E, with 2 g/kg cholesterol. The CAT-regimen did not affect weight gain, feed intake or organ weights. BHT did not affect feed intake but lowered the body weight and the amount of liver lipids and increased the weights of livers and lungs relative to the body weight. Rats consuming CAT had 2.5-3.5-fold increased plasma, liver and lung alpha-T concentrations, but C concentrations remained unchanged. BHT-feeding resulted in 2.4- and 1.7-fold elevation in alpha-T but approximately 50% decrease in gamma-T concentrations in blood plasma and liver, respectively. BHT also lowered total C in the liver without affecting the concentration of C in the liver lipids. To investigate whether the alpha-T-sparing action of the studied compounds was due to the inhibition of tocopherol-omega-hydroxylase, HepG2 cells were incubated with CAT or BHT in the presence of delta-tocopherol (delta-T) and the 3'- and 5'-delta-carboxychromanol metabolites in the media were analyzed by GC/MS. Neither CAT nor BHT inhibited tocopherol-omega-hydroxylase activity in hepatocyte cultures; CAT was also inactive in a rat microsomal assay. In conclusion, both dietary CAT and BHT markedly increased alpha-T concentrations in plasma and organs of Sprague-Dawley rats by a mechanism that apparently does not involve inhibition of tocopherol-omega-hydroxylase, a key enzyme in tocopherol catabolism.     

Effects of gamma-tocopherol supplementation on thrombotic risk factors

OBJECTIVE: The antioxidant activity of vitamin E is derived primarily from alpha-tocopherol (alpha-T) and gamma-tocopherol (gamma-T). Results of epidemiological studies have demonstrated an inverse relationship between vitamin E intake and coronary disease. However, the results of clinical trials using alpha-T are equivocal. We determined the effect of 5 weeks of 100 mg/d or 200 mg/d gamma-T supplementation on thrombotic markers such as platelet reactivity, lipid profile and the inflammation marker C-reactive protein (CRP). METHODS AND RESULTS: Fourteen healthy subjects consumed 100 mg/day while 13 consumed 200 mg/d of gamma-T and 12 received placebo (soybean capsules with less than 5 mg/d gamma-T) in a double-blinded parallel study design. Fasting pre and post dose blood samples were analysed. Blood gamma-T concentrations increased significantly (p<0.05) relative to dose during the intervention period. Both groups receiving active ingredients showed significantly lower platelet activation after supplementation (p<0.05). Subjects consuming 100 mg/d gamma-T had significantly decreased LDL cholesterol, platelet aggregation and mean platelet volume (MPV) (p<0.05). Little effect of gamma-T was observed on other parameters. CONCLUSIONS: These data suggest that gamma-T supplementation may have a permissive role in decreasing the risk of thrombotic events by improving lipid profile and reducing platelet activity.     

Inhibitory effect of delta-tocotrienol,a HMG CoA reductase inhibitor,on monocyte-endothelial cell adhesion

We have previously shown that alpha-tocotrienol (alpha-T3), a vitamin E analogue and HMG CoA reductase (HMGR) inhibitor, markedly inhibited monocyte-endothelial cell adhesion, a process that was reversed with the addition of mevalonate intermediates involved in protein prenylation. Since delta-T3 and gamma-T3 possess greater HMGR inhibition than alpha-T3, we postulated that these analogues might have a greater effect on protein prenylation, and thus on monocyte adhesion and endothelial adhesion molecule expression in comparison to alpha-T3. Hence, we pursued to investigate the effect of various analogues of tocotrienol (alpha, gamma, delta) on monocytic cell adhesion and expression of adhesion molecules using a human umbilical vein endothelial cell-line, EA.hy926, as the model system. Relative to alpha-T3, delta-T3 displayed a more profound inhibitory effect on monocytic cell adherence using a 15 micromol/L concentration within 24 h (delta: 42 +/- 5%; alpha: 26 +/- 8% vs. control). This inhibitory action was reversed by co-incubation with farnesol and geranylgeraniol, suggesting a role for prenylated proteins in the regulation of monocyte adhesion. To further evaluate the effect of tocotrienols on the vascular endothelium, we measured the surface expression of adhesion molecules. Compared to alpha-T3, delta-T3 markedly inhibited the expression of VCAM-1 (delta: 57 +/- 6%; alpha: 37 +/- 10% vs. control) and E-selection (delta: 36 +/- 3%; alpha: 18 +/- 6% vs. control) in TNF-alpha activated endothelial cells. The above result suggests that delta-T3 is a potent and effective agent for the reduction of cellular adhesion molecule expression and monocytic cell adherence.     

Cigarette smoke alters human vitamin E requirements

Vitamin E is a lipophilic chain-breaking antioxidant that prevents lipid peroxidation. Although cigarette smoke is a potent source of oxidative stress that depletes vitamin E in vitro, it is unclear whether it has a similar effect in vivo, particularly in humans. Therefore, this review will discuss the role of cigarette smoke on gamma-tocopherol (gamma-T) nitration, its effect on alpha-tocopherol (alpha-T) biokinetics in smokers, and the changes in the synthesis, plasma concentrations, and urinary excretion of the vitamin E metabolite (CEHC; carboxy-ethyl-hydroxy-chroman). Last, the possibility of CEHC as a biomarker of vitamin E status will be assessed as will the question whether smokers have increased dietary requirements of vitamin E.     

Walnuts reduce aortic ET-1 mRNA levels in hamsters fed a high-fat, atherogenic diet

Walnut consumption is associated with reduced coronary vascular disease (CVD) risk; however, the mechanisms responsible remain incompletely understood. Recent clinical studies suggested that these mechanisms involve non-plasma lipid-related effects on endothelial function. Male Golden Syrian hamsters (12 groups, n=10-15) were fed for 26 wk atherosclerotic, high-fat, hyperlipidemic diets with increasing concentrations of whole walnuts (61-150 g/kg diet), or alpha-tocopherol (alpha-T, 8.1-81 mg/kg diet) and single diets with either walnut oil (32 g/kg diet) or pure gamma-tocopherol (gamma-T; 81 mg/kg diet) added. Aortic endothelin 1 (ET-1), an important endothelial regulator, was assayed as mRNA. Aortic cholesterol ester (CE) concentration along with other vascular stress markers (Cu/Zn and Mn superoxide dismutase, biliverdin reductase) and plasma lipid concentrations were determined. Hyperlipidemia (plasma LDL cholesterol approximately 6 times normal) occurred in all groups. Aortic CE concentration, a measure of atherosclerotic plaque, was highest in the lowest alpha-T only group and declined significantly with increasing alpha-T. The aortic CE of all walnut groups was decreased significantly relative to the lowest alpha-T only group but showed no dose response. The diets did not produce changes in the other vascular stress markers, whereas aortic ET-1 mRNA levels declined dramatically with increasing dietary walnuts (to a 75% reduction in the highest walnut content group compared with the lowest alpha-T group) but were unaltered in the alpha-T groups or gamma-T group. The study results are consistent with those of human walnut feeding studies and suggest that the mechanisms underlying those results are mediated in part by ET-1-dependent mechanisms. The contrasting results between the alpha-tocopherol or gamma-tocopherol diets and the walnut diets also make it unlikely that the non-plasma lipid-related CVD effects of walnuts are due to their alpha-tocopherol or gamma-tocopherol content. Finally, the results indicate that the walnut fat compartment is a likely location for the components responsible for the reduced aortic CE concentration.     

Supplementation of diets with alpha-tocopherol reduces serum concentrations of gamma- and delta-tocopherol in humans

Despite promising evidence from in vitro experiments and observational studies, supplementation of diets with alpha-tocopherol has not reduced the risk of cardiovascular disease and cancer in most large-scale clinical trials. One plausible explanation is that the potential health benefits of alpha-tocopherol supplements are offset by deleterious changes in the bioavailability and/or bioactivity of other nutrients. We studied the effects of supplementing diets with RRR-alpha-tocopheryl acetate (400 IU/d) on serum concentrations of gamma- and delta-tocopherol in a randomized, placebo-controlled trial in 184 adult nonsmokers. Outcomes were changes in serum concentrations of gamma- and delta-tocopherol from baseline to the end of the 2-mo experimental period. Compared with placebo, supplementation with alpha-tocopherol reduced serum gamma-tocopherol concentrations by a median change of 58% [95% CI = (51%, 66%), P < 0.0001], and reduced the number of individuals with detectable delta-tocopherol concentrations (P < 0.0001). Consistent with trial results were the results from baseline cross-sectional analyses, in which prior vitamin E supplement users had significantly lower serum gamma-tocopherol than nonusers. In view of the potential benefits of gamma- and delta-tocopherol, the efficacy of alpha-tocopherol supplementation may be reduced due to decreases in serum gamma- and delta-tocopherol levels. Additional research is clearly warranted.     

维生素E琥珀酸酯抑制人胃癌细胞生长的研究

采用体外细胞培养的方法,观察维生素Ｅ琥珀酸酯（ＶＥＳ）对人胃癌细胞（ＳＧＣ－７９０１）生长的影响,并利用ＭＴＴ法比较ＶＥＳ和α－生育酚对胃部细胞生长的作用效果。结果表明：ＶＥＳ（５、１０和２０μｇ／ｍｌ）对胃癌细胞生长有明显抑制作用,而α－生育酚对同样剂量对肿瘤细胞生长无抑制作用。这提示ＶＥＳ具有不同于维生素原抗癌机理。     

Supplementation with mixed tocopherols increases serum and blood cell gamma-tocopherol but does not alter biomarkers of platelet activation in subjects with type 2 diabetes

BACKGROUND: Some studies have shown potential benefit of vitamin E on platelet function, but several clinical trials failed to show improved cardiovascular outcome with alpha-tocopherol supplementation. Gamma-tocopherol, a major dietary form of vitamin E, may have protective properties different from those of alpha-tocopherol. OBJECTIVE: We compared the effects of supplementation with alpha-tocopherol (500 mg) and a gamma-tocopherol-rich compound (500 mg, containing 60% gamma-tocopherol) on serum and cellular tocopherol concentrations, urinary tocopherol metabolite excretion, and in vivo platelet activation in subjects with type 2 diabetes. DESIGN: Fifty-eight subjects were randomly assigned to receive either 500 mg alpha-tocopherol/d, 500 mg mixed tocopherols/d, or matching placebo. Serum, erythrocyte, and platelet tocopherol and urinary metabolite concentrations were measured at baseline and after the 6-wk intervention. Soluble CD40 ligand, urinary 11-dehydro-thromboxane B2, serum thromboxane B2, soluble P-selectin, and von Willebrand factor were measured as biomarkers of in vivo platelet activation. RESULTS: Serum alpha-tocopherol increased with both tocopherol treatments. Serum and cellular gamma-tocopherol increased 4-fold (P < 0.001) in the mixed tocopherol group, whereas red blood cell gamma-tocopherol decreased significantly after alpha-tocopherol supplementation. Excretion of alpha-carboxyethyl-hydroxychroman increased significantly after supplementation with alpha-tocopherol and mixed tocopherols. Excretion of gamma-carboxyethyl-hydroxychroman increased significantly after supplementation with mixed tocopherols and after that with alpha-tocopherol, which may reflect the displacement of gamma-tocopherol by alpha-tocopherol due to incorporation of the latter into lipoproteins in the liver. Neither treatment had any significant effect on markers of platelet activation. CONCLUSIONS: Supplementation with alpha-tocopherol decreased red blood cell gamma-tocopherol, whereas mixed tocopherols increased both serum alpha-tocopherol and serum and cellular gamma-tocopherol. Changes in serum tocopherol closely reflect changes in cellular concentrations of tocopherols after supplementation.