赖氨熊果酸增溶及对肝癌细胞增殖抑制作用

目的制备赖氨熊果酸并探讨其增溶作用及对肝癌Bel7402细胞增殖的抑制作用。方法采用共研磨法制备赖氨熊果酸,采用粉末X射线衍射法(PXRD)、红外光谱(IR)法对赖氨熊果酸进行表征,采用高效液相色谱法(HPLC)测定赖氨熊果酸的增溶作用,采用噻唑蓝(MTT)法观察赖氨熊果酸对肝癌Bel7402细胞增殖的抑制效果。结果共研磨法制备赖氨熊果酸得率为95.2%,纯度为98.2%,赖氨熊果酸的溶解度为49.69μg/m L,明显高于赖氨酸-熊果酸物理混合物在水中的溶解度1.87μg/m L;赖氨熊果酸对肝癌Bel7402细胞增殖的抑制作用明显增强,在浓度10、20μmol/L时,与相同浓度5-FU(阳性对照)比较,赖氨熊果酸对肝癌Bel7402细胞的抑制作用差异无统计学意义(P0.05)。结论赖氨熊果酸增溶作用明显,对肝癌细胞增殖具有明显抑制作用,作为新一代高效低毒的抗肿瘤药物具有一定开发价值。     

Bel7402肝癌细胞中共轭亚油酸异构体通过不同机制下调COX-2表达

[目的]采用体外细胞培养方法,研究c9,t11-CLA和t10,c12-CLA对肝癌细胞Bel7402的COX-2表达的影响和机制。[方法]采用逆转录聚合酶链反应(RT-PCR)和蛋白免疫印迹(Westernblot)方法检测c9,t11-CLA和t10,c12-CLA处理后的Bel7402细胞中COX-2 mRNA和蛋白质的表达以及PPARγ配体罗格列酮和GW9662对COX-2蛋白水平的影响。[结果]在100μmol/L的浓度下,c9,t11-CLA和t10,c12-CLA都可以降低COX-2mRNA和蛋白质的表达,罗格列酮可以下调COX-2蛋白水平,而PPARγ抑制性配体GW9662可以逆转c9,t11-CLA对于COX-2蛋白水平的下调作用,但是对于t10,c12-CLA的下调作用则没有影响。[结论]两种异构体对COX-2表达的下调作用很可能具有不同的机制。     

不同形态硒对结肠癌Lovo细胞粘附及迁移功能的影响

目的探讨L-硒甲基硒代半胱氨酸(L-SeMSC)和亚硒酸钠(SS)对结肠癌Lovo细胞粘附及迁移能力的影响。方法采用MTT法、划痕实验、粘附实验分别检测不同形态硒对Lovo细胞增殖、迁移、粘附能力的影响;采用Western blot检测CDC37、MMP2和MMP9蛋白的表达。结果两种硒具有抑制Lovo细胞增殖作用,且随着药物浓度的增加,抑制作用增强,相同浓度下,L-SeMSC对细胞抑制作用更强(P<0.05)。两种硒均能缩短Lovo细胞的迁移距离、降低细胞粘附率,与SS相比,在中(0.1 mg/L)、高(0.2 mg/L)浓度组,L-SeMSC的抑制Lovo细胞迁移和粘附的能力更强。不同浓度L-SeMSC均能抑制CDC37和MMP2的表达,且具有浓度依赖性;高浓度(0.2 mg/L)SS能抑制CDC37、MMP2和MMP9的表达,但作用弱于相同浓度L-SeMSC。结论 L-SeMSC和SS均能通过降低CDC37、MMP2和MMP9的表达抑制结肠癌Lovo细胞的粘附和迁移能力,相同浓度L-SeMSC作用强于SS。     

白毛藤对人肝癌Bel7402细胞增殖抑制及凋亡作用

目的 探讨白毛藤体外对人肝癌Bel7402细胞增殖与凋亡的影响及其作用机制.方法 采用四甲基偶氮噻唑蓝(MTT)法和荧光显微镜观察白毛藤对人肝癌Bel7402细胞的增殖抑制作用和其诱导Bel7402细胞凋亡情况;用反转录-聚合酶链式反应(RT-PCR)检测survivin、bax基因表达量的变化.结果 随白毛藤浓度增加、作用时间延长,对人肝癌Bel7402细胞的增殖抑制作用均明显增强(P＜0.01);4,8,16 mg/mL白毛藤组人肝癌Bel7402细胞凋亡率分别为10.98%、17.02%、26.78%,均高于对照组6.18%(P＜0.01);作用48 h后,4,8,16 mg/mL白毛藤组人肝癌Bel7402细胞中survivin基因灰度比值分别为(0.43±0.01)、(0.38±0.02)、(0.29±0.01).均低于对照组(0.65±0.05),差异均有统计学意义(P＜0.01);4,8,16 mg/mL,白毛藤组人肝癌Bel7402细胞中bax基因灰度比值分别为(0.47±0.05)、(0.56±0.04)、(0.73±0.06),均高于对照组(0.36±0.02),差异均有统计学意义(P＜0.01).结论 白毛藤可抑制人肝癌Bel7402细胞增殖、诱导其细胞凋亡,并能调控survivin及bax基因的表达.     

共轭亚油酸异构体的抗乳腺癌活性研究

目的 :　研究共轭亚油酸 (CLA)两种主要异构体 (9,1 1 - CLA和 1 0 ,1 2 - CLA)对两种人乳腺癌细胞 (MCF- 7,MDA- MB- 2 31 )和大鼠扩散乳腺癌细胞 (F3 )活性的异同。方法 :　采用MTT方法研究两种主要 CLA异构体在不同浓度、不同作用时间及不同给药方式下 ,抑制上述几种癌细胞增殖的能力。结果 :　 1 0 ,1 2 - CLA对所有的试验癌细胞均呈现较强的抑制活性 ,而 9,1 1 -CLA仅对 MCF- 7呈现抑制活性 ,对 MDA- MB- 2 31和 F3 基本无活性 ;1 0 ,1 2 - CLA对 F3 的抑制活性与处理时间和给药方式密切相关 ,加药后不更换培养液处理 72 h可使其抑制能力达到最大值 ;细胞周期研究显示 1 0 ,1 2 - CLA对 MCF- 7呈微弱的 G0 / G1期抑制 ,而对 F3 细胞周期则无影响。结论 :　 CLA的两种主要异构体在该实验条件下对上述三种癌细胞增殖抑制能力和作用机制有着明显的差异 ,其中 1 0 ,1 2 - CLA活性明显强于 9,1 1 - CLA。 1 0 ,1 2 - CLA是通过其代谢物对 F3 的增殖起抑制作用     

环氧合酶-2在c9,t11-共轭亚油酸抑制胃腺癌细胞增殖中的作用

目的　采用体外细胞培养方法,研究不同浓度c9,t11 共轭亚油酸对人胃腺癌细胞(SGC- 790 1)的亚油酸代谢途径中限速酶———环氧合酶(COX -2 )及其产物前列腺素E2 (PGE2 )的影响。方法　采用逆转录聚合酶链反应(RT- PCR)和Westernblot方法检测不同浓度c9,t11 CLA处理后的SGC 790 1细胞中COX- 2mRNA和蛋白的表达,放射免疫方法检测细胞分泌的PGE2 浓度。结果　在2 5、5 0、10 0和2 0 0 μmol L浓度时,c9,t11 CLA均可下调COX 2mRNA和蛋白的表达,同时降低细胞中PGE2 的分泌。结论　环氧合酶2是c9,t11 CLA抑制肿瘤细胞增殖的作用靶点。     

苦参碱对肝癌细胞增殖抑制作用研究

目的在体外观察苦参碱对BEL-7402肝癌细胞增殖的影响。方法用含不同浓度（0～3.2mmol/L）苦参碱的培养液在体外培养BEL-7402细胞24h,通过MTT试验测定其对BEL-7402细胞增殖的影响作用。结果在上述使用浓度范围内,苦参碱对BEL-7402细胞增殖的抑制率为0～36.7%。结论苦参碱在体外可抑制BEL-7402细胞的增殖,且呈剂量依赖性。     

刺儿菜提取物抗BEL-7402肿瘤细胞活性的研究

目的研究刺儿菜提取物对BEL-7402肿瘤细胞生长的抑制作用,观察提取物作用下肿瘤细胞的凋亡情况。方法利用MTT比色法测定提取物对BEL-7402肝癌细胞生长的抑制率;采用透射电镜观察法和TUNEL染色法研究了刺儿菜提取物诱导肝癌细胞凋亡。结果刺儿菜提取物10,20,40,80mg/L对肝癌细胞有明显的抑制作用,随提取物浓度的增加和培养时间的延长,抑制作用逐渐增强;对该提取液处理培养24,48,72,96h的细胞凋亡指数有显著差异。结论刺儿菜提取物对肿瘤细胞生长有抑制作用。     

L-硒-甲基硒代半胱氨酸和亚硒酸钠对大肠癌Lovo细胞生长及凋亡的影响

目的通过有机硒—L-硒甲基硒代半胱氨酸(L-Se-methylselenocysteine,L-SeMSC)和无机硒—亚硒酸钠(sodium selenite,SS)对大肠癌Lovo细胞的作用,研究不同硒对Lovo细胞活力、生长及凋亡相关蛋白的影响。方法将不同硒与Lovo细胞作用24h后,MTT法测定细胞活力,Western blot测定细胞生长相关蛋白CDK2、CDK4、CyclinD1及细胞凋亡相关蛋白Caspase-3,Caspase-8,Caspase-9蛋白的表达量。结果两种硒均能抑制Lovo细胞活力,且相同浓度L-SeMSC与SS相比,对细胞活力抑制作用更强(P0.05);两种硒均能降低CDK4及CyclinD1蛋白表达,且L-SeSMC能降低CDK2表达,但0.2mg/LSS却提高CKD2表达;两种硒均能提升Caspase-3,Caspase-8,Caspase-9的表达,相同浓度(0.1mg/L与0.2mg/L)L-SeMSC与SS相比,对Caspase-3和Caspase-8提升作用更显著(P0.05)。结论与SS相比,L-SeMSC对Lovo细胞具有更强的活性抑制作用和促凋亡作用。     

共轭亚油酸对乳腺癌细胞系SKBR 3增殖的抑制作用

目的比较共轭亚油酸(CLA)和亚油酸(LA)对乳腺癌细胞生长的影响,以探讨共轭亚油酸的抗肿瘤作用.方法采用体外培养乳腺癌细胞系SKBR 3,用细胞生长曲线和MTT试验观察细胞的生长状况,透射电镜观察细胞形态学变化,流式细胞仪检测细胞周期.结果细胞生长曲线和MTT试验均表明,CLA对SKBR 3细胞生长有明显的抑制作用,且随着作用浓度和作用时间的增加,抑制作用增强.细胞生长曲线显示,CLA各浓度组(200、100、50 μmol·L-1)作用5 d后抑制率分别为75.0%、57.9%、33.1%;LA各浓度组(200、100、50 μmol·L-1)作用5 d后抑制率分别为38.4%、25.4%、10.1%.电镜观察可见CLA可以引起细胞凋亡,而LA未观察到明显的病理变化.流式细胞仪检测表明,200 μmol·L-1可使细胞周期分布发生明显改变.作用6 d后G1期为72.2%,并出现凋亡峰,而LA组及对照组G1期分别为为56.6%,50.6%.结论 CLA可能通过抑制细胞增殖,诱导细胞凋亡等途径抑制癌症的发展.     

共轭亚油酸抑制苯并(a)芘诱导小鼠前胃癌的研究

目的：探讨不同的构成的共轭亚油酸（CLA）对苯并（a)芘[B(a)P]诱导的小鼠前胃癌的抑制作用及可能机制。方法：用B(a)P在昆明种小鼠体内建立前胃癌模型,观察不同构成的CLA对小鼠前胃癌形成的抑制作用,同时采用蛋白印迹法分析小鼠前胃组织中的蛋白表达情况。结果：B(a)P组、75％纯度C9,T11-CAL组、98％纯度c9,t11-CAL组、98％纯度t10,c12-CLA组的前胃肿瘤发生率分别为100．0％、75．0％、69．2％、53．8％;蛋白印迹法分析结果表明,CAL抑制ERK－1的表达,促进MKP－1的表达,而对MEK－1的表达无明显影响,结论：不同构成的CAL对B(a)P诱导小鼠前胃癌均具有抑制作用;CAL影响MAPKs级联反应ERKs及此途径负调控子MKP－1蛋白的表达可能是其抑制肿瘤作用的机制之一。     

Mechanisms of Anticancer Activity of a Fatty Acid Mixture Extracted from Hen Egg Yolks Enriched in Conjugated Linoleic Acid Diene (CLA) against WM793 Melanoma Cells

The conjugated linoleic acid (CLA) diene is a biologically active compound with proven health-promoting effects. In terms of anticancer properties, it has been shown that CLA reduces the proliferation of cancer cells. In this study, it has been demonstrated that a mixture of fatty acids, isolated from chicken egg yolk enriched in CLA isomers by biofortification, reduces (by 30.5%) the proliferation of human melanoma cancer cells line WM793 to a greater extent than a mixture of fatty acids not containing these isomers. At the same time, the tested fatty acid mixtures show no effect on human normal BJ fibroblast cells. For the first time, the genes with increased expression have been identified and the proteins have been activated by the fatty acid mixture of CLA-enriched egg yolk, mainly responsible for mitochondrial pathway-dependent apoptosis.     

Conjugated linoleic acid isomers inhibit platelet-derived growth factor-induced NF-κB transactivation and collagen formation in human vascular smooth muscle cells

Background Atherosclerosis is characterized by extensive thickening of the arterial intima partially resulting from deposition of collagen by vascular smooth muscle cells (SMCs). Polyunsaturated fatty acids stimulate collagen formation through NF-κB activation. Aim of the study The present study aimed to explore the effect of conjugated linoleic acids (CLAs) which are known to inhibit NF-κB activation on collagen formation by SMCs. Methods Vascular SMCs were cultured with 50 μmol/l of CLA isomers (c9t11-CLA, t10c12-CLA) or linoleic acid (LA) and analysed for collagen formation and NF-κB p50 transactivation. Results Treatment with CLA isomers but not LA significantly reduced PDGF-stimulated [³H] proline incorporation into cell layer protein of SMCs without altering cell proliferation. Simultaneous treatment with the PPARγ inhibitor T0070907 abrogated this effect. Treatment of SMCs with c9t11-CLA and t10c12-CLA significantly reduced PDGF-induced NF-κB p50 activation. Conclusions CLA isomers inhibit PDGF-stimulated collagen production by vascular SMCs, which is considered to be a hallmark of atherosclerosis, in a PPARγ-dependent manner. Whether inhibition of the NF-κB-pathway is of significance for the reduction of collagen formation by CLA isomers needs further investigation.     

Intestinal bifidobacteria that produce trans-9, trans-11 conjugated linoleic acid: a fatty acid with antiproliferative activity against human colon SW480 and HT-29 cancer cells

Bifidobacterium breve species of human intestinal origin have the ability to synthesize cis-9, trans-11 (c9, t11) conjugated linoleic acid (CLA) from free linoleic acid. In this study, the ability of Bifidobacterium species to isomerize C(18) polyunsaturated fatty acids was investigated, and the antiproliferative activities of the two main microbially produced CLA isomers were assessed. Linoleic acid was converted principally to c9, t11 CLA and lesser amounts of t9, t11 CLA, whereas c9, t11 CLA was converted mainly to t9, t11 CLA. Likewise, t10, c12 CLA was converted principally to t9, t11 CLA, which was incorporated into the bacterial cell membranes. To examine the antiproliferative effect of the two main CLA isomers formed, SW480 and HT-29 human colon cancer cells were cultured in the presence of c9, t11 CLA and t9, t11 CLA. The t9, t11 CLA had a more potent antiproliferative effect than c9, t11 CLA. It is tempting to suggest that the ability of Bifidobacterium to produce such bioactive metabolites may be associated with the beneficial effects of bifidobacteria present in the human gastrointestinal tract.     

trans-10, cis-12 conjugated linoleic acid reduces insulin-like growth factor-II secretion in HT-29 human colon cancer cells

We previously demonstrated that a mixture of conjugated linoleic acid (CLA) isomers decreases colon cancer incidence in rats treated with 1,2-dimethylhydrazine. Our in vitro studies have also shown that CLA inhibits the growth of HT-29 cells, a human colon cancer cell line. When we compared the individual potencies of the two main isomers found in the mixture of CLA isomers (e.g., cis-9, trans-11 [c9t11] and trans-10, cis-12 [t10c12]), t10c12 CLA decreased viable cell numbers in a dose-dependent manner. By contrast, c9t11 CLA had no effect. Therefore, the present study examined whether the decreased cell growth is related to changes in secretion of insulin-like growth factor (IGF)-II and/or IGF-binding proteins (IGFBPs) that have been shown to regulate HT-29 cell proliferation. Cells were incubated in serum-free medium with various concentrations of the individual CLA isomers, and immunoblot analysis of 24-hour, serum-free, conditioned media using a monoclonal anti-IGF-II antibody was performed. HT-29 cells secreted both mature 7,500 apparent molecular weight (M(r)) and higher-M(r) forms of IGF-II. t10c12 CLA decreased the levels of the higher-M(r) and the mature form of IGF-II in a dose-dependent manner, whereas c9t11 CLA had no effect. Ligand blot analysis of conditioned medium using (125)I-IGF-II revealed that the production of IGFBP-2 and IGFBP-4 was also decreased by t10c12 CLA, whereas c9t11 CLA had no effect. Exogenous IGF-II abrogated the growth inhibition induced by t10c12 CLA. These results indicate that inhibition of HT-29 cell growth by t10c12 CLA may be mediated by decreasing IGF-II secretion in these cells.     

共轭亚油酸营养再分配作用及其安全性

共轭亚油酸(CLA)是一组具有共轭不饱和双键的亚油酸的位置和结构异构体,具有降脂增肌、促进生长、抗癌、抗动脉粥样硬化及调节免疫等生理功能。本文总结了CLA营养再分配活性及其安全性评价的研究成果,从分子药理水平,分析了CLA的作用机制与作用方式。     

Effect of CLA isomers and their mixture on aging C57Bl/6J mice

Background  

Dietary supplements containing conjugated linoleic acid (CLA) are widely promoted for weight loss management over the counter. Recently, FDA approved the CLA as Generally Recognized as Safe category so that it can be used in various food and beverages. The combined effect of CLA isomers have been studied extensively in animals and humans, however, the role of individual isomers remains unraveled.     

Beef conjugated linoleic acid isomers reduce human cancer cell growth even when associated with other beef fatty acids

Although many data are available concerning anticarcinogenic effects of industrial conjugated linoleic acid (CLA), few studies have reported the antitumour properties of CLA mixtures originating from ruminant products. The aim of the present study was to investigate the in vitro antiproliferative effects of beef CLA mixtures on breast, lung, colon, melanoma and ovarian human cancer cell lines. For this purpose, four fatty acid (FA) extracts prepared from beef lipid and varying in their CLA composition, their corresponding purified CLA-enriched fractions, and mixtures of pure synthetic CLA, the composition of which reproduced that of the four selected beef samples, were tested on cancer cell lines. Cancer cells were exposed for 48 h to medium containing 100 microm-FA and their proliferation was determined by quantifying cellular DNA content (Hoechst 33342 dye). Compared with cells incubated without FA, the number of cancer cells was reduced from 25 to 67 % (P<0.0001) following FA treatment. Antiproliferative effects of CLA mixtures varied in magnitude according to the source of FA, the CLA composition and the cell lines. CLA mixtures naturally present in beef inhibited the proliferation of human cancer cell lines, a high content in cis-trans isomers allowing the most important antiproliferative effect. Beef total FA exhibited a greater growth-inhibitory activity than their corresponding CLA-enriched fractions. These results suggested that either beef FA other than beef CLA could possess antiproliferative properties and/or the existence of complementary effects of non-conjugated FA and CLA, which could favour the antiproliferative properties of beef total FA.