矽肺患者巨噬细胞培养上清通过p38MAPK对人胚肺成纤维细胞增殖的影响

目的 探讨矽肺患者巨噬细胞(AM)培养上清是否通过激活p38丝裂原活化蛋白激酶(p38MAPK)信号转导通路促进人胚肺成纤维细胞(HELF)的增殖作用,进而参与矽肺纤维化的发生发展过程.方法 以支气管肺泡灌洗法收集矽肺患者AM,在体外用含有SiO2(50 μg/ml)的DMEM培养基和不含SiO2的DMEM培养基培养18h,然后收集培养的AM上清液.用组织贴块法获取培养HELF,与AM上清液共同孵育,将HELF分为空白对照组、AM组、SiO2+AM组、SB203580+SiO2+AM组,用噻唑蓝(MTT)法和流式细胞仪法分别检测各组HELF的增殖情况和细胞周期.结果 SiO2+AM组细胞的平均吸光度值为0.48±0.03,与空白对照组(0.29±0.01)、AM组(0.38±0.02)、SB203580+SiO2+AM组(0.33±0.03)的差异有统计学意义(P<0.05);SiO2+AM组细胞增殖指数为18.12±0.82,与空白对照组(9.24±0.48)、AM组(14.76±0.43)、SB203580+SiO2+AM组(11.71±0.70)的差异有统计学意义(P<0.05).SB203580+SiO2+AM组的细胞吸光度值和细胞增殖指数均明显降低.结论 经SiO2刺激的矽肺患者AM培养上清通过激活p38MAPK信号转导通路可以促进HELF增殖.     

c9,t11-共轭亚油酸对巨噬细胞因子表达的影响

目的 研究c9，t11-共轭亚油酸(Conjugated linoleic acid，CLA)对C57小鼠巨噬细胞杀伤黑色索瘤细胞(B16-MB)能力的影响。方法用0，25，50，75，100μmol／L CLA处理巨噬细胞24h后，分别用MTT法检测CLA处理的巨噬细胞对B16-MB细胞的杀伤能力；RT-PCR方法检测C57小鼠巨噬细胞细胞因子IL-6、TNF-α和iNOS mRNA表达。结果 在100，75μmol／L CLA处理后，巨噬细胞对肿瘤的杀伤率分别为18％和14．5％：同时，IL-6、TNF-α和iNOS mRNA表达增加。结论 c9，tll-CLA增强C57小鼠巨噬细胞对B16-MB细胞的杀伤能力，并与其诱导IL-6、TNF-α和iNOS mRNA的表达有关。推测CLA发挥抗肿瘤作用可能与其参与机体免疫调节作用有关．     

电磁脉冲对BV-2细胞形态和分泌功能的影响及其机制探讨

目的 研究电磁脉冲( electromagnetic pulse,EMP)对小鼠BV-2小胶质细胞形态及分泌功能的影响,并初步探讨其作用机制.方法 离体培养的BV-2细胞经200 kV/m EMP辐照200次,分别在辐照后1、6、12、24h收集细胞培养上清及细胞.倒置显微镜下观察细胞形态变化,ELISA法检测培养上清中肿瘤坏死因子-α(TN F-α)、白细胞介素(IL)-1β、IL-10等细胞因子水平的变化,硝酸还原酶法检测培养上清中一氧化氮(NO)水平,DCFH-DA探针检测活性氧,免疫印迹(Western-blot)法检测细胞外信号调节激酶(ERK)、c -Jun氨基末端激酶(JNK)、p38磷酸化水平和蛋白表达量的变化.应用p38抑制剂( SB203580)预处理细胞后再进行EMP辐照,然后检测培养上清中NO水平和活性氧的生成.结果 EMP辐照后1、6和12h,部分小胶质细胞出现胞体变大、突触变粗变短,且活化细胞比例与假辐照组相比明显增加,差异有统计学意义(P＜0.05);EMP辐照后细胞培养上清中TNF-α、IL-1β、IL-10等细胞因子水平未发生明显改变,但活性氧检测结果显示,与假辐照组(小胶质细胞平均荧光强度10.34)相比,EMP辐照后1h小胶质细胞荧光强度(平均荧光强度21.56)明显增加,6h达峰值(平均值为32.46),12h开始恢复(平均荧光强度24.36),差异均有统计学意义(P＜0.05),24h恢复至假辐照水平;EMP辐照后NO水平的变化与活性氧一致,辐照后1h开始增加,6h达峰值,12h开始恢复,24h恢复至假辐照组水平;蛋白杂交结果显示,EMP辐照后1、6h,p38的磷酸化水平和蛋白水平较假辐照组明显增加,差异有统计学意义(P＜0.05),ERK和JNK无明显变化.应用p38抑制剂SB203580预处理细胞,明显抑制了EMP诱导的小胶质细胞对活性氧和NO的产生,活性氧水平除6h组未恢复至假辐照水平外,其他各组均恢复至假辐照水平,NO水平各组均恢复至假辐照组水平.结论 EMP辐照可活化小胶质细胞并且促进其对NO和活性氧的生成,p38信号通路参与了此过程.     

共轭亚油酸对肿瘤细胞亚油酸代谢途径中限速酶的影响

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(c9,t11-CLA)对人胃腺癌细胞(SGC-7901)中亚油酸代谢途径的限速酶的影响。方法用200、100、50和25μmol／L浓度的c9,t11-CLA处理SGC-7901细胞24h,四甲基偶氮唑盐实验检测c9,t11-CLA对SGC-7901细胞增殖的抑制作用,采用逆转录聚合酶链反应检测c9,t11-CLA对SGC-7901细胞中亚油酸代谢途径的Δ6-脱氢酶、△5-脱氢酶、环氧合酶(COX)-1、COX-2和5-脂氧合酶(5-LOX)mRNA表达的影响。结果在200、100、50和25μmol／L浓度时,c9,t11-CLA对SGC-7901增殖的抑制率分别为54．3％、20．5％、10．5％、2．93％;均可下调COX-2mRNA的表达,上调Δ6-脱氢酶、COX-1mRNA的表达,但对Δ5-脱氢酶和5-LOXmRNA表达的影响不显著。结论c9,t11-CLA可通过调节Δ6-脱氢酶和COX的表达抑制肿瘤细胞的增殖,说明c9,t11-CLA通过影响亚油酸代谢途径中限速酶的基因表达而改变类二十碳烷酸的形成,推测c9,t11-CLA影响肿瘤细胞中亚油酸代谢途径的限速酶是其发挥抗癌活性的另一作用机制。     

丝裂原活化蛋白激酶信号途径在苯并[a]芘影响内皮细胞热休克蛋白70基因表达中的作用

目的研究丝裂原活化蛋白激酶(mitogen activated protein kinase,MAPK)信号途径在苯并[a]芘(BaP)影响内皮细胞热休克蛋白70(HSP70)基因表达中的作用.方法猪主动脉内皮细胞以不同浓度BaP(0、0.1、0.5、1.0、5.0、10.0 μmol/L)染毒24 h,或以PD98059(10μmol/L)或SB203580(20μmol/L)预处理1 h后再加一定剂量的BaP共同孵育24 h.分别以Western-blot法检测各组细胞磷酸化细胞外信号调节蛋白激酶(extracellularsignal regulated protein kinase,ERK)、磷酸化c-Jun氨基末端激酶(c-Jun amino-terminal kinase,JNK)、磷酸化p38和HSP70表达水平的改变.结果随BaP浓度的升高,MAPK途径的3种磷酸化激酶表达水平出现不同程度的改变,其中磷酸化ERK1表达改变最明显,呈剂量反应性增加.BaP抑制内皮细胞HSP70表达,在中、高剂量(≥1.0μmol/L)暴露时,HSP70表达明显低于对照组,差异有显著性(P＜0.05);尽管ERK途径抑制剂PD98059可部分减弱BaP对HSP70表达的抑制效应,但经PD98059预处理后内皮细胞HSP70表达水平仍明显低于正常状态下的表达水平,差异亦有显著性(P＜0.05).结论BaP可能通过ERK1途径而影响内皮细胞HSP70基因表达,同时可能还有其他信号途径在BaP影响HSP70基因表达过程中起作用.     

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.     

共轭亚油酸对人胃腺癌细胞中环氧合酶的影响

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(CLA)对人胃腺癌细胞(SGC-7901)的亚油酸代谢途径中限速酶-环氧合酶(COX)表达的影响.方法采用逆转录聚合酶链反应(RT-PCR)和蛋白免疫印迹(Western blot)方法检测不同浓度c9,t11-CLA处理后的SGC-7901细胞中环氧合酶(COX)-1、(COX)-2mRNA和蛋白的表达.结果在200,100,50和25μmol/L浓度时,c9,t11-CLA均可下调COX-2 mRNA和蛋白的表达,与共轭亚油酸(CLA)浓度呈负相关;上调COX-1 mRNA的表达,并与CLA浓度呈正相关.结论 c9,t11-CLA的抗癌活性与其影响COX的表达有关.     

共轭亚油酸对胰岛素抵抗大鼠葡萄糖运载体4蛋白表达影响的研究

目的通过研究共轭亚油酸（conjugated linoleic acid,CLA）对饮食诱导胰岛素抵抗大鼠葡萄糖运载体4（glucose transporter4,GLUT4）蛋白表达的影响,探讨CLA抗糖尿病作用的机制。方法选用雄性Wistar大鼠,用随机数字表法按体重随机分为对照组、高脂组、高脂＋CLA组（每100g饲料含CLA分别为0．75g、1．50g．3．00g）,每组动物10只,观察CLA对胰岛素抵抗大鼠胰岛素、血糖水平的影响,并应用Western blot方法检测大鼠骨骼肌GLUT4蛋白的表达水平。结果高脂组大鼠血清胰岛素和糖血水平分别为（11．11±2．73）μU／ml、（5．09±0．66）mmol／L,CLA可降低肥胖大鼠的高胰岛素、高糖血症,低、中、高剂量组胰岛素水平分别为（6．99±1．77）μU／ml、（7．36±1．48）μU／ml、（7．85±1．60）μU／ml,血糖水平分别为（4．28±0．72）mmol／L、（4．18±0．55）mmol／L、（4．06±0．63）mmol／L,且高脂组大鼠骨骼肌GLUT4蛋白的表达水平较基础组降低,CLA可增加高脂组大鼠骨骼肌GLUT4蛋白的表达水平。结论CLA可通过增加胰岛素抵抗大鼠骨骼肌GLUT4蛋白的表达水平,改善胰岛素抵抗。     

共轭亚油酸强化乳对小鼠体脂及血脂的影响

目的:探讨共轭亚油酸(CLA)强化乳对小鼠体脂及血脂的影响。方法:选4w龄的雄性昆明种小鼠40只,随机分成四组,每组10只,分别在牛奶中添加0%、0.1%、0.5%、1.0%的CLA喂养小鼠4w后,测定体重、腹脂重、体脂含量、饲料利用率及血浆甘油三酯(TG)、总胆固醇(TC)、高密度脂蛋白胆固醇含量(HDL)与脂蛋白脂酶(LPL)活性。结果:小鼠的体增重、腹脂重、体脂含量及饲料利用率都随着乳中CLA的添加量增加而降低,与对照组比,当添加0.5%时达到显著差异;在乳中添加CLA可以降低小鼠血浆中的TG与TC含量,提高HDL含量和LPL活性,降低动脉硬化指数(TC-HDL/HDL)。TC含量在添加0.5%CLA时最低,其它的血脂指标在添加1.0%CLA时效果最好。结论:共轭亚油酸强化乳有降低体脂和血脂的作用。     

共轭亚油酸对鼠黑色素瘤细胞转移特性的影响

:采用体外培养技术 ,模拟体内肿瘤转移过程 ,研究共轭亚油酸 (conjugated linoliec acid,CL A)对鼠黑色素瘤 B16 - MB高转移细胞系的转移特性 ,从而探讨 CL A预防肿瘤转移的可能途径。研究结果表明 ,剂量为 10 0 μmol/ L、2 0 0 μmol/ L 的 CL A可抑制肿瘤细胞增殖、恢复肿瘤细胞间通讯功能、抑制肿瘤细胞向细胞外基质成分 [层粘连蛋白 (fibronectin,FN)和纤维粘连蛋白 (laminin,L N) ]的黏附。     

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。     

共轭亚油酸对人胃腺癌细胞侵袭及转移相关基因表达的影响

目的 研究c9,t11－共轭亚油酸（CLA）对人胃腺癌细胞（SGC－7901）侵袭能力的影响,探讨其抑制肿瘤转移的可能机制。方法 用重组基底膜侵袭实验评价癌细胞侵袭能力;用逆转录聚合酶链反应（RT－PCR）检测SGC－7901细胞中组织基质金属蛋白酶抑制剂（TIMP）－1、TIMP－2和nm23-H1mRNA的表达。结果 在200、100和50μmol/L浓度时,c9,t11-CLA对SGC－7901细胞侵袭重组基底膜的抑制率分别为53.7%、40.9%和29.3%。c9,t11-CLA可诱导SGC－7901细胞中TIMP－1、TIMP－2和nm23-H1mRNA的表达。结论 c9,t11-CLA抑制SGC-7901细胞侵袭重组基底膜。c9,t11-CLA的抗侵袭活性与诱导肿瘤细胞中TIMP－1、TIMP－2和nm23-H1mRNA的表达等有关。     

共轭亚油酸异构体对人类癌细胞增殖的抑制作用研究

目的研究不同共轭亚油酸对人类胃癌细胞AGS、肝癌细胞Bel7402和肠癌细胞Lovo细胞增殖的影响。方法采用体外培养的AGS、Bel7402和Lovo细胞为模型,测定不同处理后活细胞数量。结果研究采用的5种共轭亚油酸异构体都对AGS、Bel7402和Lovo细胞增殖表现出了抑制作用,对AGS的抑制作用要普遍高于对Bel7402和Lovo的抑制作用,并且浓度不同,抑制作用也具有差异。结论五种共轭亚油酸异构体都具有抑制癌细胞增殖的作用,并且这种作用受到共轭亚油酸浓度的影响。     

The effects of conjugated linoleic acid on human health-related outcomes

Conjugated linoleic acid (CLA) is a collective term for a mixture of positional and geometric isomers of conjugated dienoic derivatives of linoleic acid. CLA has received considerable attention as a result of animal experiments that report anti-carcinogenic, anti-atherogenic and anti-diabetic properties, and modulation of body composition and immune function. Several studies of CLA supplementation in human subjects have now been published, but in contrast to animal studies there has been marked variation between reports on the health-related outcomes. The consensus from seventeen published studies in human subjects is that CLA does not affect body weight or body composition. Some detrimental effects of the trans-10,cis-12 CLA isomer have also been reported in terms of altered blood lipid composition and impaired insulin sensitivity. Finally, CLA has only limited effects on immune functions in man. However, there have been reports of some interesting isomer-specific effects of CLA on the blood lipid profile, but not on immune function. These isomer-specific effects need further investigation. Until more is known, CLA supplementation in man should be considered with caution.     

共轭亚油酸对人乳腺癌细胞生长的抑制作用

目的 研究共轭亚油酸（c9,t11-CLA)对人乳腺癌细胞（MCF－7）生长的影响。方法 采用细胞核分裂指数、细胞生长曲线、细胞集落形成试验、^3H-TdR掺入试验和软琼脂培养方法,所设剂量（μmol/L）为25,50,100,200,以96％乙醇为溶剂对照。结果 在细胞核分裂指数、细胞生长曲线和细胞集落形成试验中,可见c9,t11-CLA对MCF－7细胞生长有明显的抑制作用,其作用于MCF－7细胞8d后的抑制率（％）分别为27．18、35．43、91．05和92．86;在^3H-TdR掺入试验中,可见随着c9,t11-CLA剂量的增加,^3H-TdR掺入到MCF－7细胞中明显的减少,与阴性对照组相比差异有显性;由软琼脂培养试验结果可见,随着c9,t11-CLA剂量的增加,MCF－7细胞的集落形成逐渐降低,除了25 μmol/L剂量组外与阴性对照组相比差异有显性。结论 c9,t11-CLA对MCF－7细胞的生长有明显的抑制作用。     

The effects of conjugated linoleic acid supplementation on immune function in healthy volunteers

OBJECTIVE: To assess the effects of dietary supplementation using two isomeric blends of conjugated linoleic acid (CLA) on immune function in healthy human volunteers. DESIGN: Double-blind, randomised, placebo-controlled intervention trial. SUBJECTS AND INTERVENTION: A total of 55 healthy volunteers (n=20 males, n=35 females) were randomised into one of three study groups who received 3 g/day of a fatty acid blend containing a 50:50 cis-9, trans-11: trans-10, cis-12 CLA isomer blend (2 g CLA), and 80:20 cis-9, trans-11: trans-10, cis-12 (80:20) CLA isomer blend (1.76 g CLA) or linoleic acid (control, 2 g linoleic acid) for 8 weeks. RESULTS: Supplementation with the 80:20 CLA isomer blend significantly (P< or =0.05) enhanced PHA-induced lymphocyte proliferation. CLA decreased basal interleukin (IL)-2 secretion (P< or =0.01) and increased PHA-induced IL-2 and tumor necrosis factor alpha (TNF(alpha)) production (P< or =0.01). However, these effects were not solely attributable to CLA as similar results were observed with linoleic acid. CLA supplementation had no significant effect on peripheral blood mononuclear cells IL-4 production, or on serum-soluble intercellular adhesion molecule-1 (sICAM-1) or plasma prostaglandin E2 (PGE2) or leukotreine B4 (LTB4) concentrations. CONCLUSIONS: This study shows that CLA supplementation had a minimal effect on the markers of human immune function. Furthermore, supplementation with CLA had no immunological benefit compared with linoleic acid.     

Downregulation of inflammatory markers by conjugated linoleic acid isomers in human cultured astrocytes

Objectives: Conjugated linoleic acid (CLA) isomers have been shown to possess anti-inflammatory activity in the central nervous system. In this study, we aimed to evaluate whether modulation of the fatty acid profile by the CLA isomers c9,t11 or t10,c12CLA was associated with changes in the expression of pro-inflammatory molecules in human astrocytes.

Methods: Cultured astrocytes were treated for 6 days with 100?µM fatty acids (c9,t11CLA or t10,c12CLA or oleic acid). Following the treatment, the fatty acid profile of the cell and pro-inflammatory molecule expression were assessed.

Results: Only the t10,c12CLA isomer induced a significant decrease in arachidonic acid and increased the ratio of docosahexaenoic acid/eicosapentaenoic acid, which constitutes indirect evidence of peroxisome proliferator-activated receptor alpha activation. Inhibition of tumour necrosis factor-α, interleukin-1β, and RANTES expression was observed in astrocytes treated with c9,t11CLA and t10,c12CLA.

Discussion: Current data demonstrate that CLA isomers, particularly t10,c12, may affect neuroinflammation by reducing the pro-inflammatory molecules in cultured astrocytes, suggesting a potential nutritional role of CLA isomers in modulating the astrocyte inflammatory response.     

Enrichment of n‐3 fatty acids and conjugated linoleic acid in beef: ProSafeBeef

ProSafeBeef is a 5‐year integrated project funded by the European Commission under the sixth Framework Programme. The overall aim is to advance beef safety and quality across Europe and the work programme spans seven integrated “pillars”. Pillar 3 is concerned with producing safe beef and beef products with enhanced nutritional and eating quality characteristics. A particular focus is on the development of strategies to enhance the concentrations in beef of those fatty acids considered to be of benefit to human health, without causing a detrimental effect on the appearance, shelf‐life or eating quality of the beef. There is accumulating evidence of the importance of long‐chain n‐3 (omega‐3) polyunsaturated fatty acids (PUFAs) for human health and disease prevention, and also evidence from experimental studies that has shown anticarcinogenic, antiatherogenic, and anti‐obesity effects of two isomers of conjugated linoleic acid (CLA). Based on this evidence, the major focus of research efforts to improve the nutritional value of beef has been on increasing the concentration of the n‐3 PUFAs and CLA. Considerable progress has been made within pillar 3 of ProSafeBeef to meet this aim, primarily by manipulating the diet of cattle. Fundamental information on ruminal lipid metabolism and on the control and/or prevention of ruminal hydrogenation of dietary lipids arising from research within ProSafeBeef will facilitate the production of beef with a ‘healthier’ fatty acid profile. Moreover, strategies will be defined for industry on how to optimise nutritional and sensory properties and oxidative quality of beef products, by combining the nutritional enhancement made in the live animal together with target levels of functional ingredients to be added during processing.