海地瓜硫酸软骨素通过Wnt信号通路抑制3T3-L1前脂肪细胞分化的研究

目的 探究海地瓜硫酸软骨素(Acaudina Molpadioides chondroitin sulfate, AM-CHS)抑制3T3-L1前脂肪细胞分化的分子机制。方法 采用传统的鸡尾酒诱导剂诱导分化3T3-L1前脂肪细胞,分别采用油红O染色和甘油三酯(Triglycerides, TG)含量测定法评价AM-CHS对3T3-L1前脂肪细胞分化的影响。采用RT-PCR法检测脂肪细胞中Wnt信号通路关键基因和脂肪合成关键基因mRNA的表达水平。结果 AM-CHS能够显著抑制3T3-L1细胞脂滴的形成,拮抗罗格列酮(Rosiglitazone, RSG)的促分化作用,并上调Wnt信号通路中的关键受体Frz和LRP5的mRNA表达,下调分化转录因子PPARγ、C/EBPα和SREBP-1c mRNA的表达,抑制脂质合成基因FAS和GPAT mRNA的表达,不影响脂质分解基因HSL和ATGL mRNA的表达。结论 AM-CHS能够显著抑制3T3-L1前脂肪细胞的分化,抑制成熟脂肪细胞的脂质合成,并拮抗罗格列酮(Rosiglitazone, RSG)的促脂质合成作用,其作用机制与激活Wnt信号通路有关。     

催产素对3T3-L1脂肪细胞糖脂代谢的影响

目的 观察催产素对3T3-L1脂肪细胞糖脂代谢的影响。方法 3T3-L1前脂肪细胞体外培养,并诱导其分化成熟为脂肪细胞。研究催产素对脂肪细胞葡萄糖消耗量以及三酰甘油、游离脂肪酸和甘油的影响。采用实时荧光定量PCR法检测糖脂代谢相关基因GLUT-1、GLUT-4、ATGT、HSL的m RNA表达。结果 与对照组比较,催产素20、50、100μg/m L组葡萄糖消耗量有所增加,且表现出剂量相关。催产素组较对照组的三酰甘油降低,而甘油和游离脂肪酸增高。催产素50μg/m L组中脂代谢相关基因HSL表达明显高于对照组,糖代谢相关基因GLUT-4 m RNA表达水平增加。结论 催产素处理可减少3T3-L1细胞脂质合成、增加脂质分解作用,并可明显改善脂质积聚。     

ATGL/HSL角度下解析Neu-p11改善胰岛素抵抗作用机制

目的探讨脂肪组织甘油三酯酶(adipose triglyceridelipase,ATGL)及激素敏感性脂肪酶(hormone-sensitive lipase,HSL)在褪黑素非选择性受体激动剂Neu-p11改善高糖高胰岛素(high glucose and insulin,HGI)诱导的3T3-L1脂肪细胞胰岛素抵抗(insulin resistance,IR)中的作用及机制。方法培养3T3-L1脂肪细胞,HGI诱导IR模型。以葡萄糖消耗量及细胞内甘油三酯(triglyceride,TG)定量测定作为检测指标,Western blot检测蛋白水平的表达情况。结果 HGI孵育减少脂肪细胞葡萄糖摄取,促进细胞内TG积聚,同时伴有ATGL及HSL的蛋白表达下调。Neu-p11干预逆转了HGI对脂肪细胞的作用效应,而MT2竞争性拮抗剂luzindole却拮抗了Neu-p11的上述效应。结论 Neu-p11以MT2受体依赖性方式抑制IR脂肪细胞TG沉积,可能与其上调AT-GL、HSL蛋白的表达,促进TG水解相关。     

蕨菜超临界萃取物化学成分及其抑制3T3-L1前脂肪细胞分化作用研究

目的分析蕨菜超临界CO_2萃取物(SFE-PA)的化学成分,并探究萃取物对3T3-L1前脂肪细胞分化的抑制作用及机制。方法采用GC-MS分析SFE-PA主要化学成分。体外培养3T3-L1前脂肪细胞,构建脂肪细胞分化的细胞模型,MTT法测定细胞毒性,采用油红O染色法分析SFE-PA对脂肪细胞分化的抑制作用,利用qRT-PCR检测脂肪细胞分化中相关基因的mRNA表达水平。结果从SFE-PA中共鉴定出10种化合物,主要成分有4,4,5,7,8-五甲基-二氢香豆素(36.07%)、β-谷甾醇(25.66%)、4-氨基-7-二乙氨基-香豆素(8.26%)、棕榈酸(5.05%)等。SFE-PA浓度依赖性地抑制3T3-L1前脂肪细胞分化,并减少细胞中脂质的沉积。同时,SFE-PA明显下调PPARγ、CEBPα、SREBP-1c、FAS、ACC的mRNA表达水平。结论 SFE-PA具有明显抑制3T3-L1前脂肪细胞分化的作用,这一作用与其调控脂肪细胞分化转录调控因子和脂质合成相关基因等密切相关,而高含量的酚类、醇类化合物可能是其发挥作用的物质基础。     

荭草素抑制3T3-L1前脂肪细胞分化及其改善胰岛素抵抗的作用研究

目的研究荭草素对3T3-L1前脂肪细胞分化及对脂肪细胞胰岛素抵抗的影响,并探讨其作用机制。方法传统鸡尾酒法诱导分化3T3-L1前脂肪细胞,MTT法检测荭草素对前脂肪细胞活力的影响,油红O染色法检测脂质积累,酶法检测细胞内甘油三酯(triglyceride,TG)的含量。地塞米松诱导成熟的脂肪细胞,建立胰岛素抵抗模型,荧光标记2-脱氧葡萄糖(2-[N-(7-nitrobenz-2-oxa-1,3-diaxol-4-yl)amino]-2-deoxyglucose,2-NBDG)摄入法观察脂肪细胞对葡萄糖的摄取能力;Western blot检测腺苷酸活化蛋白激酶(adenosine monophosphate activated protein kinase,AMPK)、乙酰辅酶A羧化酶(acetyl Co A carboxylase,ACC)的磷酸化水平及葡萄糖转运体4(glucose transporter type 4,GLUT4)的表达水平,免疫荧光法检测GLUT4的向膜转位能力。结果荭草素可浓度依赖性地减少细胞脂滴的积累及细胞内TG的含量,但对细胞活力无明显的影响(P>0.05)。胰岛素抵抗状态下,荭草素明显增加脂肪细胞对2-NBDG的摄取(P<0.05),明显上调AMPK、ACC的磷酸化水平(P<0.05),促进GLUT4的向膜转位及表达。结论荭草素抑制前脂肪细胞的分化,同时,荭草素通过上调AMPK/GLUT4信号途径相关蛋白的表达,促进了细胞对葡萄糖的摄取,达到改善胰岛素抵抗的作用。     

胰高血糖素样肽-1对胰岛素抵抗3T3-L1脂肪细胞脂肪酸代谢的作用及机制

目的研究胰高血糖素样肽-1(GLP 1)对胰岛素抵抗3T3 L1脂肪细胞脂肪酸代谢的作用及机制。方法采用高糖高胰岛素造成胰岛素抵抗3T3 L1脂肪细胞模型,通过ELISA及Western blot等方法观察GLP 1对此模型脂肪酸代谢的影响及机制。结果 ELISA结果显示,GLP 1对胰岛素抵抗3T3 L1脂肪细胞中游离脂肪酸(FFA)的含量影响与胰岛素相关:在有胰岛素(100 nmol.L-1)存在时,GLP 1可增加上清液中FFA含量;而无胰岛素存在时,GLP 1可减少上清液中FFA含量。GLP 1升高细胞中脂肪酸合成酶(FAS)表达量的作用也必须依赖胰岛素的存在。Western blot结果显示在有胰岛素存在时,GLP 1可促进蛋白激酶B(PKB)磷酸化;而无胰岛素存在时则无此作用。PKB磷酸化的抑制剂LY294002或Wortmannin可阻断胰岛素存在时GLP 1对PKB磷酸化的促进作用及对上清液FFA含量的升高作用。另外,在有(无)胰岛素存在时,GLP 1均可降低激素敏感性脂肪酶(HSL)的蛋白表达量。结论 GLP 1可增强胰岛素抵抗3T3 L1脂肪细胞对胰岛素的敏感性并降低HSL的含量;胰岛素可影响GLP 1对胰岛素抵抗3T3 L1脂肪细胞脂肪酸代谢的调节作用。     

大豆黄素衍生物LRXH609的减肥作用及机理探讨

目的：探讨大豆黄素衍生物IRXH609（LRX）的减肥作用和可能的机理。方法：以高脂饮食诱导肥胖大鼠,测量体重、Lee’s指数、腹腔脂肪重量、摄食量、血脂和血糖,检验LRX灌服30d后的减肥效果;体外培养诱导分化3T3-L1前脂肪细胞,观察药物对脂肪细胞增殖、脂质合成和分解的影响。结果：LRX显著降低高脂饮食诱导的肥胖大鼠体重、Lee’s指数、腹腔脂肪重量;降低血液中TC和游离脂肪酸（FFA）含量;抑制3T3-L1前脂肪细胞增殖;显著提高3T3-L1前脂肪细胞内激素敏感脂酶（HSL）活性,促进脂肪分解释放甘油（Gly）,减少细胞内TG含量。结论：LRXH609有显著的减肥和调血脂作用,可能的机理是通过抑制前脂肪细胞增殖和分化,激活HSL促进脂肪细胞内TG分解,降低脂肪细胞内TG存储量。     

3T3-L1前脂肪细胞分化过程中miRNAs基因的表达变化

目的:探讨3T3-L1前脂肪细胞诱导分化过程中miRNAs基因的表达变化。方法:运用经典的"激素鸡尾酒"法建立3T3-L1前脂肪细胞分化模型。采用实时定量聚合酶链式反应方法检测不同时相点(0d、2h、2d、4d)细胞分化相关标志物的表达和20种miRNAs的表达变化,采用生物信息学方法对变化显著的miRNAs进行靶基因预测。结果:成功建立3T3-L1前脂肪细胞分化模型。不同时相点细胞分化相关标志物基因水平提示3T3-L1前脂肪细胞分化为脂肪细胞。20种miRNAs中以miR-126、miR-214、miR-320、miR-351的表达最为显著。靶基因预测结果显示,miR-126靶基因有影响细胞增殖的转录因子Crk、代谢相关基因Irs1等;miR-214靶基因有转录因子Zbtb20、Kpna1等;miR-320靶基因有影响代谢相关基因Igf2bp3、Gxylt1等;miR-351靶基因有Myt1、Mcl1、Bmf等。结论:miR-126、miR-214、miR-320、miR-351可通过影响基因转录及细胞代谢水平,从而对3T3-L1前脂肪细胞分化发挥作用,为开发治疗肥胖等疾病的新药提供了新的研究思路。     

海地瓜硫酸软骨素对3T3-L1前脂肪细胞增殖和分化的影响

目的研究海地瓜硫酸软骨素(Acaudina Molpadioideschondroitin sulfate,AM-CHS)对3T3-L1前脂肪细胞增殖和分化的影响,并探讨其作用机制。方法采用传统的鸡尾酒诱导剂诱导分化3T3-L1前脂肪细胞,以MTT法检测AM-CHS对3T3-L1前脂肪细胞及不同分化阶段3T3-L1细胞增殖活性的影响;分别采用油红O染色和甘油三酯(triglycerides,TG)含量测定法评价其对3T3-L1前脂肪细胞分化的影响。采用RT-PCR法检测脂肪细胞中过氧化物酶体增殖体激活受体γ(peroxisome proliferators-activated receptors gamma,PPARγ)、CCAAT增强子结合蛋白α(CCAAT/enhancer bind-ing protein alpha,C/EBPα)、固醇调节元件结合蛋白-1c(ste-rol regulatory element binding protein-1c,SREBP-1c)等分化相关基因的mRNA表达水平。结果 AM-CHS能明显抑制3T3-L1前脂肪细胞和成熟脂肪细胞的增殖,抑制3T3-L1前脂肪细胞的分化过程,以对分化早期的抑制作用最强。RT-PCR结果表明,AM-CHS能明显降低脂肪细胞PPARγ、C/EBPα和SREBP-1c mRNA的表达。结论海地瓜AM-CHS能明显抑制3T3-L1前脂肪细胞的增殖和分化,其作用机制与下调分化相关基因PPARγ、C/EBPα和SREBP-1c的表达有关。     

NYGGF4基因表达沉默对3T3-L1脂肪细胞线粒体动力学的影响

目的 探讨NYGGF4基因表达沉默对3T3-L1成熟脂肪细胞线粒体动力学的影响.方法 以3T3-L1前体脂肪细胞为工具,分别构建NYGGF4基因表达沉默细胞株和空载对照细胞株,通过地塞米松(DEX)、1-甲基-3-异丁基黄嘌呤(MIX)、胰岛素联合方案将前体脂肪细胞诱导分化为成熟脂肪细胞.采用实时荧光定量PCR检测成熟脂肪细胞中线粒体融合基因(Mfn) 1/2、线粒体动态相关基因(Drp)1、线粒体分裂基因(Fis)1的mRNA表达水平和线粒体DNA(mtDNA)拷贝数.结果 NYGGF4基因表达沉默成熟脂肪细胞的Mfn1/2、Drp1表达水平与对照组相仿.NYGGF4基因表达沉默成熟脂肪细胞的Fisl的表达水平显著高于对照组(P＜0.05).结论 在3T3-L1脂肪细胞中,NYGGF4基因表达沉默可显著上调Fisl基因的表达水平,提示NYGGF4基因表达沉默可能通过影响线粒体分裂从而影响成熟脂肪细胞线粒体动力学.     

Synthese von γ,γ'-Dihydroxysulfonen und γ-Hydroxy-γ'-ketosulfonen

Syntheses of γ,γ'-Dihydroxysulfones and γ-Hydroxy-γ'-ketosulfones Reduction of γ,γ'-diketosulfones 1 with dimethylaminoborane leads to γ,γ'-dihydroxysulfones 3 via γ-hydroxy-γ'-ketosulfones 2 . The influence of substituents on the ratio of the yields of 2 and 3 is investigated.     

Analysis of γβ, βγ, γγ, ββ multiple turns in proteins

Abstract: The number of γ‐turns in a representative protein dataset selected from the current Protein Data Bank has increased almost seven times during the past decade. Eighty percent classic γ‐turns and 57% inverse γ‐turns are associated as multiple turns with either another γ‐turn or a β‐turn. We refer to these as multiple turns of the (γβ)_1,2,3 or (βγ)_1,2,3 type, depending upon whether the γ‐turn is before or after the β‐turn along the protein chain, respectively. However, for multiple turns involving only γ‐turns, we follow the nomenclature analogous to that proposed earlier for the multiple (or double) β‐turns. Fifty‐eight per cent β‐turns are associated as multiple turns with another β‐turn. We extracted multiple turns from the protein dataset and classified them on the basis of individual γ‐ or β‐turn types and the number of overlapping residues. Furthermore, we evaluated the amino acid positional potentials and determined the statistically significant amino acid preferences, hydrogen bond/side‐chain interaction preferences in the multiple turns and secondary structure preferences for residues immediately flanking these turns. The results of our analysis would be useful in the modeling, prediction or design of multiple turns in proteins. The amino acid sequence corresponding to the multiple turn, position in the protein chain, PDB Code/chain in which multiple turn is present and the individual turn types constituting the multiple turns are available from our website and this information would also be integrated in our Database of Structural Motifs in Proteins ( http://www.cdfd.org.in/dsmp.html ).     

Analysis of γβ, βγ, γγ, ββ continuous turns in proteins

Abstract: We report the observation of continuous turns in proteins which comprise individual γ‐turns or β‐turns or both that are situated immediately one after the other along the polypeptide chain. The continuous turns were identified from a representative data set of three‐dimensional protein crystal structures. The γβ/βγ, γγ and ββ continuous turns represent peptides of varying amino acid residue lengths and conformations. The continuous turns frequently observed in proteins were: γβ, between a coil and a strand; βγ, between a helix and a strand; γγ, between coils; and ββ, either between a strand and a coil or between strands or coils. We determined the statistically significant amino acid residue preferences at individual positions in the turn, calculated amino acid positional potentials and analyzed main chain hydrogen bonds and side‐chain interactions likely to stabilize the continuous turns. The data on continuous turns have been integrated in the database of structural motifs in proteins (DSMP) on our web server at ( http://www.cdfd.org.in/dsmp.html ). This is useful to make queries on sequences compatible with different continuous turns.     

3-O-demethyl-2,3-di-epi-fortimicins and 3-O-demethyl-3-epi-fortimicins

Syntheses of the 3-O-demethyl-2,3-di-epi-fortimicins A and B and the 3-O-demethyl-3-epi-fortimicins A and B have been accomplished in processes the key steps of which were solvolyses of 4-N-acetyl-3-O-demethyl-3-O-methanesulfonylfortimicin derivatives. Antibacterial activities of the new antibiotics are reported.     

3',3'-Difluoro-3'-deoxythymidine: comparison of anti-HIV activity to 3'-fluoro-3'-deoxythymidine.

3',3'-Difluoro-3'-deoxythymidine (3) has been synthesized in four steps from thymidine, and characterized by 1H NMR and NOE experiments. The JHF coupling constants support a conformation in solution that is predominantly 2'-endo (S). Although conformationally and sterically nucleoside 3 may resemble other thymidine analogs which are active against HIV-1, 3 is virtually inactive.     

Lactone, 10. Mitt. Zur Synthese N-heterocyclisch substituierter γ,γ-Diphenyl-γ-butyrolactone

Lactones, X: Synthesis of γ,γ-Diphenyl-γ-butyrolactones Substituted by N-Heterocycles Starting from 3 and 16 , syntheses of γ,γ-diphenyl-γ-butyrolactones with N-heterocycles at the α-position are described. Scope and limitation of the syntheses, tautomerism and stereochemistry of the products as well as spectroscopic results which are inconsistent with data from the literature are discussed.