EGCG通过PTEN增强ATRA对早幼粒白血病细胞株的分化作用

探讨PTEN在表没食子儿茶素没食子酸酯(EGCG)增强维甲酸(ATRA)诱导的急性早幼粒细胞白血病(APL)细胞分化中的作用。我们分别将ATRA、EGCG及两种药物联合应用于NB4与HL-60细胞72 h,从形态学上观察细胞核的变化;Western blotting检测PTEN及髓系分化标志CD11b的表达;免疫荧光检测PTEN的入核情况;CCK-8检测细胞增殖率。结果显示,ATRA与EGCG联合作用于NB4与HL-60细胞后,PTEN与CD11b的表达较单独应用ATRA时增加。PTEN特异性抑制剂SF1670作用于NB4后,细胞分化明显减少;PI3K抑制剂LY294002作用后,PTEN及CD11b表达增加。这些均表明EGCG能够增强ATRA诱导APL细胞株的分化作用,且这种促进作用与PTEN的增加是相关的。     

NLS-RARα对人白血病细胞NB4分化抑制及其机制的研究

该文旨在探讨带核定位信号的维甲酸受体α(nuclear localization signal retinoic acid receptor alpha,NLS-RARα)对人急性早幼粒白血病(acute promyelocytic leukemia,APL)细胞株NB4分化的影响及其机制。免疫印迹实验检测全反式维甲酸(all-trans retinoic acid,ATRA)诱导的NB4细胞分化标志物C/EBPβ、CD11b和p38α蛋白质水平;利用慢病毒介导的NLS-RARα基因过表达,进一步用免疫印迹实验验证过表达效率并检测NLS-RARα对NB4细胞分化标志物C/EBPβ、CD11b和p38α蛋白质水平的影响;间接免疫荧光实验分析NLS-RARα与p38α的空间共定位;免疫共沉淀实验分析NLS-RARα与p38α的相互作用。结果显示,生理浓度和药理浓度的ATRA促进NB4细胞分化的同时也激活了p38α,且p38α的活性变化与髓系分化标志物C/EBPβ变化一致;髓系分化表面标志物CD11b表达量在药理浓度ATRA(1μmol/L)处理下达到最高;NLS-RARα抑制NB4细胞的分化,且只有在ATRA存在的条件下,NLS-RARα抑制NB4细胞的分化与下调p38α活性相关;NLSRARα与p38α存在空间共定位且NLS-RARα与p38α直接相互作用。该研究结果提示,当存在ATRA诱导时,NLS-RARα与p38α直接相互作用后下调p38α的活性进而抑制NB4细胞的分化。     

miR-382-5p通过靶基因PTEN阻滞NB4细胞分化

该文主要研究在急性早幼粒细胞白血病NB4细胞中, miR-382-5p通过调控靶基因PTEN(phosphatase and tensin homologue)抑制了全反式维甲酸(all-trans retinoic acid, ATRA)诱导的急性早幼粒细胞分化。我们运用ATRA(1μmol/L)诱导细胞分化; Western blot检测PTEN及髓系分化标志物CD11b的蛋白质水平;实时荧光定量PCR检测miR-382-5p的表达水平;过表达PTEN的慢病毒载体分别感染NB4细胞和HL-60、THP-1细胞;脂质体转染miR-382-5p的模拟剂(mimics)和特异性抑制剂(inhibitors)NB4细胞。结果显示, PTEN促进ATRA诱导的NB4细胞分化,而在HL-60和THP-1细胞中并无明显促分化效应。NB4细胞中,脂质体转染miR-382-5p mimics在mRNA和蛋白水平均抑制了PTEN的表达,并且抑制了ATRA诱导的分化;转染miR-382-5p inhibitors则恢复了PTEN表达,同时促进了ATRA诱导的急性早幼粒细胞NB4细胞的分化。该文结果提示, miR-382-5p靶向抑制了PTEN的表达从而抑制ATRA诱导的NB4细胞分化。     

盐霉素抑制急性早幼粒细胞白血病细胞增殖并诱导分化

该文旨在探讨盐霉素(salinomycin,SAL)对急性早幼粒细胞白血病NB4细胞增殖和分化的影响及其可能的机制。采用CCK-8(cell counting kit-8)实验检测细胞增殖,瑞氏染色观察细胞形态学变化,流式细胞术检测粒细胞分化标志物CD11b的表达,Western blot检测相关蛋白质水平变化。结果显示,SAL抑制了细胞增殖;SAL作用72 h后,细胞呈现典型分化形态学改变。随着SAL的浓度升高,CD11b阳性的细胞比例以及CD11b、C/EBPβ蛋白质水平逐渐增加。此外,SAL降低了β-catenin以及下游分子C-myc、Cyclin D1的蛋白质水平。该研究还探讨了联合使用Wnt/β-catenin信号通路的抑制剂IWR-1与SAL对细胞分化的影响。结果显示,与单独使用SAL相比,联合使用SAL和IWR-1促进了SAL诱导的NB4细胞分化。该研究结果提示,SAL可抑制NB4细胞的增殖,并可能通过抑制Wnt/β-catenin信号通路诱导细胞分化。     

MicroRNA-10b通过调控锌指蛋白KLF4阻滞白血病细胞分化

探讨microRNA-10b(miR-10b)通过调节锌指蛋白Krüppel-like factor 4(KLF4)的表达对急性白血病细胞分化的影响。Real-time PCR及Western blot分别检测不同分化程度的白血病细胞系中miR-10b与KLF4的表达;1,25-二羟基维生素D3(1,25D3)诱导人白血病细胞系HL60向单核系分化,检测此过程中miR-10b及KLF4的表达变化;利用体外合成的寡核苷酸(miR-10b mimics)转染HL60细胞,瑞氏–吉姆萨染色观察1,25D3诱导后细胞分化形态学的改变;流式细胞术检测单核细胞表面标志CD14的表达。结果显示,miR-10b在分化早期的KG-1a细胞中表达最高,在分化晚期的U937、THP-1细胞中表达最低(P<0.01),而KLF4的表达与之相反;1,25D3诱导HL60向单核系分化过程中,miR-10b表达呈时间依赖性降低,KLF4表达则逐渐增高;HL60细胞中过表达miR-10b后可抑制1,25D3诱导的细胞分化形态特征的改变及CD14的表达(P<0.05)。提示miR-10b通过负调控KLF4的表达阻滞白血病细胞HL60单核系的分化。     

CD11b/DNA双参数流式细胞术检测HL-60细胞分化的细胞周期

目的采用双参数流式细胞术研究全反式维甲酸(alltransretinoidacid,ATRA)诱导人类急性早幼粒白血病细胞HL-60细胞分化的细胞周期。方法HL-60细胞经分化诱导剂ATRA(终浓度为1μmol/L)诱导不同时间点后,利用CD11b/DNA双参数流式细胞术同时检测分化细胞表面抗原CD11b的表达及分化细胞DNA含量。结果HL-60细胞经ATRA诱导后,细胞表面分化抗原CD11b表达明显升高,细胞阻滞于G0/G1期,且CD11b阳性细胞主要位于G0/G1期。结论CD11b/DNA双参数流式细胞术能简便,快速,直观地检测细胞分化的细胞周期。     

转谷氨酰胺酶2通过mTOR通路和自噬调控全反式维甲酸诱导的白血病细胞HL60和U937的髓系分化

全反式维甲酸(ATRA)是具有融合基因PML-RARα的急性早幼粒细胞白血病(APL)特异的靶向治疗药物。此外，ATRA在无PML-RARα融合的急性髓系白血病及其它一些肿瘤中也有一定治疗效果。但ATRA治疗也会引起一些并发症或发生愈后复发。因此，对ATRA诱导分化调控机制的研究非常重要。转谷氨酰胺酶2(TGM2)是一种多功能酶，能调控mTOR信号通路和自噬等。ATRA能诱导APL细胞中TGM2表达上调，TGM2敲低抑制ATRA诱导的细胞分化。但其调控机制及涉及的信号通路尚不明确。本研究发现，在HL60和U937细胞中，ATRA能够上调CD11b和TGM2的表达(P<0.05),抑制mTOR信号通路，并增强自噬；与对照相比，敲低TGM2,mTOR信号通路增强，自噬被抑制，而ATRA诱导的CD11b表达被抑制(P<0.05),分化减弱，被ATRA抑制的mTOR信号通路得到部分恢复，而被ATRA增强的自噬适当减弱。这表明ATRA使HL60和U937细胞发生髓系分化，并诱导TGM2表达升高；而TGM2通过mTOR信号通路和自噬途径调控ATRA诱导的髓系分化。该研究将有利于更深入地...     

MiR-15b促进急性早幼粒细胞白血病细胞分化并抑制增殖

该文探讨了miR-15b在全反式维甲酸(all-trans retinoic acid, ATRA)治疗急性早幼粒细胞白血病(acute promyelocytic leukemia, APL)过程中发挥的作用。采用实时荧光定量PCR检测miR-15b的表达;流式细胞术检测细胞的分化情况; CCK-8实验检测细胞增殖;双荧光素酶报告实验检测miR-15b与CCNE1 3′UTR端的结合能力; Western blot检测下游靶基因CCNE1的表达。结果显示,ATRA促进miR-15b的表达;过表达miR-15b增强了ATRA对APL细胞的分化作用,而抑制miR-15b表达后则出现相反结果; miR-15b抑制了APL细胞的增殖能力;双荧光素酶报告实验显示miR-15b与CCNE1的3′UTR端结合; Western blot显示mi R-15b可以抑制下游靶基因CCNE1的表达。这些结果表明, miR-15b通过抑制CCNE1的表达促进APL细胞分化,抑制细胞增殖。     

维替泊芬通过抑制YAP诱导白血病NB4细胞凋亡

探讨维替泊芬对人类白血病NB4细胞活性、凋亡的影响及其作用机制。我们用CCK-8实验检测NB4细胞的增殖活性;集落形成实验检测NB4细胞的集落形成能力;细胞周期和凋亡用流式细胞术来检测;凋亡形态学用Hoechst33342染色来观察;以及Western blotting检测细胞中蛋白的表达水平。结果显示,NB4细胞经维替泊芬处理后,集落数量减少,集落大小减小;CCK-8结果提示细胞增殖活性受到显著抑制,且呈现出剂量和时间依赖性(p0.05);Hoechst 33342染色后观察到细胞凋亡形态;流式检测到G0/G1期细胞明显增多,细胞凋亡率明显增加(p0.05);Western blotting检测到YAP、GSK3β、p-AKT、cyclin D1、Bcl-2蛋白表达水平明显下调,Bax、p-GSK3β表达明显增加,裂解的PARP蛋白明显增加(p0.05)。因此本研究提示,维替泊芬能抑制人类白血病NB4细胞的增殖,诱导细胞凋亡。其机制是通过抑制YAP蛋白的表达诱导细胞凋亡,AKT/GSK3β信号参与了这一过程。     

柚皮素通过Notch1/Hes1通路抑制肺癌干细胞增殖、迁移和分化

为探讨柚皮素对肺癌干细胞增殖、迁移和分化的分子机制,本研究应用免疫磁珠法分选肺癌干细胞(A549-CSCs),并通过流式细胞术进行表面分子的鉴定;通过CCK8法检测不同浓度的柚皮素(25μg/m L,50μg/mL, 100μg/mL)对肺癌干细胞(A549-CSCs)活力的影响,Transwell检测柚皮素对A549-CSCs细胞迁移能力的影响,Q-PCR检测柚皮素对肺癌干细胞分化相关因子Sox2和Oct4 m RNA表达的影响,Western blotting法检测柚皮素对细胞内Notch1和Hes1蛋白表达的影响。流式细胞术检测结果显示,A549-CSCs细胞表面分子CD133呈阳性表达,符合肺癌干细胞特征。CCK8结果显示,与对照组(control)比较,25μg/m L、50μg/mL、100μg/mL柚皮素处理A549-CSCs 24 h,细胞活力显著降低(p<0.05);Transwell检测结果显示,与对照组比较,不同浓度柚皮素处理组A549-CSCs迁移能力显著降低(p<0.05);定量PCR (real-time polymerase chain reaction, Q-PCR)结果显示,与对照组比较,柚皮素处理组细胞Sox2和Oct4 m RNA表达水平显著降低(p<0.05);蛋白质印迹法(Western blotting)结果显示,与对照组相比柚皮素处理组细胞Notch1和Hes1蛋白表达水平均降低。本研究发现柚皮素可能通过抑制Notch1/Hes1通路抑制肺癌干细胞增殖、迁移和分化。这为柚皮素治疗肺癌提供临床依据。     

六种藓类植物茎的比较解剖学研究

通过对6种藓类植物,即褶叶青藓(Brachythecium salebrosum(Web.et Mohr.)B.S.G.)、湿地匐灯藓(Plagiomnium acutum(Lindb.)Kop.)、侧枝匐灯藓(Plagiomnium maximoviczii(Lindb.)Kop.)、大凤尾藓(Fissidensnobilis Griff.)、大羽藓(Thuidium cymbifolium(Doz.et Molk.)B.S.G.)和大灰藓(Hypnum plumaeforme Wils.)嫩茎和老茎的石蜡切片和显微观察发现,同一藓类植株的嫩茎和老茎,茎结构稳定,不同种藓类植物茎横切面具有不同特征.植物体茎横切面形状、表层细胞的层数、细胞大小和细胞壁厚薄、皮层细胞大小和形状、中轴的有无以及比例等特征可以作为藓类植物的分科分类依据之一.     

Photoregulation of seed germination of wild-type and of an aurea-mutant of tomato

Seed germination of an aurea mutant of tomato ( Lycopersicon esculentum Mill.) is promoted by continuous irradiation with red, far-red or long-wavelength far-red (758 nm) light as well as by cyclic irradiations (5 min red or 5 min far-red/25 min darkness). Far-red light applied immediately after each red does not change the germination behaviour. Seed germination of the isogenic wild-type, cv. UC-105, is promoted by continuous and cyclic red light while it is inhibited by continuous and cyclic far-red light and by continious 758 nm irradiation. Far-red irradiation reverses almost completely the promoting effect of red light. The promoting effect (in the aurea mutant) and the inhibitory effect (in the wild-type) of continuous far-red light do not show photon fluence rate dependency above 20 nmol m⁻² s⁻¹. It is concluded that phytochrome controls tomato seed germination throgh low energy responses in both the wild type and the au mutant. The promoting effect of continuous and cyclic far-red light in the au mutant can be attributed to a greater sensitivity to P_fr.     

真菌类遗传学分析的知识结构教学

本文以认知结构理论为指导,讨论了真菌类遗传分析与高等动植物遗传分析的内在联系,认为利用这种内在联系进行教学可收到好的效果并说明了作者的具体教学过程。 Abstract:In the paper, the relationship between genetic analysis of Fungi and genetic analysis of high animal and plant was discussed.A good results were obtained when we adopted this method in the teaching.     

Cause of Senescence of Nine Sorts of Flowers

The levels of endogenous phytohormones and respiratory rate in nine sorts of flowers such as Cymbidium faberi Rolfe, Nopalxochia ackermannii Kunth and others were investigated both at full bloom and senescence and meanwhile the effect of exogenous phytohormones on prolonging the blossoms and promoting ethylene production were tested. There is a high content of endogenous ethylene in all the long-lived flowere, about 3–16 folds higer than the short-lived ones. There is a high level of ABA at full blooming flowers of short-lived flowers, in which there is no or only some cytokinins in it, but the ratio of CTK (6BA+zeatin)/ABA is smaller(l.7). The endogenous ABA reached a much higher level at senescence in all nine sorts of flowers, so it is reasonable to consider that it is ABA which plays an important role of regulation in controlling flower's senescence. There is a much higher level of GA3 and zeatin in the long-lived flowers which is not demonstrated in the shortlived ones. The respiratory rate is one of the factors controtling the longevity of flowers, but it does not play a decided role. Application of 6BA and zeatin prolongs distinctly orchid’s longevity, however exogenous IAA through the promotive action on ethylene production, evidently extends the longevity of the flowers of the Nopalxochia ackermannii Kunth.     

Interconnection of parameters of regulation system of lipid peroxidation and of morphophysiological parameters of mouse liver

A complex analysis of seasonal fluctuations of the mean group parameters of the system of regulation of lipid peroxidation has been performed in liver of Balb/c mice. Association of lipid characteristics and morphophysiological parameters is studied in the Balb/c mouse liver. An inter-connection is revealed between the liver index and the amount of lysoforms of phospholipids, the scale and character of the interconnection differing essentially depending on proportion of phos-phatidylcholine in mouse liver phospholipids.     

龙胆科药用植物化学成分的研究现状

龙胆科植物在我国的分布范围很广,且多数为药用植物,其多数种属的药用植物,至今其化学成分尚未被系统研究。综述了目前龙胆科药用植物的化学成分的研究现状及一般提取方法,对近年来发现的环烯醚萜及裂环烯醚萜类化合物进行了总结,为本科药用植物的更深入研究提供了参考。     

Physiological characteristics of various species of strains of carboxydobacteria

Seven strains of aerobic carbon monoxide-oxidizing bacteria (carboxydebacteria) when growing on CO as sole source of carbon and energy had doubling times which ranged from 12–42 h. The activity profiles obtained after discontinuous sucrose density gradient centrifugation indicated that the CO-oxidizing enzymes are soluble and the hydrogenases are membrane-bound in all strains examined. The CO-oxidizing enzymes of Pseudomonas carboxydohydrogena, Pseudomonas carboxydoflava, Comamonas compransoris, and the so far unidentified strains OM2, OM3, and OM4 had a molecular weight of 230,000; that of Achromobacter carboxydus amounted to 170,000. The molecular weights of the CO-oxidizing and H₂-oxidizing enzymes turned out to be identical. The cell sonicates were shown to catalyze the oxidation of both CO and H₂ with methylene blue, thionine, phenazine methosulfate, toluylene blue, dichlorophenolindophenol, cytochrome c or ferricyanide as electron acceptors. Methyl viologen, benzyl viologen, FAD⁺, FMN⁺, and NAD(P)⁺ were not reduced. The spectrum of electron acceptors was identical for all strains tested. Neither free formate, hydrogen nor oxygen gas were involved in the CO-oxidation reaction. Methylene blue was reduced by CO at a 1:1 molar ratio. The results indicate that CO-oxidation by carboxydobacteria is catalyzed by identical or similar enzymes and that the reaction obeys the equation CO+H₂OCO₂+2H⁺+2e^- as previously shown for Pseudomonas carboxydovorans.Dedicated to Otto Kandler remembering almost three decades of enjoyable cooperation     

Modulation of allostery of pyruvate kinase by shifting of an ensemble of microstates

Since the introduction of the concepts of allostery about four decades ago, much advancement has been made in elucidating the structure-function correlation in allostery. However, there are still a number of issues that remain unresolved. In this review we used mammalian pyruvate kinase (PK) as a model system to understand the role of protein dynamics in modulating cooperativity. PK has a triosephosphate isomerase (TIM)(α/β)₈ barrel structural motif. PK is an ideal system to address basic questions regarding regulatory mechanisms about this common (α/β)₈ structural motif. The simplest model accounting for all of the solution thermodynamic and kinetic data on ligand-enzyme interactions involves two conformational states, inactive E^T and active E^R. These conformational states are represented by domain movements. Further studies provide the first evidence for a differential effect of ligand binding on the dynamics of the structural elements, not major secondary structural changes. These data are consistent with our model that allosteric regulation of PK is the consequence of perturbation of the distribution of an ensemble of states in which the inactive E^T and active E^R represent the two extreme end states. Sequence differences and ligands can modulate the distribution of states leading to alterations of functions. The future work includes: defining the network of functionally connected residues; elucidating the chemical principles governing the sequence differences which affect functions; and probing the nature of mutations on the stability of the secondary structural elements, which in turn modulate allostery.