羟自由基诱导的烟草原生质体的凋亡:流式细胞法的新证据

用1.0 mmol/L FeSO4/0.5 mmol/L H202处理烟草(Nicotiana tabacum L.cultivar BY 2)原生质体,发现羟自由基能够诱导烟草原生质体的凋亡.具体表现为细胞核皱缩、DNA Ladder、TUNEL阳性反应等典型的凋亡特征.在动物细胞凋亡过程中,线粒体起着非常重要的作用,其中膜电位(△ψm)的变化以及由其引起的位于线粒体膜上的通透性孔(PTP)的开放与Cyt c的释放有关.另外,在动物凋亡细胞中,磷脂酰丝氨酸(phosphatidyl serine,PS)会从细胞膜内侧向外翻转.为了判断植物细胞凋亡过程中膜电位的变化情况以及PS的外翻程度,我们采用了流式细胞法.结果表明,随着处理时间的延长,烟草原生质体线粒体的膜电位逐渐降低;膜内PS大量外翻.说明由羟自由基和烟草原生质体组成的凋亡体系是一种可靠的凋亡组合,可以用来对植物细胞凋亡机理做进一步研究.     

白头翁汤正丁醇提取物诱导白念珠菌生物被膜细胞凋亡

目的探讨白头翁汤正丁醇提取物(Butyl alcohol extract of Bai Tou Weng decoction,BAEB)对白念珠菌生物被膜细胞凋亡的影响。方法分别以DCFH-DA染色和JC-1染色,用流式细胞仪检测白念珠菌生物被膜细胞内活性氧(ROS)水平和线粒体膜电位(MMP)变化;Annexin V-FITC/PI染色染色荧光显微镜观察白念珠菌生物被膜细胞磷脂酰丝氨酸(phosphatidylserine,PS)外翻并检测凋亡率;FITC-VAD-FMK染色观察白念珠菌生物被膜细胞metacaspase活性;DAPI染色观察白念珠菌生物被膜细胞核形态。结果≥128μg/mL BAEB处理后白念珠菌生物被膜细胞内活性氧水平显著升高,线粒体膜电位显著降低,PS外翻增加,凋亡率明显升高,metacaspase酶活性显著升高,细胞核出现固缩和碎裂。结论白头翁汤正丁醇提取物可诱导白念珠菌生物被膜细胞凋亡。     

1-甲基-4-苯基-吡啶盐致多巴胺能细胞系MES23.5的死亡机制

通过测定环境毒素1-甲基-4-苯基-吡啶盐(MPP )作用于多巴胺能细胞系MES23．5后细胞存活率的变化及细胞线粒体膜电位(△ψM)、活性氧(ROS)、羟自由基、超氧化物岐化酶(SOD)的变化,发现MPP^ 作用于多巴胺能细胞系MES23．5,可导致细胞存活率显著性减少,浓度达到200mol／L以上后,细胞存活率的下降呈时间与MPP^ 浓度依赖;以200μmol/L MPP^ 作用细胞6∽48h后,△ψM逐渐下降、ROS、羟自由基逐渐增加,48h后SOD开始显著性减少。结果表明早期线粒体能量代谢障碍和膜电位变化导致ROS(尤其是羟自由基)含量增加是MPP^ 导致多巴胺能细胞氧化应激的原因,而细胞内自由基的清除机制受损,则最终导致细胞变性死亡。     

喜树碱诱导甜菜夜蛾细胞凋亡线粒体途径的MPTP依赖性

【目的】线粒体通透性转换孔(MPTP)的开放可以导致线粒体膜通透性改变,与细胞凋亡关系密切。本研究旨在探索MPTP在喜树碱诱导的昆虫细胞凋亡中的作用,以进一步揭示喜树碱(CPT)诱导昆虫细胞凋亡的机制。【方法】环孢菌素A(CsA)为MPTP开放抑制剂,通过预加入20μmol/L CsA,应用流式细胞仪测定其对CPT和羟基喜树碱(HCPT)诱导的甜菜夜蛾Spodoptera exigua细胞(IOZCAS-SPEX-Ⅱ)凋亡作用的影响,包括细胞内Ca~(2+)浓度变化,线粒体膜电位变化以及活性氧簇(ROS)变化,从而分析MPTP在CPT和HCPT诱导细胞凋亡的作用。【结果】结果显示,10μmol/LCPT和HCPT处理IOZCAS-SPEX-Ⅱ细胞6 h和12 h时,与0.1%DMSO对照组相比,甜菜夜蛾细胞发生凋亡,胞质Ca~(2+)浓度增大,线粒体膜电位降低或丧失,ROS增加,即CPT和HCPT诱导甜菜夜蛾细胞发生凋亡,为线粒体内途径。但经过20μmol/L CsA预处理2 h后再加入CPT和HCPT处理6 h,与0.1%DMSO组相比,细胞凋亡率、胞质Ca~(2+)浓度、线粒体膜电位及ROS产生均无显著差异(P0.05),即CsA抑制了MPTP的开放,从而抑制了CPT和HCPT诱导的甜菜夜蛾细胞凋亡;而加入CPT和HCPT处理12 h时,CsA对MPTP开放的抑制作用显著降低,与单CPT和HCPT处理组相比,细胞凋亡率、胞质Ca~(2+)浓度、线粒体膜电位及ROS差异不显著,即CPT和HCPT诱导的细胞凋亡如常发生。【结论】本研究证实喜树碱和羟基喜树碱诱导甜菜夜蛾细胞凋亡线粒体途径具有MPTP开放依赖性,且首次明确这种依赖性具有时间性。     

PUMA对高糖诱导的大鼠H9C2心肌细胞凋亡的作用及机制

目的:观察促凋亡蛋白p53上调凋亡调控因子(PUMA)在高糖所致H9C2心肌细胞凋亡中的作用及机制。方法:H9C2心肌细胞随机分为对照组(使用5.5mmol/L葡萄糖作用于细胞)和高糖组(使用35 mmol/L葡萄糖作用于细胞,HG组)分别刺激6 h, 12 h, 24 h和48 h,每组设复孔5个,TUNEL染色检测细胞凋亡率;RT-PCR及Western blot法分别测定PUMA mRNA及蛋白表达情况;JC-1法检测线粒体膜电位;Western blot测定caspase-3表达和细胞色素c(Cyt C)释放。H9C2细胞随机分为四组,对照组、高糖(35 mmol/L)、HG+si-scramble组(使用si-scramble转染心肌细胞24 h,使用35mmol/L葡萄糖作用于细胞)和Si-PUMA组(使用si-PUMA转染心肌细胞24 h,使用35mmol/L葡萄糖作用于细胞),观察抑制PUMA表达对高糖诱导细胞凋亡率、线粒体膜电位、Cyt C的影响。结果:与对照组相比,高糖刺激心肌细胞组TUNEL染色阳性率、活化caspase-3和PUMA表达明显升高(P<0.0...     

二烯丙基三硫通过线粒体依赖性途径诱导人肝癌HepG2细胞凋亡

二烯丙基三硫(diallyl trisulfide,DATS)对多种肿瘤有抗癌作用,但机制尚不完全清楚．为探讨DATS对人肝癌细胞系HepG2细胞凋亡的影响,用丫啶橙/溴化乙锭(AO/EB)法观察细胞凋亡情况,在显微镜下计数凋亡细胞数．JC-1荧光染色观察线粒体膜电势变化．Western blot法检测细胞色素c蛋白分布,ELISA法检测caspase-3活性． 结果显示,DATS诱导HepG2细胞凋亡,用 50 μmol/L 与100 μmol/L DATS处理48 h,细胞凋亡率分别达到60.33%和93.67%,并引起HepG2细胞线粒体膜电势降低．Western blot显示,DATS能诱导胞浆细胞色素c增加,与此同时,线粒体细胞色素c减少,诱导HepG2细胞caspase-3活化．提示：DATS可通过降低线粒体膜电势,促进细胞色素c由线粒体膜释放到胞浆中,激活caspase-3途径诱导人肝癌细胞系HepG2细胞凋亡．     

甘蔗和烟草叶原生质体分离期间的膜损伤及有关酶活性变化

甘蔗和烟草叶原生质体分离期间的膜损伤及有关酶活性变化何若天,覃伟,李任强（广西农业大学实验中心,南宁５３０００５）关键词：原生质体,超氧阴离子自由基（Ｏ_２~－）,膜损伤,甘蔗,烟草植物原生质体分离期间,所用细胞壁降解酶和高渗介质等对细胞生理有深刻影响...     

全反式维甲酸拮抗地塞米松抑制人气道上皮细胞16HBE凋亡

该研究探讨全反式维甲酸(all-trans Retinoic Acid, ATRA)对GCs诱导人气道上皮细胞过度凋亡的拮抗作用。将人支气管上皮细胞16HBE作为研究对象,使用10μmol/L地塞米松(Dex)、1μmol/L ATRA处理细胞,体外培养16HBE细胞,分为Dex组、ATRA组、Dex+ATRA组和Control组。采用TUNEL法检测16HBE细胞凋亡情况, Annexin V/PI双染法检测膜磷脂酰丝氨酸外翻的情况,荧光探针JC-1检测线粒体膜电位变化, Western blot检测细胞的凋亡蛋白Caspase-3和其水解片段Cleaved-caspase-3的水平。相较于Control组, Dex组的TUNEL阳性细胞明显增多, Annexin V阳性蛋白明显上调,线粒体膜电位明显下降, Caspase-3和Cleaved-caspase-3的水平明显增高;联合ATRA则可以显著减轻Dex引起的这个趋势。因此, Dex可以导致人支气管上皮16HBE细胞的过度凋亡,而ATRA可以削弱Dex的这种作用。     

低温诱导淡水白鲳尾鳍细胞系早期凋亡

淡水白鲳属喜温性鱼类,本文以其尾鳍细胞系CBT为研究材料,探讨低温胁迫(10℃)对细胞的影响。实验采用MTT法测定细胞活力,透射电子显微镜观察超微结构,激光扫描共聚焦显微镜检测磷脂酰丝氨酸(PS)对翻和线粒体膜电位的变化。结果显示:低温处理导致细胞活力和线粒体膜电位显著下降(P<0.01);电镜下可见核质过聚等现象;10℃处理3d,Annexin-V-FITC/PI双染表明87.7%的细胞进入早期凋亡。由此认为,冷胁迫淡水白鲳CBT细胞致死的途径是细胞凋亡。     

鸟苷酸二钠体外抗氧化能力的研究

本文研究了鸟苷酸二钠（GMPNa2）对羟自由基和DPPH&#183;自由基的清除效果,同时建立了H2O2氧化损伤体外培养脾细胞模型,用MTT法检测GMPN％的修复及增殖作用,并分析了GMPNa2对细胞抗氧化体系及抗氧化能力的影响,以评价其抗氧化活性。结果表明：GMPNa2具有剂量依赖性的体外抗氧化和清除活性氧能力,10mmol／LGMPNa2的羟自由基清除能力高达96．644％,但DPPH清除率为6．589％。GMPNa2清除羟自由基与同浓度Vc相近,而其清除DPPH自由基的能力很弱。添加0．5、1、5和10mmol／LGMPNa2均能显著修复H2O2诱导的脾细胞氧化损伤（P〈0．05）,提高总抗氧化能力和抗氧化酶类活力（P〈0．01）。GMPNa2添加量大于1mmol／L时,可显著降低MDA含量（P〈0．01）。说明GMPNa2具有显著的抗氧化作用。     

Recent advances in the study of Kaposi's sarcoma-associated herpesvirus replication and pathogenesis

It has now been over twenty years since a novel herpesviral genome was identified in Kaposi's sarcoma biopsies. Since then, the cumulative research effort by molecular biologists, virologists, clinicians, and epidemiologists alike has led to the extensive characterization of this tumor virus, Kaposi's sarcoma-associated herpesvirus(KSHV; also known as human herpesvirus 8(HHV-8)), and its associated diseases. Here we review the current knowledge of KSHV biology and pathogenesis, with a particular emphasis on new and exciting advances in the field of epigenetics. We also discuss the development and practicality of various cell culture and animal model systems to study KSHV replication and pathogenesis.     

The structure, reproduction and taxonomy of Vidalia and Osmundaria (Rhodophyta, Rhodomelaceae)

Comprises species occurring mostly in subtidal habitats in tropical, subtropical and warm-temperate areas of the world. An analysis of the type species, V. spiralis (Sonder) Lamouroux ex J. Agardh, a species from Australia, establishes basic characters for distinguishing species in the genus. These characters are (1) branching patterns of thalli, (2) flat blades that may be spiralled on their axis, (3) width of the blade, (4) primary or secondary derivation of sterile and fertile branchlets and (5) position of sterile and fertile branchlets on the thalli. Application of the latter two characters provides an important basic method for separation of species into three major groups. Osmundaria , a genus known only in southern Australia, was studied in relation to Vidalia , and its separation from the Vidalia assemblage is not accepted. Species of Vidalia therefore are transferred to the older genus name, Osmundaria. Two new species, Osmundaria papenfussii and Osmundaria oliveae are described from Natal. Confusion in the usage of the epithet, Vidalia fimbriala Brown ex Turner has been clarified, and Vidalia gregaria Falkenberg, described as an epiphyte on Osmundaria pro/ifera Lamouroux, is revealed to be young branches of the host, Osmundaria prolifera.     

New or rare chromosome counts in Artemisia L. (Asteraceae, Anthemideae) and related genera from Kazakhstan

Fifteen chromosome counts of six Artemisia taxa and one species of each of the genera Brachanthemum, Hippolytia, Kaschgaria, Lepidolopsis and Turaniphytum are reported from Kazakhstan. Three of them are new reports, two are not consistent with previous counts and the remainder are confirmations of very scarce (one to four) earlier records. All the populations studied have the same basic chromosome number, x = 9, with ploidy levels ranging from 2x to 6x. Some correlations between ploidy level, morphological characters and distribution are noted.     

肝癌中HBV和HCV基因和抗原的分布及意义

采用原位分子杂交方法检测HCV RNA及HBV X基因;采用免疫组织化学方法研究HCV核心抗原,非结构区C33c抗原及HBxAg在肝细胞肝癌中的定位及分布.结果表明(1)HCV RNA、HBV X基因在肝细胞肝癌组织检出率分别为40%(55/136)和82%(112/136).HCV RNA定位于癌细胞的胞浆内,阳性细胞呈散在、灶状及弥漫分布三种形式;HBV X基因在肝癌细胞中的分布呈胞浆型、核型及核浆型,阳性细胞也呈上述三种分布形式;(2)HCV C33c抗原、核心抗原在肝细胞肝癌中的阳性率为81%(133/164)及86%(141/164).C33c抗原定位于癌细胞及肝细胞的胞浆内;核心抗原既定位于癌细胞核中,又可定位于胞浆中.C33c抗原阳性细胞以灶状分布为主;而核心抗原阳性细     

Selection with two alleles and complete dominance

For a plant selection model with frequency-independent viabilities, fertilities and selfing rates, it is shown that apart from global fixation, for certain parameter combinations a protected polymorphism and facultative fixation (either allele may become fixed according to initial frequencies) may both occur. Facultative fixation requires different selling rates for the dominant and recessive type. Protection of the polymorphism requires resource allocation for male and female function. In this connection the problem of purely genetically caused population extinction is discussed.
For general frequency dependence and regular segregation, the chances for establishment of a completely recessive gene are compared to those of a completely dominant gene. It is proven that the process of establishment of the recessive gene, despite a fitness advantage, may be considerably endangered by drift effects if random mating prevails. The recessive gene may reach the same effectivity in establishment as a dominant gene, only if the recessive homozygote mates exclusively with its own type during the period of establishment.