姜黄素通过促进ROS产生诱导人脑胶质瘤细胞凋亡

本文用姜黄素处理人脑胶质瘤U87细胞,以CCK-8法检测姜黄素对细胞增殖的影响,在光学显微镜下观察姜黄素处理后的细胞形态学变化;酶联免疫法测定NADPH氧化酶的含量;用DCFH-DA荧光探针检测细胞ROS含量,并对氧化应激指标总氧化力(T-AOC)、丙二醛(MDA)、超氧化物歧化酶(SOD)和谷胱甘肽(GSH)进行了检测。用Annexinv/PI双染后流式细胞术检测和AO/EB荧光染色检测细胞凋亡,并通过免疫印迹法检测了凋亡相关蛋白信号通路。结果表明,姜黄素通过提高细胞NADPH氧化酶活性促进ROS产生,引起细胞内T-AOC降低,MDA含量升高,GSH含量下降,SOD活性升高,使细胞处于氧化胁迫,并可能通过ROS的升高触发细胞信号通路,下调NF-κB/p65蛋白的表达,最终通过凋亡执行分子Caspase-3促使细胞凋亡。     

氧化还原与细胞凋亡的关联

细胞内氧化还原状态与细胞凋亡相互关联的机理仍然存在很大争议。细胞内氧化还原状态的改变促进了氧自由基(ROS)的产生和凋亡诱导因子的激活,致使细胞凋亡的同时又加剧了细胞内氧化还原状态的改变。通过激活细胞凋亡信号激酶(ASK-1)、氧化还原转录因子NF-κB、AP-1及Caspase激活,揭示了细胞内氧化还原状态伴随细胞凋亡的不同阶段。     

活性氧对NF-κB活性及JNK信号通路的调节

活性氧（ROS）是生物体有氧代谢过程中产生的一类活性含氧化合物的总称,机体细胞可通过多种途径维持ROS产生与降解的动态平衡。研究表明,活性氧可作为第二信使调节与细胞增殖、分化、凋亡相关的信号转导通路。c-JunN端激酶（JNK）通路可以介导氧化应激、细胞因子、紫外照射等引起的细胞凋亡。另外,κ基因结合核因子（NF-κB）是氧化应激调节的靶因子之一,同样也能诱导促进细胞内的氧化应激反应,还可通过活性氧蓄积抑制JNK的激活。简要综述活性氧对NF-κB和JNK信号通路的调节。     

齐墩果酸抑制肿瘤坏死因子-α诱导的成纤维细胞样滑膜细胞炎症因子表达及其机制研究

探讨齐墩果酸(Oleanolic acid,OA)对肿瘤坏死因子-α(TNF-α)诱导成纤维细胞样滑膜细胞的炎症因子表达的影响及其机制。首先复苏培养人成纤维细胞样滑膜细胞(FLS),通过RT-PCR检测细胞IL-6及IL-1βmRNA表达,采用Western blot方法检测p38MAPK及NF-κB蛋白表达变化,通过ELISA法检测细胞上清液中IL-6及IL-1β浓度。与对照组比较,TNF-α明显诱导FLS细胞IL-6及IL-1βmRNA的表达及上清液中IL-6及IL-1β的分泌(P0.05),同时磷酸化p38蛋白和核NF-κB明显增加(P0.05),且p38MAPK阻断剂SB203580能抑制TNF-α诱导的核NF-κB增加。OA呈浓度依赖性抑制TNF-α诱导的FLS细胞p38蛋白磷酸化和核NF-κB增加(P0.05)。且OA、p38MAPK通路抑制剂SB203580或NF-κB阻断剂BAY 11-7082均能抑制TNF-α诱导的IL-6及IL-1β分泌增加(P0.05)。综上所述,OA能抑制TNF-α诱导的FLS细胞炎症因子IL-6及IL-1β的产生,其机制可能与抑制p38MAPK/NF-κB信号通路有关。     

细胞水平筛选鉴定激活核因子κB通路的新基因TMEM9B

核因子κB(NF-κB)是细胞内重要的转录因子,其介导的细胞信号转导通路在细胞凋亡中的作用是国内外研究的热点．为了筛选NF-κB通路相关新基因,建立了基于细胞水平的报告基因高通量筛选模型．利用双荧光素酶报告系统检测报告基因荧光素酶活性,通过对构建的439个人类未知功能基因的筛选,获得了一批激活NF-κB信号通路的功能基因,其中基因TMEM9B可以明显激活NF-κB通路．进一步实验显示TMEM9B激活NF-κB通路呈明显剂量依赖性,Western blot及EMSA实验证实,TMEM9B能够促进胞质内NF-κB的抑制分子IκBα的降解,并促使NF-κB由胞质向胞核转移,同时流式细胞术实验发现TMEM9B可引起293T和HeLa细胞的凋亡．总之,所建立的基于细胞水平的NF-κB通路筛选模型稳定高效,筛选并验证TMEM9B可明显激活NF-κB信号转导通路,并从而引起细胞凋亡．     

延龄草苷对过氧化氢诱导的PC12细胞氧化损伤和炎症因子表达的影响

探讨延龄草苷对过氧化氢(H_2O_2)诱导的PC12细胞氧化损伤和炎症因子表达的影响。采用MTT法和LDH活性测定观察延龄草苷对PC12细胞模型的影响,采用相关试剂盒检测活性氧(ROS)和丙二醛(MDA)含量,超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GSH-Px)活力,Western blot检测Sirt1、NF-κB和TNF-α的蛋白表达。延龄草苷(5~20μM)能够显著提高H_2O_2损伤的PC12细胞的活力,提高细胞抗氧化能力,并上调Sirt1的表达,下调NF-κB和TNF-α的表达,其保护作用可能与提高抗氧化能力,降低炎症因子损伤,调控Sirt1/NF-κB信号通路有关。     

氧化应激对腺病毒E1A蛋白活化NF-κB转录活性的影响及其机制研究

目的:探讨腺病毒E1A蛋白对细胞内抗氧化物质谷胱甘肽(GSH)水平的影响及氧化应激对腺病毒E1A蛋白介导的核因子-κB(NF-κB)转录活化的影响。方法:构建稳定表达E1A蛋白的大鼠肺泡上皮细胞(E1A组)及对照质粒转染细胞(对照组),每组5×105个细胞,试验重复3次。采用H2O2刺激细胞,检测细胞内GSH水平。脂多糖(LPS)和肿瘤坏死因子-α(TNF-α)进行刺激,丁胱亚磺酰亚胺(BSO)进行干预,Western blot法检测NF-κB和AP-1蛋白的表达。结果:E1A+细胞内GSH基础水平与E1A-比较无显著差异,但在H2O2作用后没有诱导出GSH含量的上升,表现为下降而低平的趋势,去除氧化剂后,仍低于正常水平。E1A-细胞在氧化剂的作用后呈明显的上升趋势,去除氧化剂后仍高于基础水平。细胞内NF-κB蛋白表达(积分吸光度值):刺激前E1A+细胞分别为79.3±4.6和80.3±3.8,在LPS和TNF-α刺激后分别为81.8±3.9～89.9±1.6和94.1±1.9～99.8±1.6,均明显高于对照组(刺激前分别为68.3±3.8和69.4±4.3,刺激后分别为70.1±2.8～80.8±3.6和73.4±4.9～83.2±6.7)。给予BSO预处理后再用LPS和TNF-α刺激,E1A-细胞NF-κB蛋白表达的积分吸光度值(1.22±0.16和1.75±0.13)与LPS或TNF-α单独作用组(1.25±0.18和1.69±0.19)无明显差别;E1A+细胞NF-κB蛋白表达的积分吸光度值(1.75±0.10和2.26±0.21)明显高于LPS或TNF-α单独作用组(1.35±0.12和1.80±0.14)。结论:腺病毒潜伏感染持续表达E1A蛋白可以影响细胞GSH,降低细胞对氧化应激的耐受性,并可能通过此机制造成NF-κB异常转录活化,而GSH的降低又进一步放大了E1A介导的NF-κB转录活化作用。     

TNF-α诱导的NF-κB慢病毒报告基因系统的构建

目的:构建一种肿瘤坏死因子α(TNF-α)诱导的NF-κB慢病毒报告基因系统。方法:以NF-κB信号通路为基础,设计并构建含有NF-κB响应元件和mCherry报告基因序列的表达载体,并进一步建立受NF-κB调控表达mCherry的细胞株,最后使用TNF-α进行刺激验证。结果:质粒经过双酶切鉴定,得到约1500 bp的目的片段,符合预计大小且测序分析正确,表明质粒构建成功;在TNF-α刺激实验中,NF-κB信号通路的刺激物TNF-α作用于构建的NF-κB调控表达mCherry荧光蛋白的细胞株后出现特异性荧光反应,TNF-α诱导组的细胞有较强的mCherry表达,阳性率约为90%,而未加TNF-α组细胞mCherry阳性表达率约为10%。结论:构建了受NF-κB调控稳定表达mCherry的慢病毒报告基因系统,可用于对NF-κB活化效果的检测及筛选,具有普适性的应用价值。     

Apelin 通过调节ROS 水平抑制TNF-α 诱导的HepG2 细胞凋亡

目的:探讨apelin在肿瘤坏死因子(tumor necrosis factor-α,TNF-α)诱导的肝细胞凋亡中的作用及可能机制。方法:PCR检测HepG2细胞和原代小鼠肝细胞中APJ受体的表达;采用Hoechst 33342染色检测TNF-α诱导的HepG2细胞凋亡;用活性氧(ROS)检测试剂盒结合流式细胞术测定细胞内ROS水平;通过Western blot检测信号分子JNK的磷酸化水平;比较给予apelin处理对上述指标的影响。结果:HepG2细胞和原代小鼠肝细胞均表达APJ受体;apelin可抑制TNF-α导致的细胞内ROS生成增多和JNK磷酸化水平升高并减少TNF-α诱导的HepG2细胞凋亡。结论:Apelin可能通过拮抗TNF-α诱导的细胞内ROS水平升高,使JNK信号失活,从而抑制HepG2细胞凋亡。     

A Unique Protease Cleavage Site Predicted in the Spike Protein of the Novel Pneumonia Coronavirus (2019-nCoV) Potentially Related to Viral Transmissibility

正Dear Editor,In December 2019, a novel human coronavirus caused an epidemic of severe pneumonia(Coronavirus Disease 2019,COVID-19) in Wuhan, Hubei, China(Wu et al. 2020; Zhu et al. 2020). So far, this virus has spread to all areas of China and even to other countries. The epidemic has caused 67,102 confirmed infections with 1526 fatal cases     

Curcuminoids as inhibitors of thioredoxin reductase: A receptor based pharmacophore study with distance mapping of the active site

Curcumin is the yellow pigment of turmeric that interacts irreversibly forming an adduct with thioredoxin reductase (TrxR), an enzyme responsible for redox control of cell and defence against oxidative stress. Docking at both the active sites of TrxR was performed to compare the potency of three naturally occurring curcuminoids, namely curcumin, demethoxy curcumin and bis-demethoxy curcumin. Results show that active sites of TrxR occur at the junction of E and F chains. Volume and area of both cavities is predicted. It has been concluded by distance mapping of the most active conformations that Se atom of catalytic residue SeCYS498, is at a distance of 3.56 from C13 of demethoxy curcumin at the E chain active site, whereas C13 carbon atom forms adduct with Se atom of SeCys 498. We report that at least one methoxy group in curcuminoids is necessary for interation with catalytic residues of thioredoxin. Pharmacophore of both active sites of the TrxR receptor for curcumin and demethoxy curcumin molecules has been drawn and proposed for design and synthesis of most probable potent antiproliferative synthetic drugs.     

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.     

Comparative morphology of the carpel in the Liliaceae: Hewardieae, Petrosavieae, and Tricyrteae

The young pistils in the melanthioid tribes, Hewardieae, Petrosavieae and Tricyrteae, are uniformly tricarpellate and syncarpous. They lack raphide idioblasts. All are multiovulate, with bitegmic ovules. The Petrosavieae are marked by the presence of septal glands and incomplete syncarpy. Tepals and stamens adhere to the ovary in the Hewardieae and the Petrosavieae but not in the Tricyrteae. Two vascular bundles occur in the stamens of the Hewartlieae and Tricyrtis latifolia. Ventral bundles in the upper part of the ovary of the Hewardieae are continuous with compound septal bundles and placental bundles in the lower part. Putative ventral bundles occur in the alternate position in the Tricyrteae and putative placental bundles in the opposite. position in the Petrosavieae. The dichtomously branched stigma in each carpel of the Tricyrteae is supplied by a bifurcated dorsal bundle.     

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