水杉愈伤组织诱导及植株再生

通过愈伤组织诱导器官发生途径,建立了水杉（Metasequoia glyptostroboides）的植株再生体系,探讨了不同外植体（种胚、幼叶切块、茎段、根段）和植物生长调节剂对不定芽直接再生和愈伤组织诱导器官发生的影响。结果表明：以种胚、无菌苗叶片、茎段和根作为外植体,在MS补加2,4-D、NAA和6-BA不同组合的培养基上都能诱导得到愈伤组织,其中种胚诱导愈伤组织效果最好,诱导率可达100%,茎诱导效果次之,诱导率为97.1%。诱导愈伤组织效果较好的培养基有：MS＋1.0mg·L-12,4-D＋0.5mg·L-16-BA、MS＋0.1mg·L-16-BA＋1.0mg·L-1NAA、MS＋0.5mg·L-16-BA＋1.0mg·L-1NAA、MS＋1.0mg·L-16-BA＋1.0mg·L-1NAA、MS＋0.5mg·L-16-BA＋2.0mg·L-1NAA、MS＋1.0mg·L-16-BA＋2.0mg·L-1NAA和MS＋0.5mg·L-12,4-D＋0.5mg·L-1NAA。以愈伤组织在MS培养基上植株再生效果最好,再生率为62.5%。     

单核细胞增多性李斯特菌氨基肽酶Lmo1711体外表达及其酶活分析

对单核细胞增多性李斯特菌(简称单增李斯特菌)lmo1711基因编码的氨基肽酶进行克隆表达与纯化,并研究该重组蛋白的体外酶学特性。首先通过生物信息学分析预测Lmo1711与氨基肽酶家族成员的亲缘关系及关键活性位点的保守性。利用SWISS-MODEL模拟预测该蛋白的空间结构;构建Lmo1711原核表达载体并转化入E.coli Rosetta中,诱导表达重组目的蛋白,并利用镍离子亲和层析方法纯化目的蛋白;以氨基酸-对硝基苯胺偶联物为底物,Lmo1711通过水解底物N端氨基酸残基产生游离对硝基苯胺单体,405 nm处检测吸光值对该产物进行检测从而分析Lmo1711的酶学特性。在此基础上系统研究Lmo1711对不同氨基酸残基底物的催化特异性,及不同金属离子对该酶活性的影响。经原核表达纯化获得49.3 kDa的重组Lmo1711蛋白,与预测分子量一致;生物信息学分析推测Lmo1711属于M29氨基肽酶家族,且存在保守关键氨基酸活性位点(Glu250、Glu316、His345、Tyr352、His378、Asp380);酶活分析显示,Lmo1711具有较强的氨基肽酶活性,针对不同底物的结合和催化能力差异较大,对亮氨酸残基的亲和程度最高;Lmo1711氨基肽酶活性具有金属离子依赖性,Co~(2+)、Cd~(2+)、Zn~(2+)等多种金属离子均能显著增强其活性,其中Co~(2+)的激活效应最显著。本试验首次发现并证实,单增李斯特菌Lmo1711属于M29氨基肽酶家族成员,具有较强的催化活性,且对金属离子具有不同程度的依赖性。     

Long non‐coding RNA highly up‐regulated in liver cancer promotes epithelial‐to‐mesenchymal transition process in oral squamous cell carcinoma

Oral squamous cell carcinoma (OSCC) is an oral and maxillofacial malignancy that exhibits high incidence worldwide. In diverse human cancers, the long non‐coding RNA (lncRNA) highly up‐regulated in liver cancer (HULC) is aberrantly expressed, but how HULC affects OSCC development and progression has remained mostly unknown. We report that HULC was abnormally up‐regulated in oral cancer tissues and OSCC cell lines, and that suppression of HULC expression in OSCC cells not only inhibited the proliferation, drug tolerance, migration and invasion of the cancer cells, but also increased their apoptosis rate. Notably, in a mouse xenograft model, HULC depletion reduced tumorigenicity and inhibited the epithelial‐to‐mesenchymal transition process. Collectively, our findings reveal a crucial role of the lncRNA HULC in regulating oral cancer carcinogenesis and tumour progression, and thus suggest that HULC could serve as a novel therapeutic target for OSCC.     

通过和玉米杂交诱导硬粒小麦单倍体

硬粒小麦（Ｔｒｉｔｉｃｕｍ ｄｕｒｕｍ Ｄｅｓｆ．）ＤＲ１４７授以超甜玉米（Ｚｅａ ｍ ａｙｓ Ｌ．） ｓｓ ７７００的花粉后,在８３．４％ 的柱头上观察到花粉萌发及花粉管长入胚囊,有９．９％ 的子房发生了卵细胞的单受精,１．９％ 的子房发生了中央细胞的单受精,３２．７％ 的子房发生了双受精。尽管双受精后可同时形成胚和胚乳,但胚乳往往发育迟缓,甚至败育。硬粒小麦×玉米形成的杂合子核型高度不稳定,在最初的几次细胞分裂中,来自父本玉米的染色体逐步被排除,最后形成硬粒小麦单倍体胚。在授以玉米花粉４ ｈ 后用１００ ｐｐｍ ２,４－Ｄ溶液浸蘸硬粒小麦穗部,可以有效地促进幼胚在缺乏胚乳或胚乳败育情况下的生长和发育。授粉９—１３ ｄ 后由５３３个硬粒小麦子房解剖出２５个胚,得胚率为４．７％ 。通过幼胚拯救获得１１棵正常植株,植株获得率为２．１％ 。根尖细胞染色体计数表明,它们为单倍体（２ｎ＝ ２ｘ＝ １４）。     

Synaptic neurotransmission depression in ventral tegmental dopamine neurons and cannabinoid-associated addictive learning

Drug addiction is an association of compulsive drug use with long-term associative learning/memory. Multiple forms of learning/memory are primarily subserved by activity- or experience-dependent synaptic long-term potentiation (LTP) and long-term depression (LTD). Recent studies suggest LTP expression in locally activated glutamate synapses onto dopamine neurons (local Glu-DA synapses) of the midbrain ventral tegmental area (VTA) following a single or chronic exposure to many drugs of abuse, whereas a single exposure to cannabinoid did not significantly affect synaptic plasticity at these synapses. It is unknown whether chronic exposure of cannabis (marijuana or cannabinoids), the most commonly used illicit drug worldwide, induce LTP or LTD at these synapses. More importantly, whether such alterations in VTA synaptic plasticity causatively contribute to drug addictive behavior has not previously been addressed. Here we show in rats that chronic cannabinoid exposure activates VTA cannabinoid CB1 receptors to induce transient neurotransmission depression at VTA local Glu-DA synapses through activation of NMDA receptors and subsequent endocytosis of AMPA receptor GluR2 subunits. A GluR2-derived peptide blocks cannabinoid-induced VTA synaptic depression and conditioned place preference, i.e., learning to associate drug exposure with environmental cues. These data not only provide the first evidence, to our knowledge, that NMDA receptor-dependent synaptic depression at VTA dopamine circuitry requires GluR2 endocytosis, but also suggest an essential contribution of such synaptic depression to cannabinoid-associated addictive learning, in addition to pointing to novel pharmacological strategies for the treatment of cannabis addiction.     

血卟啉单甲醚在A549肺癌细胞内亚细胞分布的动态变化

目的:探讨血卟啉单甲醚(Hematoporphyrin monomethyl ether,HMME)在A549肺癌细胞内亚细胞分布的动态变化。方法:传代培养A549肺癌细胞,分别与光敏剂HMME孵育2 h和12 h。应用由荧光显微镜及电感耦合器材(Charge-coupled device,CCD)组成的高分辨率荧光显微成像系统,结合荧光探针标记技术,采用细胞器-细胞荧光强度比值法研究HMME在不同时间的亚细胞分布情况。结果:在2 h和12 h两个孵育时间点高尔基体的平均荧光强度比值(J1/J2)值都最高;随着孵育时间延长,A549细胞的四种细胞器J1/J2值都升高且溶酶体幅度最大。结论:孵育时间是影响HMME亚细胞分布的一个重要因素。随着孵育时间的延长,A549肺癌细胞各细胞器吸收HMME能力逐渐增强,尤以溶酶体显著。     

95例SARS患者细胞免疫功能抑制的特点及其原因

用ELISA方法测定95例SARS患者急性期、恢复期血清细胞因子水平,用流式细胞仪检测该组患者急性期、恢复期淋巴细胞亚群,并分析细胞因子水平和淋巴细胞亚群变化的关系。结果发现,IL-10和TGF—β在观察期（急性期和恢复期）持续升高,急性期CD4^＋和CD8^＋T细胞显著减少,恢复期患者外周血CD8^＋记忆T细胞减少36．78％。IL-10和TGF-β升高与淋巴细胞变化具有统计学相关。结果提示,SARS病毒感染抑制宿主的细胞免疫功能,引起急性期CD4^＋和CD8^＋T细胞显著降低和恢复期的免疫记忆细胞减少。而IL-10和TGF—β过表达可能在SARS免疫病理中起重要作用。     

亮氨酸拉链提高小鼠血浆中剪接的凝血因子Ⅷ凝血活性

培养细胞实验表明,亮氨酸拉链通过改善内含肽(intein)的蛋白质剪接效率,提高双载体转B区缺失型凝血因子Ⅷ(BDD-FⅧ)基因细胞剪接FⅧ蛋白的分泌量和活性.本文从C57BL/6小鼠门静脉注射含亮氨酸拉链和Ssp DnaB内含肽融合的BDD-FⅧ的重链和轻链基因双表达载体,48 h后,检测到血浆的重链分泌量和FⅧ活性分别为(298±67)μg/L和(1.15±0.29)U/mL,明显高于不含亮氨酸拉链的双载体转BDD-FⅧ基因对照小鼠((179±59)μg/L和(0.58±0.19)U/mL).结果表明,亮氨酸拉链通过改善蛋白质反式剪接,提高基于蛋白质剪接的双载体转BDD-FⅧ基因小鼠血浆的凝血活性,为进一步双腺相关病毒(AAV)载体转BDD-FⅧ基因的甲型血友病基因治疗研究提供了依据.