灵芝两个无孢品种中水提物成分分析及免疫活性比较

龙芝2号和鹿角灵芝均为赤芝的栽培品种,且在生长发育过程中均不产生孢子。目前,尚缺乏对两者化学成分和药理活性的系统比较研究。本研究以灵芝两个无孢品种——龙芝2号和鹿角灵芝为原料,研究对比两者子实体水提物成分差异及免疫活性的强弱。采用化学及仪器分析相结合的方法,分析两者水提物中的多糖得率、含量、重均分子量分布特征及核苷、蛋白质、氨基酸含量差异,并研究了灵芝两个无孢品种水提物样品刺激RAW 264.7细胞释放NO活性。结果表明,两者多糖得率及含量差异不大,但龙芝2号中重均分子量分布范围较广,多糖分子量较大,其含有3种多糖,分子量分别为2.021×10⁶、1.802×10⁶和4.825×10⁵,而鹿角灵芝中仅含有1种多糖,分子量为1.589×10⁴;两者中含有的核苷种类相似,但各核苷的含量存在差异;鹿角灵芝和龙芝2号中蛋白质含量分别为10.70%和10.32%,两者均不含有组氨酸,蛋氨酸在两者中含量均较高,分别达到2.556%和2.591%。从鹿角灵芝和龙芝2号中得到的水提物样品均具有体外刺激巨噬细胞释放NO的活性。     

Antigenicity and immunogenicity of SARS-CoV S protein receptor-binding domain stably expressed in CHO cells

The receptor-binding domain (RBD) of SARS coronavirus (SARS-CoV) spike (S) protein contains multiple conformation-dependent epitopes that induce neutralizing antibody responses. Here we used CHO-K1 cells to establish a cell line for stable expression of a 193-mer (residues 318-510) RBD (RBD193-CHO) and determined its antigenicity and immunogenicity. We found that RBD193-CHO reacted strongly with a panel of six monoclonal antibodies recognizing various conformational and linear epitopes in RBD, suggesting that this recombinant protein maintains intact conformation and good antigenicity. Immunization of mice with RBD193-CHO resulted in induction of high titers of RBD-specific neutralizing antibodies and potent IL-4-expressing T cell responses. RBD193-CHO induced immunity that protected a majority of the vaccinated mice from SARS-CoV challenge. These results suggest that the recombinant RBD produced in an established stable cell line maintains strong immunogenicity with high potential for use as an effective and economic subunit SARS vaccine.     

神经生长因子与神经系统老化

老化通常指生物体生长发育成熟以后,随年龄增加生理机能逐渐减退,内环境稳定性下降,组织器官逐渐发生退行性改变,最终走向衰老、死亡的过程。神经系统老化是神经元退行性病变形成的基础和条件。由于神经生长因子（nerve growth factor,NGF）与中枢神经系统胆碱能神经元的存活和可塑性调节密切相关,所以NGF在神经系统老化和神经退行性变疾病如老年性痴呆（Alzheimer’s disease,AD）的发生发展过程中发挥重要作用。本文综述了NGF在脑老化中的变化及其与AD发病机制的关系。     

细菌胞外多糖的生物合成与基因控制

细菌胞外多糖是指细菌在生长发育过程中合成并分泌到细胞外的长链,高分子糖类聚合物。细菌胞外多糖的生物合成途径涉及装配、多聚化及运输三个过程,是多种酶和转运系统的结果,其发生的部位包括胞内和胞外,有些合成过程会发生在细胞壁上,对于胞外多糖合成相关基因的报道,发现控制胞外多糖合成是一大类基因簇,不同的菌株其基因簇的数量和种类各不相同。这些研究的不断更新为将来胞外多糖的应用提供了更加广阔的前景。     

干旱条件下小麦冠层温度及其性状的关联研究

通过对小麦冠层温度和有关性状的长期观测,发现自然界存在冠层温度持续偏低的小麦,在干旱条件下,它的叶片功能期、叶绿素含量、蒸腾速率、净光合速率、可溶性蛋白含量和超氧化物歧化酶活性等重要性状明显优于冠层温度持续偏高的小麦品种,且丙二醛(MDA)积累速度慢,在籽粒灌浆期表现尤为明显。冷型小麦表现出干旱胁迫下仍然具有代谢功能较好、活力旺盛和抗早衰能力较强的特征,这进一步拓展了冷型小麦的应用范围,对于加速适应于干旱条件的优良品种选育并将其推向生产具有十分重要的意义。     

筛选鉴定一株产生抑菌活性物质的海洋放线菌

目的：分离筛选能够产生抑菌活性物质的海洋放线茵,并进行生理生化和16SrDNA鉴定。方法：用分离培养基培养海洋放线菌,并筛选出能够产生抑菌活性物质的菌株,对所筛选菌株的形态特征、生理生化特性进行鉴定分析;采用通用引物27F、1492R扩增该菌株的16SrDNA,对测序结果进行分析;采用Neighbor—Joining（N—J）法构建系统发育进化树。结果：筛选到一株对金黄色葡萄球菌、大肠杆菌、白色念珠菌具有较强抗性的海洋放线菌F1,该菌株好氧,中度嗜盐,在高氏I号培养基上呈白色绒粉状,16SrDNA序列比对表明该菌株与田无链霉菌（Streptomyces tanashiensis）NR043369的相似度为99％。结论：筛选到的菌株F1是一株海洋来源的放线菌,与田无链霉菌NR043369的同源性较高,可能属海洋链霉菌属,对金黄色葡萄球菌等病原菌具有较强的抑菌活性。     

突触泡蛋白2研究进展

突触泡蛋白2(SV2)是一类跨膜糖蛋白,定位于脊椎动物神经元及内分泌细胞,与神经递质的释放、内分泌泡胞吐作用、突触泡稳态的维持、神经肌肉接头的形成及肾上腺素能受体α2C的定位密切相关。最近还发现SV2是肉毒神经毒素BoNT/A的受体,介导BoNT/A进入神经元。SV2可作为突触泡标记蛋白,广泛应用于生物学研究及肿瘤诊断。此外,SV2还是抗癫痫药物的作用靶标。     

Modified heparin inhibits P-selectin-mediated cell adhesion of human colon carcinoma cells to immobilized platelets under dynamic flow conditions

Accumulating evidence indicates that the formation of tumor cell platelet emboli complexes in the blood stream is a very important step during metastases and that the anti-metastasis effects of heparin are partially due to a blockade of P-selectin on platelets. In this study, heparin and chemically modified heparins were tested as inhibitors of three human colon carcinoma cell lines (COLO320, LS174T, and CW-2) binding to P-selectin, adhering to CHO cells expressing a transfected human P-selectin cDNA, and adhering to surface-anchored platelets expressing P-selectin under static and flow conditions. The aim was to screen for heparin derivatives with high anti-adhesion activity but negligible anticoagulant activity. In this study, four modified heparins with high anti-adhesion activity were identified including RO-heparin, CR-heparin, 2/3ODS-heparin, and N/2/3DS-heparin. NMR analysis proved the reliability of structure of the four modified heparins. Our findings suggested that the 6-O-sulfate group of glucosamine units in heparin is critical for the inhibition of P-selectin-mediated tumor cell adhesion. Heparan sulfate-like proteoglycans on these tumor cell surfaces are implicated in adhesion of the tumor cells to P-selectin. Some chemically modified heparins with low anticoagulant activities, such as 2/3ODS-heparin, may have potential value as therapeutic agents that block P-selectin-mediated cell adhesion and prevent tumor metastasis.     

条件性缺氧诱导因子-1α RNAi转基因小鼠模型的建立

缺氧诱导因子1a (hypoxia inducible factor-1 a, HIF-1 a)是细胞在缺氧等条件下稳定表达的具有转录活性的蛋白,通过与多种靶基因调控区的缺氧反应元件(hypoxia response element, HRE)结合, 调控靶基因表达, 使机体对缺氧、缺血等病理生理过程产生适应性反应。为从整体动物水平研究HIF-1 a的作用, 需要建立HIF-1 a相关遗传修饰小鼠。分别针对HIF-1 a mRNA序列的两个靶位点合成两对互补的寡核苷酸链, 构建可诱导的RNA干扰真核表达载体HIF-AB和HIF-CD。分别将CRE重组酶真核表达载体CRE-ERT2与HIF-AB或HIF-CD转染入RAW264.7细胞, 筛选得到稳定表达CRE-ERT2与HIF-AB, 或CRE-ERT2与HIF-CD的稳定细胞系。在用4-HT诱导去除上述细胞系中HIF-AB或HIF-CD所含的Neo基因后, 用CoCl2诱导HIF-1 a表达, 采用半定量RT-PCR检测HIF-AB或HIF-CD对HIF-1 a 基因表达的影响。结果发现干扰载体(HIF-AB和HIF-CD)对HIF-1 a mRNA序列的沉默效果分别为85%和72%。选择干扰效率较高的表达载体HIF-AB经显微注射获得HIF-1 a基因敲低小鼠模型, 经PCR以及测序验证获得2个转基因阳性小鼠(Founders, G0代)。G0代雄鼠与FVB/N雌鼠交配后获得2只F1代(first filial generation)转基因阳性小鼠, 经与EIIA-Cre转基因小鼠交配, 得到EIIA-Cre; HIFRNAiflox/+小鼠, RT-PCR结果显示, EIIA-Cre; HIFRNAiflox/+小鼠肝、肺、肾等组织的HIF-1 a mRNA水平明显降低, 分别约为正常对照的44%、38.2%和23.5%。该小鼠模型的建立为进一步研究HIF-1 a的功能及作用机制提供了新的手段。