超声波用于海藻糖载入血小板

研究超声波对海藻糖载入人血小板过程的影响.将浓度约1×109个/ml的人血小板在含50 mmol/L海藻糖、100 mmol/L NaCl、10 mmol/L KCI,10 mmol/L EGTA、10 mmol/L咪唑的溶液中孵化,温度为37℃,持续时间4 h.孵化期间对各试样分别施加800 kHz、不同强度和不同时间的超声波辐射,以不施加超声波辐射的血小板样品为对照组.采用蒽酮硫酸法和分光光度计测量糖含量,并据此计算渗入血小板的海藻糖浓度.结果表明,在超声波辐射强度1=0.8 W/cm2、辐射时间1 h时海藻糖载入血小板的浓度最高,达(17.40±2.90)mmol/L,比对照组(11.27±2.53)mmol/L高54.3%.在光学显微镜下观察经超声波法载入海藻糖后的血小板,发现形态保持完好,与新鲜血小板形态几乎无差别,合适的超声波辐射能够有效强化海藻糖载入人血小板的过程,同时为进一步的血小板冻干研究提供了基础.     

海藻糖载入红细胞及其冷冻干燥的实验研究

冷冻干燥保存是长期保存人体红细胞的理想方案之一。冻干保护剂海藻糖渗入细胞内后,对细胞膜和细胞内物质有保护作用,其中的一个作用是增加细胞质的浓度,使冻干过程容易形成稳定的玻璃态。应用高渗法处理红细胞,通过考察胞内海藻糖含量、红细胞冻干后的存活率、腺苷三磷酸酶(ATPase)、超氧化物歧化酶(SOD)活力以及细胞形态变化,研究胞内海藻糖含量对红细胞冻干后活性的影响。结果显示:海藻糖对红细胞冻干具有明显的保护作用,随胞内海藻糖浓度升高,其保护性能逐渐增强;43.8mmol/L的胞内海藻糖浓度对红细胞保护最好,细胞存活率达到53.6%,形态保持良好,ATP和SOD活力均在正常的范围内。     

小菜蛾海藻糖酶酶学特性及四种杀虫剂对其活性的抑制作用

本试验以4龄小菜蛾Plutella xylostella(L.)体内海藻糖酶为研究对象,研究了其酶学特性以及4种常用杀虫剂在离体条件下对其活性的影响。结果表明,小菜蛾体内海藻糖酶的最适反应pH为6.0,温度50℃,其米氏常数(Km)为(12.57±0.99)mmol/L,最大反应速度(Vm)为(0.775±0.04)mmol/(min·g pro)。在药剂浓度为40mg/L时,4种杀虫剂(丙溴磷、灭多威、仲丁威和多杀霉素)对4龄幼虫海藻糖酶活性均有不同程度的抑制作用,其抑制率分别为25.83%、20.19%、18.99%和18.62%;而且随着药剂浓度的增加,上述4种杀虫剂对小菜蛾海藻糖酶活性的抑制率也逐渐增大。     

稀释液中添加海藻糖对山羊精子功能和膜完整性的影响

选用5只年龄为3～4岁的波尔山羊公羊研究在稀释液中添加海藻糖对山羊精子功能和膜完整性的影响。山羊精子分别用含6.6 mmol/L、13.2 mmol/L、19.8 mmol/L、26.4 mmol/L、39.6 mmol/L、52.9 mmol/L、66.1mmol/L、79.3 mmol/L的不同海藻糖的Tris-柠檬酸-葡糖糖(TCG)稀释液(卵黄:18%;甘油:6%)稀释和冷冻。结果表明:39.6 mmol/L、52.9 mmol/L、66.1 mmol/L、79.3 mmol/L组降温后的精子活率显著(P<0.05)降低;52.9 mmol/L、66.1 mmol/L、79.3 mmol/L组降温后的精子畸形率和39.6 mmol/L组降温后的膨胀精子率显著(P<0.05)提高。26.4 mmol/L组和39.6 mmol/L组冻融后的精子活率显著(P<0.05)高于对照组;66.1mmol/L和79.3 mmol/L组冻融后的精子活率、畸形率分别显著(P<0.05)低于和高于对照组。19.8 mmol/L、26.4 mmol/L、39.6 mmol/L组冻融后精子获能率显著(P<0.05)低于对照组。39.6 mmol/L组冻融后顶体完整率和膨胀精子率显著(P<0.05)高于对照组,而66.1 mmol/L组和79.3 mmol/L组显著(P<0.05)低于对照组。39.6 mmol/L组的受胎率显著(P<0.05)高于对照组,而66.1mmol/L组和79.3 mmol/L组的受胎率显著(P<0.05)低于对照组。结果表明,在含18%的卵黄(v/v)、6%甘油(v/v)的TCG稀释液中,添加适宜浓度(26.4mmol/L和39.6 mmol/L)海藻糖,可显著提高山羊精子功能和膜的完整性。     

酵母中海藻糖提取的响应曲面优化及纯化工艺

目的:针对现有技术存在的步骤复杂、环境污染和效率不高等问题,探索干酵母中海藻糖的高效提取纯化新工艺。方法:以提取温度、提取时间和氧化钙浓度为考察因素,以海藻糖浓度为响应值,采用响应曲面法优化海藻糖的干酵母热纯水提取工艺。以硫酸锌为絮凝基质,以氢氧化钡为pH调节剂,考察了硫酸锌浓度对一步法过滤纯化的影响。结果:当提取温度为78℃、时间为1.3 h、氧化钙浓度为12.6 g/L时,得到了海藻糖最大提取浓度为0.238 g/g干酵母。当硫酸锌浓度为35.71 g/L时,蛋白质脱除率达到99.1%,溶液电导率减小至569 us/cm,更有利于后期结晶。结论:整个过滤纯化工艺的海藻糖损失率仅为7.7%,远低于传统提取工艺。     

高浓度葡萄糖对昆明小鼠早期胚胎发育的影响

建立昆明小鼠受孕模型,分离并体外培养胚胎细胞.检测了各培养浓度下的细胞增殖、分化与凋亡.胚胎细胞在0.2mmol/L和5.56mmol/L葡萄糖浓度的KSOM培养基中能正常发育和孵化;而在浓度为15.56mmol/L和25.56mmol/L葡萄糖培养基中胚胎发育和孵化均受到损害(P<0.005),且总细胞数和内细胞团细胞数也明显减少(P<0.01),但其细胞凋亡率与0.2mmol/L和5.56mmol/L葡萄糖浓度下胚胎细胞凋亡率无显著性差异(P>0.05).随着葡萄糖浓度的增高,胚泡总的表面积无明显变化,但胚胎细胞密度呈增加趋势.高血糖对早期胚胎的发育具有毒性作用,提示高糖可能导致妊娠合并糖尿病患者的流产和胎儿畸形率升高.     

腾冲嗜热杆菌热稳定海藻糖磷酸化酶的基因表达与酶的分子生物学性质研究

分离克隆了腾冲嗜热杆菌(Thermoanaerobacter tengcongensis)海藻糖磷酸化酶(TreP)的编码基因(treP), 该酶可催化以葡萄糖和α-1-磷酸葡萄糖为底物的海藻糖合成反应及其逆向的分解反应. 反向mRNA点杂交实验表明, 腾冲嗜热杆菌中treP基因在高盐胁迫条件下表达量增加, 而在海藻糖诱导条件下表达量降低. 将该基因导入不含TreP的大肠杆菌中进行诱导表达, SDS-PAGE表明, 异源表达的TreP分子量约为90 kD, 与预期值相同. 通过葡萄糖氧化酶法测定分解产物葡萄糖的产率表明: TreP催化海藻糖分解反应的最适温度是70℃, 最适pH值为7.0; 通过HPLC检测合成产物海藻糖的产率表明: TreP催化合成反应的最适温度为70℃, 最适pH值为6.0. 在最适反应条件下, 50 μg的TreP粗酶可催化25 mmol/L α-1-磷酸葡萄糖与葡萄糖在30 min合成11.6 mmol/L海藻糖; 而同量的酶在同样时间内仅能将250 mmol/L海藻糖分解生成1.5 mmol/L葡萄糖. 以上体内胁迫和诱导表达分析及体外酶学性质分析均证明该酶的主要功能是催化海藻糖的合成反应. 热稳定性实验表明, 该酶性质比较稳定, 在50℃下温育7 h还能保持77%以上的活性, 是一个有潜在工业用途的新的热稳定海藻糖合成酶.     

海洋中海藻糖产生菌的筛选及发酵条件优化

从180余份海水、海泥样品中筛选得到60株产海藻糖较高的菌株,编号为2-14的菌株海藻糖产量最高,为127.9mg/g cell。对2-14菌株进行形态特征、培养特征及生理生化试验,鉴定该菌株为红酵母属(Rhodotorula sp.)。研究摇瓶发酵条件对红酵母海藻糖产量的影响,结果为:初始pH5.5,发酵温度28℃,装液量75mL(250mL三角瓶中)。采用优化后发酵条件红酵母海藻糖产量为193.3mg/g cell,优化前对照值为132.1mg/g cell,优化后的结果是优化前的1.46倍。在5L发酵罐中培养得到最佳发酵时间为54h,发酵罐培养发酵液中海藻糖含量最高达2.5g/L,为摇瓶培养的1.6倍。     

二甲双胍对人脐带间充质干细胞形态、增殖、表面标志及细胞周期的影响

目的探讨不同浓度二甲双胍（METF）对人脐带间充质干细胞（hUC-MSC）形态、增殖、表面标志及细胞周期的影响。 方法取健康足月新生儿脐带在体外分离出hUC-MSC进行传代培养,至第3代（流式细胞仪分析）对细胞进行鉴定,取第6代处于对数生长期的hUC-MSC （相对老化）,将对照组与不同浓度METF （0.1,1,5,10,20?mmol/L）干预的细胞进行比较,观察不同浓度METF干预对细胞的形态、增殖率（MTT法分别于24、48、72?h检测）、及细胞表面标志和细胞周期的影响,采用One-Way ANOVA,及LSD-t检验进行统计学分析。 结果（1）METF为0.1?mmol/L、1?mmol/L,细胞形态无显著改变,当药物浓度为5?~?20?mmol/?L时,随着药物浓度增加、培养时间延长,细胞形态改变越显著。（2）METF为0.1?mmol/L（24?h：101.28±0.98,24?h：104.06±1.76,24?h：101.51±0.67）促进hUC-MSC增殖,药物浓度为1?~ 10?mmol/L在培养初期可增加间充质干细胞的增殖率,随着培养时间的延长,细胞的增殖逐渐被抑制。METF为20?mmol/L（24?h：86.64±0.66,48?h：58.38±2.52,72?h：17.75±1.35）抑制细胞增殖,抑制作用随着时间延长而增强（P?< 0.05）。（3）当METF浓度为5,10,20?mmol/L时,随着药物浓度的增加,CD105的表达逐渐减弱（F?= 17.539,P?< 0.05）。METF未对CD44、CD90产生影响。（4）METF为0.1?mmol/L时降低G0/G1期的比例（64.16±1.20,P?< 0.05）,促进间充质干细胞的增殖,随着药物浓度的增加,细胞增殖逐渐被抑制。 结论METF浓度在0.1mmol/?L促进hUC-MSC增殖,而在浓度5 ~ 20?mmol/L时抑制人脐带间充质干细胞的增殖及表面标志CD105的表达,不同浓度的METF均未对CD44、CD90的表达产生影响。     

高糖对海马神经元形态破坏和APP蛋白含量的影响

为探索高糖培养对原代海马神经元形态损伤和神经退行性相关蛋白β-淀粉样蛋白前体(amyloid β-protein precursor,APP)含量的影响,分离胎龄16 d的大鼠海马神经元,分别使用含有25 mmol/L、50 mmol/L、75 mmol/L和100 mmol/L葡萄糖浓度的Neurobasal培养基进行原代培养干预,Western-blot检测APP蛋白水平,光学显微镜下观察不同浓度葡萄糖作用后海马神经元形态的变化。结果发现:高糖培养后,胎鼠海马神经元突触变短,胞体肿胀,同时APP水平随着葡萄糖浓度的递增逐渐升高。以上信息提示高糖引发的神经元形态破坏与APP高水平有关,控制血糖可能有利于保护神经元细胞,使其免受损伤。     

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.