无CpG基序的短磷酸二酯寡核苷酸作免疫激活剂

以前发现细菌的DNA能作用于免疫效应细胞。据报道,这种免疫调节作用是因为存在有未甲基化的CpG二核苷酸,在二核苷酸的侧面分别毗邻两个5′嘌呤和两个3′嘧啶,合成的8～100个碱基的寡核苷酸至少含有这样一个CpG基序以刺激免疫系统。这些寡核苷     

免疫佐剂作用机制及CpG ODN作为新型佐剂的研究

免疫佐剂通过免疫调节作用等5种方式发挥功能,借助佐剂的非抗原特异性的多克隆激活是天然免疫细胞可能的作用机制.大部分佐剂通过直接参与天然免疫事件而间接/直接影响获得性免疫应答,不同的佐剂可针对不同的事件发挥作用.CpG ODN作为佐剂的优势在新近的研究中被逐渐发现.     

CpG 寡核苷酸的安全性实验研究

目的研究CpG 寡核苷酸(CpG ODN)作为免疫佐剂应用可行性.方法按要求配制CpG ODN检测内毒素,采用昆明小鼠和豚鼠进行异常毒性试验和超敏试验;分别以10 μg /只、100 μg /只、300 μg /只3个剂量组CpG ODN进行小鼠骨髓嗜多染红细胞微核试验;以10 μg /只、30 μg/只、90 μg/只3个剂量组的CpG ODN分别交替于双侧后肢胫前肌肌肉注射Balb/c小鼠,每周2次,连续4周,进行血液学、血生化、病理组织学检查及抗核抗体、抗双链DNA抗体和肿瘤坏死因子检测.结果内毒素检测阴性,实验期内所有动物均未出现异常毒性反应和超敏反应,实验结束后所有动物全部健存且体重增加. CpG ODN小鼠骨髓嗜多染红细胞微核试验为阴性结果;实验期间各组动物未见异常表现,体重增加各组间差异无显著性,血液学、血生化和病理组织学检查未见明显异常.未检测到ANA、ds-DNA.各剂量组肿瘤坏死因子含量在与对照组之间比较差异均有显著性.结论所选用CpG ODN作为生物制品使用是安全的;在受试剂量下微核试验阴性;在小鼠亚急性毒性实验中未引起毒性损害,安全性良好.     

CpG模体的免疫激活机制及应用

CpG模体是一些在机体内外具有强烈免疫激活功能的寡聚脱氧核苷酸短序列。章综述了近年来有关CpG模体作用机制及其应用方面的研究进展。在机制方面,着重对CpGDNA细胞激活和信号转导两方面进行了详尽的阐述;在应用方面,重点介绍了其作为疫苗佐剂和免疫性疾病治疗药物的研究现状。     

引入CpG基序的汉滩病毒G2糖蛋白基因的克隆及表达

构建汉滩病毒G2糖蛋白的真核表达载体,并加入可增强小鼠免疫刺激作用的CpG基序,检测其可否在真核细胞中表达。参照Genebank中汉滩病毒M的全基因序列设计引物,引物两端引入可增强小鼠免疫刺激作用的CpG基序及双酶切位点,通过聚合酶链反应(PCR)获得含CpG基序的G2片段,并将其与T载体pMD18-T相连,测序后克隆至真核表达载体pcDNA3.1 上,将此真核表达载体以脂质体法转染至真核细胞Vero-E6中,利用间接免疫荧光法(IFA)检测发现转染后的Vero-E6中出现特异性的绿色荧光。结果表明本实验成功构建了汉滩病毒包膜糖蛋白G2的重组体。     

CpG基序重组质粒增强小鼠免疫和抗病力的效应

为研制安全高效免疫调节剂增强动物免疫抗病能力,本实验设计合成含11个CpG基序的寡聚核苷酸,重组构建含CpG的VR1012质粒(VR1C);制备壳聚糖纳米颗粒包裹重组质粒(CNP-VR1C),肌注接种3周龄昆明小鼠;接种后28天口服大肠杆菌攻毒观察小鼠天然免疫的变化和对强毒感染的抵抗力,Sandwich ELISA测定血清免疫球蛋白和白细胞介素含量。结果表明:CNP-VR1C能显著提高小鼠的天然免疫水平,增强对强毒感染的抵抗力,具有研制为高效安全分子免疫增强剂的应用潜力。     

Toll样受体与新型佐剂—佐剂研究的新趋势

佐剂是疫苗开发的一部分。Toll样受体是一类重要的模式识别受体,能通过识别不同病原体的PAMP在连接固有免疫与特异性免疫的关键环节发挥着极为重要的作用。最早发现并作为佐剂使用的TLR配体是CpG。CpG通过TLR9介导对免疫系统的激活作用,从而发挥诱导Th1偏倚的Th1/Th2混合反应的佐剂效应,已经受到了广泛的关注和应用。近年来,研究发现一类具有免疫调节作用的抗病毒治疗的药物Imidazoquiline(咪唑并奎琳衍生物)是合成的TLR7/8的配体,也具有佐剂活性,但效果还存在争议。TLR配体作为佐剂的使用为佐剂的研发开拓出一条新的思路,带来了新的希望。     

猪白细胞介素4,6融合基因和CpG基序对猪蓝耳病疫苗免疫的强化效应

为探索新型高效安全的猪蓝耳病疫苗免疫佐剂,本实验用离子交联法制备壳聚糖纳米颗粒包裹猪融合基因PIL-46和CpG基序的真核表达质粒(CNP-(VRIL-46 VR1C)),将CNP-(VRIL-46 VR1C)肌注接种过猪蓝耳病灭活苗的长白川白杂交猪,接种后1、2、4、6和8周采取实验猪静脉血,用Sandwich ELISA检测血清中白细胞介素2(IL-2)、IL-4、IL-6和IgG、IgA、IgM及特异抗体的含量,同时检测外周血液免疫细胞数量的变化。结果发现实验猪接种CNP-(VRIL-46 VR1C)后1～8周内,其血清中IL-2、IL-4、IL-6和IgG、IgA、IgM及特异抗体的含量较对照组均显著增多(P<0.05),淋巴细胞和单核细胞数量也明显升高(P<0.05)。这些表明接种CNP-(VRIL-46 VR1C)的实验猪的特异体液和细胞免疫水平均显著多于对照组,提示PIL-46基因和CpG基序组合能显著增强猪的特异体液和细胞免疫机能,明显提高其免疫防御力,具有研制为高效安全的分子免疫增强剂的应用前景。     

MK──一种新发现的细胞因子

MK是一种新发现的细胞因子,属于肝素结合因子家族,MK是一种小分子多肽,其基因表达仅见于胚胎中期及成年期肾脏,在许多肿瘤细胞中也可见MK基因不同程度的表达,MK能够促进正常细胞的生长和分化,特别是促进神经细胞的发育,它还可以抑制某些肿瘤细胞的生长．MK基因在成年肾脏中表达的原因尚未阐明．     

一种新型可用于DNA分子量标准的质粒构建

将首尾带有EcoR Ⅰ酶切位点的2．0kb、1．5kb、1．0kb、0．75kb、0．5kb、0．25kb六个长度的片段逐步连接到pGEM-3zf( )质粒上的B．am H I位点中,构建的质粒用EcoR Ⅰ进行单酶切,经电泳可以获得七条DNA带与设计结果完全一致,可用于DNA电泳试验中分子量标准。     

抗溃疡性结肠炎新药IPSALAZIDE的合成

以对硝基苯甲酸为原料,经氯化后,被氨基乙酸取代,得到对硝基苯甲酰氨基乙酸,再经还原,重氮化后,与水杨酸偶合,合成了抗溃疡性结肠炎新药－５［［４－［（羧甲基）甲酰氨］苯］偶氮基］水杨酸（Ｉｐｓａｌａｚｉｄｅ）。并做了红外光谱鉴定。     

抗溃疡性结肠炎新药—Balsalazide的合成

以对硝基苯甲酸为原料,经氯化后,被β—氨基丙酸取代,得到对硝基苯甲酰—β—氨基丙酸,再经还原、重氮化后,与水杨酸偶合,合成了抗溃疡性结肠炎新药—５—〔［４［（β－羧乙基）甲酰氨］苯］偶氮基〕水杨酸（Ｂａｌｓａｌａｚｉｄｅ）,并做了红外光谱鉴定。     

A New Kind of Informational Suppression in the Nematode Caenorhabditis Elegans

Independent reversions of mutations affecting three different Caenorhabditis elegans genes have each yielded representatives of the same set of extragenic suppressors. Mutations at any one of six loci act as allele-specific recessive suppressors of certain allels of unc-54 (a myosin heavy chain gene), lin-29 (a heterochronic gene), and tra-2 (a sex determination gene). The same mutations also suppress certain alleles of another sex determination gene, tra-1, and of a morphogenetic gene, dpy-5. In addition to their suppression phenotype, the suppressor mutations cause abnormal morphogenesis of the male bursa and the hermaphrodite vulva. We name these genes smg-1 through smg-6 (suppressor with morphogenetic effect on genitalia), in order to distinguish them from mab (male abnormal) genes that can mutate to produce abnormal genitalia but which do not act as suppressors (smg-1 and smg-2 are new names for two previously described genes, mab-1 and mab-11). The patterns of suppression, and the interactions between the different smg genes, are described and discussed. In general, suppression is recessive and incomplete, and at least some of the suppressed mutations are hypomorphic in nature. A suppressible allele of unc-54 contains a deletion in the 3' noncoding region of the gene; the protein coding region of the gene is apparently unaffected. This suggests that the smg suppressors affect a process other than translation, for example mRNA processing, transport, or stability.     

A New Turgor/Membrane Potential Probe Simultaneously Measures Turgor and Electrical Membrane Potential

Abstract: A new combined turgor/membrane potential probe (T-EP probe) monitored cell turgor and membrane potential simultaneously in single giant cells. The new probe consisted of a silicone oil-filled micropipette (oil-microelectrode), which conducted electric current. Measurements of turgor and hydraulic conductivity were performed as with the conventional cell pressure probe besides the membrane potential. In internodal cells of Chara corallina, steady state turgor (0.5-0.7 MPa) and resting potentials (-200 to ?220 mV) in APW, and hydraulic conductivity (0.07 to 0.21 × 10～⁵ m s^?1 MPa^?1) were measured with the new probe, and cells exhibited healthy cytoplasmic streaming for at least 24 h during measurements. When internodal cells of Chara corallina were treated with 30, 20, 10, and 5 mM KCI, turgor responded immediately to all concentrations, and the osmotic changes in the medium were measured. Action potentials, which brought the membrane potential to a steady depolarization that measured the concentration difference of K⁺ in the medium, were induced in a concentration — dependent delay and occurred only 30, 20, and 10 mM of KCl. When the solution was changed back to APW, the repolarization of membrane potential consisted of a quick and a following slow phase. During the quick phase, which took place immediately and lasted 1 to 3 min, the plasma membrane remained activated. The membrane was gradually deactivated in the slow phase, and entirely deactivated when the membrane potential recovered to the resting potential in APW. Although the activated plasma membrane was permeable to K⁺, no major ion channels were activated on the tonoplast, and therefore, internodal cells of Chara corallina did not regulate turgor when osmotic potential changed in the surrounding medium.