β3 Adrenergic Receptor Polymorphism and Obesity‐Related Phenotypes in Hypertensive Patients

Objectives: Obesity is a complex trait that is affected by both environmental and genetic risk factors. The β₃ adrenergic receptor (ADRB3) is expressed in adipose tissue and plays a role in energy metabolism. A missense mutation on codon 64 of this gene (W64R) is associated with receptor malfunction. Previous studies examining the relation between this polymorphism and obesity produced inconsistent findings. The current study assessed the association between the W64R genotype and obesity‐related phenotypes, including body weight, BMI, and serum triglycerides, cholesterol, and glucose. Research Methods and Procedures: We determined the ADRB3 W64R genotypes and fasting serum lipid and glucose concentrations for 695 hypertensive adults (336 men, 359 women) from a rural county in Anhui Province, China. Multivariate linear regression models were fit to detect associations between the genetic polymorphism and obesity‐related phenotypes. Results: The ADRB3 W64R polymorphism was significantly associated with body weight and BMI in men but not in women. After controlling for potential confounding variables, men who were homozygous for the R64 allele were 11.8 kg heavier (p < 0.001) and had a BMI that was 3.7 kg/m² greater (p = 0.001) than men who were homozygous for the W64 allele. Serum concentrations of lipids and glucose were found not associated with the genetic polymorphism. Discussion: The ADRB3 R64 allele was associated with increased body weight and BMI in men but not in women. The genetic association was not modified by triglyceride, cholesterol, blood glucose, or blood pressure levels of the subjects.     

鳜传染性脾肾坏死病毒主要衣壳蛋白基因的原核表达

根据鳜鱼传染性脾肾坏死病毒(Infectious spleen and kidney necrosis virus,ISKNV)丰要衣壳蚩白(Major Capsid protein,MCP)基因(mcp)序列设计引物,PCR扩增得到一长约1400bp的DNA片段,将其克隆到pGEM-T Easy Vector。氨基酸亲水性分析表明,在150—250位氨基酸之间亲水性很高,可构成主要抗原决定簇及形成跨膜区。mcp基因经PCR改造后克隆至原核表达裁体pBV220,构建表达MCP的大肠杆菌基因工程菌,该工程菌经42℃诱导,SDS—PAGE检测,在约50kDa处有一特异蛋白带,含量约为菌体总蛋白的23％。用纯化和复性后的蛋白免疫新西兰大白兔制备抗血清,Western—blotting分忻显示,重组MCP制备的抗血清能与ISKNV MCP特异结合,说明表达产物具有与ISKNV MCP相似的抗原特性。     

OEP及卵黄浓度对蓝狐冻融精子质量的影响

人工采取 6只优质芬兰雄性蓝狐精液 ,利用不同OEP及卵黄含量的Tris 果糖 -柠檬酸钠稀释液稀释 ,制成细管冻精 ,透射电镜下观察精子冷冻前后质膜和顶体超微结构 ,荧光免疫方法检测不同培养时间冻融精子的质量。结果表明 ,蓝狐精子顶体外膜双层膜的厚度为 0 0 2 0 μm ,冷冻 -解冻过程中易发生质膜膨胀、顶体外膜融合现象。顶体产生的囊泡分两种类型 ,一种是体积较大的中空囊泡 ,平均直径为 1 2 5 μm。另一种是体积较小的实体囊泡 ,内充满顶体内容物 ,平均直径为 0 83μm ,两种囊泡的数量不定。OEP能有效抑制顶体囊泡形成 ,影响顶体囊泡类型、体积大小及囊泡数量 ,添加适宜剂量OEP能使顶体囊泡的体积明显缩小 ,囊泡的总数及中空囊泡的数量显著降低。蓝狐冻融精子质量与OEP及卵黄剂量有关 ,在卵黄存在的前提下 ,OEP有利于维持冻融过程中质膜 (5 6 3% )、顶体的完整性 (5 7 8% ) ,显著提高冻融精子活力 (5 4 7% )。在蓝狐精液稀释液中 ,OEP、卵黄的适宜含量分别为 1 %、 2 0 %     

春雷霉素高产菌株US95#的选育及大罐发酵工艺控制

紫外线与光修复交替处理春雷霉素产生菌得到了高产突变株,再结合自然分离选出了US95^#。该菌株不但产量高,还具有稳定的遗传性能,并在大罐良好发酵工艺控制和条件下,罐发酵水平在春雷霉素生产上有了重大突破,发酵水平比历史最高水平提高12．9％。     

猪流感病毒抗原表位基因表达载体构建与免疫原性分析

根据猪流感病毒血凝素蛋白基因(Heamuglutinine, HA)的核苷酸序列, 设计、筛选HA蛋白氨基酸序列的主要表位多肽4个, 将4个片段以柔性连接串联成模拟蛋白, 核苷酸约为300 bp, 体外扩增该模拟蛋白基因, 插入到原核表达载体pET30a(+)中, 转染宿主菌诱导表达, 结果获得分子量为20 kD的表达蛋白, 该蛋白可与抗His-tag抗体、抗猪流感病毒H1N1、H3N2亚型高免血清发生免疫学反应。纯化后免疫小鼠, ELISA及血凝抑制(Heamuglutinine inhibitor, HI)试验检测, 小鼠产生针对多肽抗原的血清抗体, 同时还可检测到H1N1、H3N2亚型SIV血凝抗体。流氏细胞仪检测免疫组外周血淋巴细胞高于对照组, 说明该模拟蛋白具有与H1N1、H3N2亚型猪流感病毒相似的免疫原性及反应原性, 为H1N1、H3N2血清亚型猪流感病毒疫苗研制提供了新手段。     

Inhibitory effect of obestatin on glucose-induced insulin secretion in rats

Obestatin is a bioactive peptide encoded by the same gene that encodes ghrelin. Our aim was to investigate the effect of obestatin on insulin secretion. We evaluated the effects of obestatin on insulin secretion from rat islet cells which had been incubated overnight in the presence of 8.3, 11.1, and 22.2 mmol/l of glucose. In vivo, the serum levels of glucose and insulin were measured 0, 1, 5, 10, 20, 40, and 60 min after the intravenous administration of saline or glucose (1 g/kg), with or without obestatin, and the area under the 60 min curve of insulin concentration (AUC_insulin) was calculated. Obestatin (0.01-100 nmol/l) inhibited insulin secretion from rat islets in a dose-dependent fashion. In vivo, when administered intravenously to rats together with glucose, obestatin (10, 50, and 250 nmol/kg) inhibited both the rapid 1-min insulin response and the AUC_insulin in a dose-dependent fashion. Our data demonstrate that under glucose-stimulated conditions, exogenous obestatin acts as a potent inhibitor of insulin secretion in anaesthetized rats in vivo as well as in cultured islets in vitro.     

Co-segregation of the T1095C with the A1555G mutation of the mitochondrial 12S rRNA gene in a patient with non-syndromic hearing loss

We reported here the clinical and molecular characterization of a Chinese subject with childhood-onset hearing impairment. Clinical evaluations showed that the patient suffered from profound and non-syndromic sensorineural hearing loss with flat configurations. Sequence analysis of the mitochondrial 12S rRNA and tRNA^Ser(UCN) genes led to the identification of double deafness-associated mutations of A1555G and T1095C in the 12S rRNA gene which apparently in the homoplasmic forms. In additional, there was no other functionally significant nucleotide variants found in this subject. As previous studies have indicated that the A1555G mutation was a primary contributing factor underlying the development of deafness but not sufficient to produce clinical phenotype, the co-segregation of two mitochondrial DNA mutations raises the possibility that the T to C transition at position 1095 plays a role in the phenotypic expression of deafness-associated A1555G mutation. Actually, the T1095C mutation disrupted an evolutionarily conserved base-pair at stem-loop of helix 25 of 12S rRNA, resulting in impaired translation in mitochondrial protein synthesis and a significant reduction of cytochrome c oxidase activity. As a result, it may enhance the biochemical defect in patient carrying the A1555G mutation, thus changing the age of onset and the severity of hearing impairment.     

枸杞原生质体培养及高效成株体系的建立

由枸杞髓部组织诱导出胚性愈伤组织,并由此愈伤组织建立起稳定的细胞悬浮系。从悬浮细胞游离的原生质体在改良KM培养基(1.5 mg/L 6_BA,0.5 mg/L NAA和0.5 mg/L 2,4_D)中进行液体浅层培养,3～4 d后出现第一次分裂,第7 d统计分裂频率为50.3%,15 d左右可形成细胞团,3～4周后形成肉眼可见的愈伤组织,愈伤组织植板率为1.25%。将细胞团转移到液体分化培养基(MS+6_BA 1.5 mg/L+2,4_D 0.2 mg/L) 8～10 d可形成大量胚状体,及时将胚性愈伤组织块转移到固体分化培养基上(MS+6_BA 0.2 mg/L),可形成大量绿芽,分化率54.17%。绿芽在生根培养基（MS+NAA 0.2 mg/L）可形成完整植株,移栽后成活良好。     

红鲫近交系的建立

目的应用人工雌核发育繁殖技术建立红鲫近交系.方法红鲫卵子被经紫外线照射的鲤鱼精子激活后,在0～4℃的温度下进行冷休克处理30min,再在常温下静水孵化;以红体色为遗传标记,在连续2代雌核发育子代中选择二倍体红鲫,放置水泥池中常规饲养;用同样的人工雌核发育繁殖技术进行保种,随机交配繁殖1～2代供应实验;用常规方法测量形态学指标.结果红鲫卵子激活率在91%以上;实验组2中摄食期仔鱼成活率为15.20%,80%的成鱼为二倍体红鲫,其中有较高比例的遗传上真实的雄性个体.鳞式主要为27(5)/(6)28;鳍式主要为D.i,17～18;A.i,6;V.i,7～8;P.i,15～16;C.24～25;其他外部形态指标与封闭群红鲫近似.结论本实验建立的人工雌核发育繁殖技术,设备要求简单,操作安全可靠,并且省时节资,完全能够用于培育鱼类近交系,本实验结果完全可以满足育种繁殖的要求;所获得的二倍体红鲫,经同工酶电泳等检测,被证实是纯合的,具备鱼类近交系品系特征;出现遗传上真实的雄性个体是正常的.     

Raft培养可诱导人正常上皮细胞分化

人的正常上皮细胞在一般的体外培养情况下很难分化成类似于机体的上皮样结构。为了得到上皮结构进行相关研究,近年来,国外建立了Raft（船式）培养方法。本文在长期从事细胞培养的工作中,摸索出了适合国内一般实验室条件的Raft细胞培养操作方案,模拟体内人皮肤结构的分化层次,以纤维细胞与胶原作为混合基质（作为上皮细胞的滋养物）,同时加入一些生长因子、激素和必需的蛋白等成分。将纤维细胞、胶原和添加成分做为基层过夜培养,次日将培养好的上皮细胞种到上面,过夜培养后即为“skin equivalent”（皮肤等价物）,将“皮肤等价物”移到一种网状金属架上以使细胞在气－液状态下培养,在这种条件下,培养中的上皮细胞与体内上皮细胞自然生长的状态、环境非常类似,在体外导致上皮细胞分化成清晰可见的层次。此项技术在临床治疗烧伤以及其它皮肤损伤的疾病方面有潜在的应用价值,也为研究皮肤的正常生理功能以及癌症的发生、发展以及治疗提供了一种模型。