BALB/c和C_(57)BL/6小鼠在基因功能检测中行为学的比较研究

目的　探索BALB c和C57BL 6两个品系在有关实验中的不同作用。方法　选取了结合随机测序与生物信息学分析设计合成的神经系统表达的一些基因的反义核酸 (anti sense)中的 2个 ,用Hamilton微量注射器将其分别定量注射到BALB c和C57BL 6小鼠的侧脑室 ,并分别设注射生理盐水和随机序列核酸 (Scramble)的对照组。每一反义核酸实验组和对照组各注射 1 0只小鼠 ,之后观察实验组与对照组在不同行为学实验中的差异。小鼠的行为学检测模型为 :考察日常代谢能力的摄食量 ,考察Locomotionactivity(移动 )的旷场行为 ,考察疼痛阈值的甩尾试验和考察记忆能力的步下法实验。结果　注射No 1基因的反义核酸后 ,两品系的实验组均在测试记忆力的步下法 (Step -downTest)试验中表现出记忆力减弱 ,且与对照组差异明显 ,说明No 1基因的功能确与记忆相关。注射No 2基因的反义核酸后 ,在测试移动能力的旷场行为 (OpenFieldBehavior)试验中 ,BALB c实验组跨格、直立行为均比对照组明显减少 ,说明受此反义核酸影响显著 ,而C57BL 6实验组则与对照组无大的差异。此外 ,在生理盐水对照组和随机序列核酸对照组的实验中以及其他行为学模型的实验中 ,两品系也存在着一定的差异。结论　用遗传背景不同的多品系进行相关实验 ,可进一步建立     

食物营养成分变化对卡氏小鼠和黄毛鼠摄食消化的影响

动物从环境中获取足够营养和能量的能力决定着其生存和繁殖。为探讨同域分布不同体型的鼠类对不同营养成分食物的摄食消化对策,以三种不同营养成分食物分别饲喂卡氏小鼠和黄毛鼠,在第10天和20天,以食物平衡法测定日摄食量、日粪便量、净摄入量和表观消化率。结果发现,在整个驯化过程中,两种鼠的不同食物组间体重无显著差异,表明均能够适应食物营养的变化维持收支平衡。在驯化第10天时,高蛋白组卡氏小鼠的日摄食量显著高于高淀粉组,但两组与普通组均无差异;高蛋白组卡氏小鼠的日粪便量显著高于高淀粉组和普通组;而净摄入量无组间差异;高淀粉组和普通组的表观消化率显著高于高蛋白组。至第20天时,三组动物日摄食量无差异;高蛋白组的日粪便量高于高淀粉组和普通组。黄毛鼠仅在第10天时高淀粉组和普通组表观消化率高于高蛋白组,三组黄毛鼠其他消化参数在第10天和第20天均无显著组间差异。结果表明,体型不同的鼠类因消化冗余能力的差异,面对相同的食物营养成分变化,会以不同的摄食消化策略调节适应。     

高纤维食物对黄毛鼠摄食和消化的影响

动物的消化生理特征和消化对策可决定其营养生态位。为揭示黄毛鼠适应高纤维食物的消化对策,在高纤维食物饲喂10 d和20 d时,以食物平衡法测定摄食量、粪便量及相应的能量学参数。在取食高纤维食物10 d时,摄食量无显著变化,但粪便量显著增加,消化率显著下降;至20 d时,摄食量显著增加,消化率仍低于对照组,但消化能与对照组无差异。高纤维组动物的体重在第10 d时显著下降,但在第20 d时未进一步下降。该结果表明,黄毛鼠能够通过增加摄食量和降低消化率的消化对策适应高纤维食物,此外,降低体重使总能量需求减少,也是其适应低质量食物的对策之一。     

影响Ⅱ型糖尿病动物模型KK／Upj—A^y／J小鼠血糖的因素

目的探讨营养,年龄及性别等因子对Ⅱ型糖尿病动物模型-KK／Upj-A^y／J小鼠血糖的影响。方法尾部取血,用血糖仪测定实验组和对照组动物的血糖并进行实验分析。结果实验组血糖值明显高于对照组;实验组发病动物的比例也明显高于对照组动物;对照组8月龄小鼠血糖值低于糖尿病的发病标准;实验组年龄对动物的血糖影响较小;实验组和对照组性别基本不影响动物的血糖。结论（1）在同样携带A^y黄色肥胖糖尿病基因的遗传作用的条件下,饲料中的营养成分是影响小鼠血糖的主要因素,（2）动物的年龄是影响血糖的另一主要因素。（3）性别对动物血糖的影响不大。     

筛选神经系统基因功能的行为学检测平台的探索

为大规模鉴定哺乳动物神经系统新基因的功能,利用反义核酸技术建立了一个小鼠行为学检测平台。从大鼠脑低丰度表达基因的cDNA亚库中挑选出在大鼠脑内表达的20个基因,设计并合成了这些基因的小鼠同源基因的特异性反义核酸。用Hamilton微量注射器将反义核酸注射到小鼠的侧脑室,并以注射生理盐水和随机序列核酸组作为对照。小鼠的行为学检测模型为;考察日常代谢能力的摄食量,考察Locomotion activity(移动）的旷场行为,考察疼痛阈值的甩尾试验和记忆能力的步下法实验,结果显示,上述20个基因中的14个在不同行为学模型中与对照组有显著性差异。     

真(鱼岁)(Phoxinus phoxinus(L.))的能量收支各组分与摄食量、体重及温度的关系

在不同的摄食水平(饥饿—最大量)及温度(5—15℃)下,对1—5g的真(鱼岁)的摄食量、排粪量、排泄量、代谢量,生长量及生化组成作了测定。真(鱼岁)的最大摄食量随体重及温度增加而增加。食物能量平均有6.5％损失于粪便中,5.1％损失于排泄物中。摄食代谢随摄食量增加而增加。在同一温度下,特定生长率与摄食量的关系是一减速增长曲线。当摄食不受限制时,生长率随温度增加而增加;当摄食受限制时,生长率随温度增加而下降。鱼体的干物质含量及能量含量随摄食量增加而增加。     

我国淡水优质草食性鱼类的营养和能量学研究 Ⅱ.草鱼、团头鲂对七种水生高等植物的最大摄食量和消化率的测定

作者采用常规生化分析法测定草食性鱼类天然食料(七种水生高等植物)的生化成分和能值。在实验室特定条件下测定了草鱼和团头鲂的系列体重组对七种水生高等植物的最大摄食量和消化率。1.七种水生高等植物的生化成分的含量和能值的分析结果表明,菹草、黄丝草、聚草、苦草、小茨草、轮叶黑藻和紫背浮萍的水分百分含量范围为80.85—92.72%、蛋白质为1.69—4.96%、脂肪为0.35—1.09%、灰分为0.98—2.6%、纤维素为0.66—2.17%、无氮浸出物为2.3—9.95%。七种水生植物的蛋白质均含有常见十七种氨基酸,包括鱼体必需氨基酸九种和非必需氨基酸八种。其中菹草的氨基酸含量最高(36.63mg/g 湿重)、苦草最低(12.92mg/g 湿重);菹草的能值最高(3.36KJ/g 湿重)、小茨藻最低(1.28KJ/g 湿重)。2.草鱼、团头鲂对七种水生植物的最大摄食量(Y,g/24h)及消化率(Y,%)与鱼体重(X,g)相关显著,关系式为 Y=aX~b。从结果表明,随着鱼体重的增加,鱼对七种食料的相对摄食量下降(绝对摄食量上升),消化率提高。草鱼的最大摄食量一般比团头鲂高,而消化率一般比团头鲂低。       

中国实验动物学报2002年目次索引2002年第10卷1～4期

第一期裸鼠肿瘤动物模型ＶＥＧＦ受体表达及其意义李钧敏　陆洪芬　田培坤等 (1)………………………………………分离和克隆小鼠ＥＳ细胞集落的主要影响因素周欣　王太一　时伟红等 (5 )………………………………………四个与小鼠摄食、摄水量及蛋白质代谢相关基因的功能研究敖红　丁澄　李晨等 (10 )………………………………2型糖尿病大鼠模型的建立及其糖代谢特征分析张芳林　李果　刘优萍等 (16 )……………………………………季节性变化对雌性恒河猴生殖功能的影响王训立　周建华　范春梅 (2 1)……………………………………………清洁…     

低氧影响蛋白质合成和分解的平衡诱导骨骼肌萎缩

暴露在低氧环境下,可能会引起胃肠功能障碍和摄食量下降,打破骨骼肌蛋白质合成和分解平衡,造成骨骼肌萎缩。为探讨低氧环境下骨骼肌的萎缩是低氧环境引起的还是低氧诱发的摄食量减少所致,本研究检测大鼠腓肠肌中低氧时蛋白质合成与分解相关基因的蛋白质表达。将21只雄性SD大鼠,随机分为3组：常氧对照组、低氧组（氧浓度为12.4%,模拟海拔4 000 m高度）和配对组（大鼠的摄食量与低氧组前1 d的摄食量相同）,每组7只,每天记录大鼠体重和摄食量。4周后,HE染色法观察腓肠肌肌纤维形态,Western印迹测试相关蛋白质水平。低氧组和配对组摄食量在低氧干预初期,较常氧对照组有显著性下降(P<0.05),干预后期差异不明显;干预期间,低氧组大鼠体重平均增加量(102.10 g)、体重(341.20 ± 16.75 g)、肌肉总量（226.83 ± 8.33 g）和腓肠肌肌纤维横截面积(12.67 ± 1.83 mm)较常氧对照组（128.00 g;377.50 ± 20.75 g;260.50 ± 9.35 g;15.78 ± 2.38 mm）和配对组（119.40 g;375.86 ± 11.30 g;262.29 ± 7.90 g;15.71 ± 2.82 mm）均显著下降,配对组较常氧对照组无显著性差异;4周干预后,与常氧对照组相比,低氧组大鼠腓肠肌中与低氧相关的HIF1α显著增加(1.42 ± 0.19, P<0.05),Akt和p-Akt/Akt显著降低 (1.44 ± 0.13; 0.47 ± 0.08, P<0.05),配对组上述3种指标相对表达量均无显著性差异;在蛋白质合成方面,低氧组mTOR较常氧对照组显著下降(0.63 ± 0.18, P<0.05),配对组较常氧对照组差异不明显;低氧组腓肠肌中,4EBP1(1.14 ± 0.14)和p70S6K1(1.14 ± 0.11)较配对组显著下降(P<0.05)。在蛋白质分解方面,低氧组p-FoxO1和p-FoxO1/FoxO1比值较常氧对照组显著下降(0.71 ± 0.15; 0.78 ± 0.14, P<0.05);低氧组大鼠腓肠肌中,Atrogin1、MuRF1、Beclin1、LC3Ⅰ及LC3Ⅱ/Ⅰ比值均高于常氧对照组(1.35 ± 0.12; 1.30 ± 0.22; 1.17 ± 0.11; 1.03 ± 0.11; 1.35 ± 0.13, P<0.05);配对组与常氧对照组间无明显差异。低氧环境下骨骼肌中蛋白质合成相关基因表达减少,蛋白质分解相关基因表达增加,造成骨骼肌萎缩,体重下降,此变化与摄食量减少无关。     

低氧影响蛋白质合成和分解的平衡诱导骨骼肌萎缩

暴露在低氧环境下,可能会引起胃肠功能障碍和摄食量下降,打破骨骼肌蛋白质合成和分解平衡,造成骨骼肌萎缩。为探讨低氧环境下骨骼肌的萎缩是低氧环境引起的还是低氧诱发的摄食量减少所致,本研究检测大鼠腓肠肌中低氧时蛋白质合成与分解相关基因的蛋白质表达。将21只雄性SD大鼠,随机分为3组：常氧对照组、低氧组（氧浓度为12.4%,模拟海拔4 000 m高度）和配对组（大鼠的摄食量与低氧组前1 d的摄食量相同）,每组7只,每天记录大鼠体重和摄食量。4周后,HE染色法观察腓肠肌肌纤维形态,Western印迹测试相关蛋白质水平。低氧组和配对组摄食量在低氧干预初期,较常氧对照组有显著性下降(P<0.05),干预后期差异不明显;干预期间,低氧组大鼠体重平均增加量(102.10 g)、体重(341.20 ± 16.75 g)、肌肉总量（226.83 ± 8.33 g）和腓肠肌肌纤维横截面积(12.67 ± 1.83 mm)较常氧对照组（128.00 g;377.50 ± 20.75 g;260.50 ± 9.35 g;15.78 ± 2.38 mm）和配对组（119.40 g;375.86 ± 11.30 g;262.29 ± 7.90 g;15.71 ± 2.82 mm）均显著下降,配对组较常氧对照组无显著性差异;4周干预后,与常氧对照组相比,低氧组大鼠腓肠肌中与低氧相关的HIF1α显著增加(1.42 ± 0.19, P<0.05),Akt和p-Akt/Akt显著降低 (1.44 ± 0.13; 0.47 ± 0.08, P<0.05),配对组上述3种指标相对表达量均无显著性差异;在蛋白质合成方面,低氧组mTOR较常氧对照组显著下降(0.63 ± 0.18, P<0.05),配对组较常氧对照组差异不明显;低氧组腓肠肌中,4EBP1(1.14 ± 0.14)和p70S6K1(1.14 ± 0.11)较配对组显著下降(P<0.05)。在蛋白质分解方面,低氧组p-FoxO1和p-FoxO1/FoxO1比值较常氧对照组显著下降(0.71 ± 0.15; 0.78 ± 0.14, P<0.05);低氧组大鼠腓肠肌中,Atrogin1、MuRF1、Beclin1、LC3Ⅰ及LC3Ⅱ/Ⅰ比值均高于常氧对照组(1.35 ± 0.12; 1.30 ± 0.22; 1.17 ± 0.11; 1.03 ± 0.11; 1.35 ± 0.13, P<0.05);配对组与常氧对照组间无明显差异。低氧环境下骨骼肌中蛋白质合成相关基因表达减少,蛋白质分解相关基因表达增加,造成骨骼肌萎缩,体重下降,此变化与摄食量减少无关。     

高黎贡山赧亢白眉长臂猿食性及日取食量

通过确定白眉长臂猿的取食特征及日取食量,探讨不同个体间和春、秋季间日取食量是否存在差异,分析影响日取食量和取食特征的因素,为白眉长臂猿日能量需求及其营养容纳量研究奠定基础。2007—2008年春、秋季,在高黎贡山赧亢采用焦点动物取样法观察和记录数据。结果发现,白眉长臂猿取食36种食物的40个部位,其果实的食物单元、取食速度、单个鲜重值等取食特征不同于嫩叶。春、秋季白眉长臂猿每日取食果实量所占百分比最高、嫩叶所占百分比次之、茎和花所占百分比最低。春、秋季家群中雌猿的日取食量均高于雄猿。与家群的雌猿相比,独猿(♀)春季的日取食量稍高。在春季,独猿取食果实持续时间约为家群的2倍,而取食嫩叶的则与家群几乎相等。相同个体秋季的日取食量高于春季的,且春、秋季日取食各部位的量所占百分比也不同。分析结果表明,食性及日取食部位的量反映了白眉长臂猿选择食物的最基本策略。雌雄个体大小差异、能量消耗、食物特征、动物生理需求等因素影响白眉长臂猿的日取食量。白眉长臂猿取食持续时间与觅食树的食物资源量相关。     

Nutritional properties of tubers of conventionally bred and transgenic lines of potato resistant to necrotic strain of Potato virus Y (PVYN)

The potential effect of genetic modification on nutritional properties of potatoes transformed to improve resistance to a necrotic strain of Potato virus Y was determined in a rat experiment. Autoclaved tubers from four transgenic lines were included to a diet in the amount of 40% and compared with the conventional cv. Irga. The experiment lasted 3 weeks and special attention was paid to nutritional properties of diets, caecal metabolism and serum indices. Genetic modification of potato had no negative effect on the chemical composition and nutritional properties of tubers, ecosystem of the caecum, activity of serum enzymes and non-specific defence mechanism of the rats. Obtained results indicate that transgenic potato with improved resistance to PVY(N): line R1F (truncated gene coding for PVY(N) polymerase in sense orientation), R2P (truncated gene coding for PVY(N) polymerase in antisense orientation), and NTR1.16 (non-translated regions of PVY(N) genome in sense orientation) are substantial and nutritional equivalence to the non-transgenic cultivar. Tubers of transgenic line NTR2.27 (non-translated regions of PVY(N) genome in antisense orientation) increased the bulk of caecal digesta and the production of SCFA as compared to tubers of the conventional cultivar and the other transgenic clones. Taking into account some deviations, it seems reasonable to undertake a long-term feeding study to confirm the nutritional properties of tubers of transgenic lines.     

Blockade of connexin 43 expression by stable transfection of antisense cDNA in cultured vascular smooth muscle cells

Gap junctional communication is involved in embryogenesis, cell growth control, and coordinated contraction of cardiac myocytes. It has been hypothesized that gap junctions coordinate responses of vascular cells to constrictor or dilator stimulation. Three connexin (Cx) proteins, 37, 40, and 43, are found in the vasculature. Cx43 gap junctions are widely distributed along the vascular tree, although a precise physiologic role in vascular function is unknown because of a lack of specific functional inhibitors and of suitable animal models. To investigate the role of Cx43 in intercellular communication among vascular smooth muscle (VSM) cells, we selectively modified the expression of the Cx43 gene using antisense cDNA stable transfections in culture. Results show that in cells stably transfected with antisense Cx43 cDNA, gene expression of Cx43 could be reduced to 20% of that observed in vector-transfected cells. In spite of the mRNA and protein reduction, the antisense Cx43 cDNA-transfected cells did not show a significant reduction in dye transfer or a difference in cell growth rate as compared with control. These results suggest either that the residual amount of Cx43 protein is sufficient for dye transfer and growth control or that the dye transfer in these cells can be mediated by Cx40 or other connexin proteins. Therefore, more potent approaches, such as dominant negative and gene knockout, are required to fully block gap junctional communication in VSM cells.     

Antisense RNA modulation of alkyl hydroperoxide reductase levels in Helicobacter pylori correlates with organic peroxide toxicity but not infectivity

Much of the gene content of the human gastric pathogen Helicobacter pylori ( approximately 1.7-Mb genome) is considered essential. This view is based on the completeness of metabolic pathways, infrequency of nutritional auxotrophies, and paucity of pathway redundancies typically found in bacteria with larger genomes. Thus, genetic analysis of gene function is often hampered by lethality. In the absence of controllable promoters, often used to titrate gene function, we investigated the feasibility of an antisense RNA interference strategy. To test the antisense approach, we targeted alkyl hydroperoxide reductase (AhpC), one of the most abundant proteins expressed by H. pylori and one whose function is essential for both in vitro growth and gastric colonization. Here, we show that antisense ahpC (as-ahpC) RNA expression from shuttle vector pDH37::as-ahpC achieved an approximately 72% knockdown of AhpC protein levels, which correlated with increased susceptibilities to hydrogen peroxide, cumene, and tert-butyl hydroperoxides but not with growth efficiency. Compensatory increases in catalase levels were not observed in the knockdowns. Expression of single-copy antisense constructs (expressed under the urease promoter and containing an fd phage terminator) from the rdxA locus of mouse-colonizing strain X47 achieved a 32% knockdown of AhpC protein levels (relative to wild-type X47 levels), which correlated with increased susceptibility to organic peroxides but not with mouse colonization efficiency. Our studies indicate that high levels of AhpC are not required for in vitro growth or for primary gastric colonization. Perhaps AhpC, like catalase, assumes a greater role in combating exogenous peroxides arising from lifelong chronic inflammation. These studies also demonstrate the utility of antisense RNA interference in the evaluation of gene function in H. pylori.     

反义adipophilin寡核苷酸降低ACAT活性(英)

已经发现,随着细胞内胆固醇酯的积聚,adipophilin的表达明显增加.在此基础上,为了寻找adipophilin在细胞内胆固醇代谢的作用点.小鼠腹腔巨噬细胞与80 mg/L OxLDL或80 mg/L OxLDL加1 mmol/L adipophilin反义寡核苷酸共孵育,在不同的时间点取样,使用免疫荧光染色,流式细胞仪分析和细胞内胆固醇测定.结果显示,72 h后,反义寡核苷酸处理组细胞内胆固醇酯显著下降到（19.9±1.9）mg/g,油红O染色显示,该组细胞质内红色脂滴明显减少.96 h的观察期间内,两组细胞adipophilin蛋白的表达量都增加.从12 h开始,未用反义寡核苷酸处理组adipophilin蛋白的表达量开始大于反义寡核苷酸处理组.在96 h时间点,统计学处理,未用反义寡核苷酸处理组明显高于反义寡核苷酸处理组,差别有显著性.使用Ox-r［CL-³H］LDL观察两组细胞摄入OxLDL的量,实验发现,两组细胞摄入Ox-r［CL-³H］LDL的量逐渐增加.但是,反义寡核苷酸处理组摄入Ox-r［CL-³H］LDL的量从24 h后明显低于未用反义寡核苷酸处理组,在48 h和96 h时间点,统计学处理,两组差别有显著性.观察ACAT的活性发现,ACAT相对活性从6 h到48 h表现为增加,但从48 h到96 h,ACAT相对活性趋于稳定.在48 h时间点,两组比较差别有显著性,反义寡核苷酸处理组的活性明显低于未用反义寡核苷酸处理组.相关分析发现,ACAT的活性与adipophilin蛋白的表达量有一定的联系,但不是直线相关.结果表明,adipophilin表达的高低与细胞摄入外源性脂蛋白,与ACAT的活性有一定的联系.提示adipophilin与脂滴的代谢功能密切相关,且ACAT有可能是其潜在的位点.     

Genetic polymorphisms of Pe and Po salivary proteins with probable linkage of their genes to the salivary protein gene complex (SPC)

Two new genetic polymorphisms (Pe and Po) are found in human parotid saliva. Each polymorphism is determined by the autosomal inheritance of one expressed (dominant) and one unexpressed (recessive) allele. Autosomal inheritance is supported by studies of 63 families including 264 children for Pe and 57 families including 242 children for Po. For randomly collected salivas, gene frequencies in 317 whites are Pe+ = 0.76 and Pe- = 0.24; in 408 whites, Po+ = 0.75 and Po- = 0.25; in 51 blacks, Pe+ = 0.76 and Pe- = 0.24; and in 59 blacks, Po+ = 0.77 and Po- = 0.23. Both Pe and Po proteins react immunologically with polyclonal antisera prepared to proline-rich proteins PRPs. The Pe protein has an isoelectric point of approximately pH 6.1-6.3, and the Po protein has an isoelectric point greater than pH 8.0. In randomly collected salivas, the Pe and Po proteins are associated with other known salivary PRPs. The Pe protein is most strongly associated with the CON 1 and Ps proteins, is less strongly associated with the Pr and Pa proteins, and is not significantly associated with the PmF, PmS, PIF, Db, Con 2, or Gl proteins. If it is assumed that the strength of these associations (presumed linkage disequilibrium) may be related in part to map distance, then these data roughly fit the linear order of PRP genes as previously determined from recombination data derived from family linkage studies. The Po protein is associated with the PmS protein. There is evidence for probable linkage of Pe and Po to the SPC (salivary protein gene complex): Pe to Pa (nine families, lod score at theta = 0 is 2.67) and Po to CON 2 (three families, lod score at theta = 0 is 2.35).     

Activation of two forms of locomotion by a previously identified trigger interneuron for swimming in the medicinal leech

Higher-order projection interneurons that function in more than one behavior have been identified in a number of preparations. In this study, we document that stimulation of cell Tr1, a previously identified trigger interneuron for swimming in the medicinal leech, can also elicit the motor program for crawling in isolated nerve cords. We also show that motor choice is independent of the firing frequency of Tr1 and amount of spiking activity recorded extracellularly at three locations along the ventral nerve cord prior to Tr1 stimulation. On the other hand, during Tr1 stimulation there is a significant difference in the amount of activity elicited in the ventral nerve cord that correlates with the motor program activated. On average, Tr1 stimulation trials that lead to crawling elicit greater amounts of activity than in trials that lead to swimming.