Ras-GRF1蛋白对小鼠学习和记忆能力的影响

通过避暗穿梭试验、跳台试验、水迷宫试验和自主活动试验观察Ras-GRF10/0鼠和野生鼠2种小鼠行为学变化,来研究Ras-GRF1蛋白对学习和记忆能力的影响。结果显示,在避暗试验中Ras-GRF10/0鼠被电击的次数多于野生鼠,且在第1天有显著差异(P0.05);在跳台试验中,野生鼠表现出的学习记忆能力也强于基因敲除鼠;在水迷宫定位航行试验中,基因敲除鼠的潜伏期均长于野生鼠,且在第2天有显著差异(P0.05),第3、4、5、6、7天都有极显著的差异(P0.01);在空间探索试验中,基因敲除鼠和野生鼠第1次穿越平台的时间和穿越平台的次数均无显著差异;在自主活动试验中,基因敲除鼠和野生鼠的活动次数没有显著性的差异,但是野生鼠的活动次数都多于敲除鼠。结果表明,野生鼠的学习和记忆能力均强于Ras-GRF10/0鼠,且野生鼠的中枢神经兴奋性高于基因敲除鼠,反映出Ras-GRF1蛋白缺失,对小鼠学习和记忆能力有一定程度的损伤。     

SPRN转基因小鼠脏器质量和动物行为学的比较分析

采用统计学计算Tg(Mosprn-B)/C57BL/6转基因小鼠与C57BL/6野生型小鼠雌雄两性的主要脏器质量与系数,并利用水迷宫观察游泳路径及时间以探讨SPRN基因对小鼠生理结构、生长发育等影响。结果表明,相同年龄的Tg(Mosprn-B)/C57BL/6转基因小鼠,雄性的体质量及心、肺、肝、肾质量显著大于雌性(P<0.01),脑质量差异比较显著(P<0.05);比较脏器系数,脑和肝的差异比较显著(P<0.05)。与C57BL/6野生型小鼠相比较,Tg(Mosprn-B)/C57BL/6转基因小鼠脑更重(P<0.01);比较脏器系数发现,脑差异极显著(P<0.01),肺、肾、脾差异较显著(P<0.05)。水迷宫定位航行结果显示,Tg(Mosprn-B)/C57BL/6转基因小鼠的逃避潜伏期小于野生型小鼠,第4天的潜伏期具有显著差异性(P<0.05);在空间搜索试验中,Tg(Mosprn-B)/C57BL/6转基因小鼠第1次穿越平台时间小于C57BL/6野生型小鼠,穿越平台的次数大于C57BL/6野生型小鼠。     

NLRP3在介导小鼠皮肤伤口修复过程中的作用

本试验进行了小鼠肌成纤维细胞及骨髓源巨噬细胞的体外培养,随后使用野生型C57BL/6J小鼠或NLRP3基因敲除鼠耳皮肤肌成纤维细胞培养上清液(myofibroblast conditioned medium, MFbCM)处理M1和M2型巨噬细胞,分别检测巨噬细胞组织修复相关极化调节分子精氨酸酶(Arginase)的活性、NO的表达量、IL-10的分泌量以及几丁质酶3样分子(Ym1)的表达。同时,通过野生型C57BL/6J小鼠和NLRP3基因敲除小鼠背部创伤模型的建立,分析皮肤伤口愈合情况。通过免疫荧光检测小鼠皮肤创面组织中Ym1和Arginase表达量。结果显示,在MFbCM诱导的条件下,骨髓源巨噬细胞的精氨酸酶活性、NO的表达、IL-10的分泌和Ym1的表达均依赖于小鼠肌成纤维细胞NLRP3的表达;当肌成纤维细胞NLRP3表达缺失后,巨噬细胞上述组织修复相关极化调节因子均出现下调现象,进而可能对伤口愈合产生不利影响。此外,与之相一致的是,NLRP3基因敲除小鼠的伤口愈合速度相比于野生型C57BL/6J小鼠显著缓慢。在创面组织中,NLRP3缺失后Ym1和Arginase表达显著下调。...     

水通道蛋白AQP3缺失对小鼠坐骨神经损伤修复的影响

为探明水通道蛋白AQP3缺失对小鼠坐骨神经损伤修复能力的影响,本文通过建立小鼠坐骨神经损伤修复模型,分别比较了野生型(AQP3+/+)和基因敲除(AQP3-/-)小鼠的坐骨神经功能指数(SFI)、神经传导速率及雪旺细胞的迁移能力。结果显示,同野生型小鼠相比,AQP3基因敲除小鼠的SFI在损伤后7 d和14 d时呈现显示性差异,神经传导速率显著降低。细胞穿孔试验表明,AQP3基因敲除小鼠同野生型小鼠相比雪旺细胞迁移能力也显著降低。研究结果表明,AQP3的缺失可导致小鼠雪旺细胞的迁移能力降低,从而影响了小鼠坐骨神经损伤后的修复能力。     

C57BL/6NcGAS基因敲除鼠的构建及对猪伪狂犬病病毒增殖的影响

胞质DNA感受器环鸟苷酸-腺苷酸合成酶(cyclic GMP-AMP synthase, cGAS)能够识别胞质中的双链DNA(double stranded DNA,dsDNA),从而激活Ⅰ型干扰素信号通路抵御外界病原微生物的感染。为研究cGAS基因对猪伪狂犬病病毒(PRV)增殖的影响，本试验首先利用CRISPR/Cas9基因编辑技术构建cGAS基因敲除小鼠，通过H&E染色检测C57BL/6N小鼠和C57BL/6N-cGAS^-/-小鼠肺脏和脑组织形态差异及感染PRV后肺脏组织中炎性浸润的影响；Q-PCR检测C57BL/6N小鼠和C57BL/6N-cGAS^-/-小鼠感染PRV后对PRV-gB、PRV-TK、IFN-β、ISG15和ISG20基因mRNA表达水平的差异；Western blot及免疫组化检测C57BL/6N小鼠和C57BL/6N-cGAS^-/-小鼠感染PRV后PRV-gB和PRV-gE蛋白表达水平；最后利用病毒PFU法检测cGAS敲除鼠对PRV子代病毒滴度的影响。Western blot检测结果显示...     

心房钠尿肽基因敲除小鼠的鉴定

研究心房钠尿肽基因敲除小鼠(ANP-/-小鼠)的生物学及病理学特性。ANP-/-小鼠与C57BL/6小鼠饲养于SPF级环境中,观察小鼠的繁育状况及其生物学特性,选取合适时间处死小鼠,通过HE染色观察脏器形态及组织结构。ANP-/-小鼠的繁殖周期比C57BL/6长,产仔率较低;ANP-/-小鼠肝脏、肺脏、脾脏较C57BL/6小鼠的有明显炎症反应,但是C57BL/6小鼠未见明显的病理改变。提示ANP基因的缺失可能与炎症发生有关。说明ANP-/-小鼠繁殖较慢、产仔率低,且正常繁殖的普通小鼠不会引起器官的病理学改变。但作为一种在心血管、代谢疾病上的动物模型,并与炎症甚至肿瘤有可能的未知联系的转基因动物,其很有研究和探索价值。     

A2a基因敲除小鼠生物学特性的初步研究

为了解A2a基因敲除小鼠的生物学物性,给研究心血管疾病及神经疾病提供部分基础资料,试验采用全自动生化分析仪及血细胞分析仪分别对60日龄C57BL/6、A2a雌雄小鼠的常用血液学指标及生化指标值进行检测,并测定了各小鼠的脏器。结果:C57BL/6和A2a小鼠血液的血小板计数、红细胞分布宽度差异显著(P0.05)。C57BL/6和A2a小鼠的丙氨酸转氨酶、肌酸激酶差异显著(P0.05),乳酸脱氢酶差异极显著(P0.01),脏器系数无显著差异(P0.05)。     

SPF级BALB/cC57BL/6小鼠繁殖性能及生长发育的比较研究

为提高SPF级实验动物的质量,选择10周龄BALB/c C57BL/6小鼠各50只（雌雄各半）,采取雌雄1？1近亲交配繁殖,并统计其生长发育和繁殖性能指标。BALB/c小鼠1～3周龄的平均窝重及离乳后的体重增长明显大于C57BL/6小鼠（p〈0.01）。第1胎交配分娩间隔,第1和第3胎离乳育成率品系间差异均有显著性（P〈0.05及P〈0.01）。BALB/c小鼠的体格比C57BL/6小鼠大,带乳能力比C57BL/6小鼠强但产仔能力比C57BL/6小鼠弱。     

TLR2~(+/+)与TLR2~(-/-)小鼠乳腺灌注乳房链球菌后血浆中抗氧化能力及细胞因子水平的比较

为探讨TLR2在乳房链球菌(Streptococcus uberis)感染小鼠乳腺后对血浆中抗氧化能力及细胞因子产生的影响,本研究以B6小鼠TLR2~(+/+)(野生型)和TLR2~(-/-)(TLR2敲除)B6小鼠为研究对象,在小鼠分娩72 h时,乳腺灌注乳房链球菌(设生理盐水灌注对照),24 h后采集血液并制备血浆。实验结果表明:TLR2~(+/+)小鼠血浆中总抗氧化能力(T-AOC)显著高于TLR2~(-/-)小鼠,乳房灌注乳房链球菌后TLR2~(+/+)及TLR2~(-/-)小鼠血浆中T-AOC水平均显著降低,同对照组相比差异显著(P0.05),且TLR2~(+/+)中也显著高于TLR2~(-/-)型;超氧化物歧化酶(SOD)活力在TLR2~(-/-)小鼠中无变化,而在野生小鼠中显著降低,丙二醛(MDA)含量在两种小鼠中均显著升高,野生型小鼠中含量低于敲除小鼠(P0.05);TLR2~(-/-)小鼠在乳房链球菌感染后血浆中肿瘤坏死因子-α(TNF-α)分泌显著增加(P0.05);野生型小鼠在感染后白细胞介素-6(IL-6)分泌显著增加(P0.05)。研究表明,TLR2参与了乳房链球菌诱发乳腺局部炎症后的机体整体水平的抗氧化能力及细胞因子分泌的变化,但不是唯一因素。     

短链菊粉与高脂饲粮对不同基因型小鼠血清葡萄糖含量及盲肠微生物区系关联性的影响

本试验旨在研究短链菊粉与高脂饲粮对不同基因型小鼠血清葡萄糖含量及盲肠微生物区系关联性的影响。将8周龄40只雄性野生型C57BL/6J小鼠与40只雄性瘦素(Leptin)基因敲除小鼠分为8组，每组10只，单笼饲养。1～4组为野生型小鼠的不同处理组：野生型对照组(CT组)、野生型+10%短链菊粉组(CI组)、野生型+高脂组(CH组)、野生型+高脂+10%短链菊粉组(CHI组);5～8组为Leptin基因敲除小鼠的不同处理组：Leptin基因敲除对照组(OT组)、Leptin基因敲除+10%短链菊粉组(OI组)、Leptin基因敲除+高脂组(OH组)、Leptin基因敲除+高脂+10%短链菊粉组(OHI组)。试验期为8周，期间监测采食量与体增重。于第8周腹腔注射葡萄糖，注射后15、30、60和120 min测定血清葡萄糖含量。之后脱臼处死，测定体重与体长，取肝脏、生殖腺脂肪称重。取盲肠段及内容物测定微生物群落组成。结果表明：1)第12天CHI组体重显著低于CI组(P<0.05),第42～54天CH组体重显著低于CI组(P<0.05),Leptin基因敲除组间体重无显著差异(P&g...     

Alymphoplasia mice are resistant to prion infection via oral route

The major cause of infection in animal prion diseases is thought to be consumption of prion-contaminated stuff. There is evidence that the enteric nerve system (ENS) and gut-associated lymphoid tissues (GATL) are involved in the establishment of prion infection through alimentary tract. To elucidate the initial entry port for prion, we inoculated prion to alymphoplasia (aly) mice showing a deficiency in systemic lymph nodes and Peyer's patches. The aly/aly mice were susceptible to prion infection by intra-cranial inoculation and there were no differences in incubation periods between aly/aly mice and wild-type C57BL/6J mice. Incubation periods in aly/aly mice were about 20 days longer than those in C57BL/6J mice with the intra-peritoneal inoculation. The aly/aly mice were completely resistant to prion infection by per os administration, while C57BL/6J mice were sensitive as they entered the terminal stage of disease around 300 days post inoculation. PrP(Sc) were detected in the intestine and spleen of C57BL/6J mice inoculated with prion intraperitoneally or orally; however PrP(Sc) was not detected in the spleen and intestine of aly/aly mice. Prion infectivity was detected in the intestines and spleens of prion-inoculated C57BL/6J mice, even after the early stages of exposure, while no infectivity was detected in these tissues of prion-inoculated aly/aly mice. No apparent differences were observed in the organization of the enteric nerve system between wild-type and aly/aly mice. These results indicate that GALT rather than ENS acts as the primary entry port for prion after oral exposure.     

Emerging therapeutic agents for transmissible spongiform encephalopathies: a review

Transmissible spongiform encephalopathies (TSEs) are a group of fatal neurodegenerative disorders associated with misfolding of prion protein, from PrPC to PrPSc. Different types of experimental studies have resulted in a better understanding of the pathogenesis of the prion diseases. Genetic and molecular properties of PrP isoforms have been explained but the conformational conversion of the PrPC isoform to the PrPSc isoform has not yet been entirely elucidated. However, a number of possible therapeutic agents have been tried and some have proven to be effective against TSEs but most have limitations in terms of toxicity and pharmacokinetics. Congo red (CR), anthracyclines, and polyanionic dextran sulfate have limited ability to cross the blood-brain barrier and may be toxic. The efficacy of polyene antibiotics seems to be restricted to certain scrapie strains. Tetrapyrroles and tetracyclines with low toxicities and favorable pharmacokinetics could be useful in preventing PrPSc accumulation. Compounds like branched polyamines, Cp-60, analogs of CR, quinacrine and chlorpromazine, beta-sheet breaker peptides and inhibitory peptides, active immunization using recombinant PrP and passive immunization with anti-PrP antibodies, have potential use as therapeutic agents but would need further research and clinical trials.     

牛源金黄色葡萄球菌黏附素纤连结合蛋白A分泌型DNA疫苗诱导小鼠免疫效应的研究

采用PCR方法对编码金黄色葡萄球菌黏附素纤连结合蛋白A(FnbpA)的A区基因片段进行了特异性扩增,构建了真核表达载体pVAX-SFn,转染BHK-21细胞后经ELISA可检测出分泌表达的FnbpA蛋白。将真核重组表达质粒肌肉注射C57BL/6小鼠,免疫后检测小鼠血清抗体效价、淋巴细胞增殖及对试验小鼠攻毒试验。结果表明,该重组表达载体诱导细胞和体液免疫应答的强度均明显超过对照组。对小鼠的攻毒试验结果提示该重组DNA经肌肉注射途径接种可对小鼠产生免疫保护。本试验的结果对该DNA疫苗今后在实际中的应用奠定了良好的试验基础。     

SPRN0/0型小鼠与C57BL/6野生型小鼠小脑中microRNA的差异表达

The purpose of this study was to identify different expression of microRNAs between cerebellum of SPRN^0/0 mice and C57BL/6 wild mice, and investigate relative expression results of microRNA in cerebrum of SPRN^0/0 mice by Real-time PCR technology. The results showed that compared with cerebellum of C57BL/6 wild mice, eight microRNAs were decreased significantly:miR-135a, miR-24-2*, miR-146a, miR-7a, miR-380, miR-448, miR-128 and miR-152. miR-24-2* fold change to more than 10, five microRNAs including miR-448,miR-146a,miR-128,miR-380 and miR-152 fold change to more than 2, miR-135a and miR-7a fold change to more than 1. Thus, we concluded that these changes of microRNAs expression might be involved in Shadoo protein function and the establishment of target genes of microRNAs might provide new ideas for researching Shadoo protein function.     

低温环境下禁食对小鼠自发运动及腓肠肌中PGC-lα、ERRα表达的影响

应用旷场试验测定不同时长4℃冷暴露下采食和禁食C57BL/6小鼠的自发运动,并结合骨骼肌中PGC-lα与ERRα表达水平,探讨低温环境下禁食对C57BL/6小鼠自发运动的影响。结果显示,低温环境下禁食对C57BL/6小鼠的自发运动并无显著性影响,但焦虑情绪12 h差异显著(P<0.05),探索行为8 h和12 h差异极显著(P<0.01);腓肠肌中PGC-1α表达在4,8 h时差异极显著(P<0.01),12 h差异显著(P<0.05);ERRα表达在4,6 h时差异极显著(P<0.01)。结果表明,低温环境下禁食对C57BL/6小鼠自发运动未出现明显影响,而对小鼠焦虑情绪、探索行为和腓肠肌中PGC-lα、ERRα表达存在不同程度的影响,相比于禁食而言,冷刺激对小鼠机体的影响更大。     

Immunisation strategies against prion diseases: prime-boost immunisation with a PrP DNA vaccine containing foreign helper T-cell epitopes does not prevent mouse scrapie

Vaccination against prion diseases constitutes a promising approach for the treatment and prevention of the disease. Passive immunisation with antibodies binding to the cellular prion protein (PrP(C)) can protect against prion disease. However, immunotherapeutic strategies with active immunisation are limited due to the immune tolerance against the self-antigen. In order to develop an anti-prion vaccine, we designed a novel DNA fusion vaccine composed of mouse PrP and immune stimulatory helper T-cell epitopes of the tetanus toxin that have previously been reported to break tolerance to other self-antigens. This approach provoked a strong PrP(C)-specific humoral and cellular immune response in PrP null mice, but only low antibody titres were found in vaccinated wild-type mice. Furthermore, prime-boost immunisation with the DNA vaccine and recombinant PrP protein increased antibody titres in PrP null mice, but failed to protect wild-type mice from mouse scrapie.     

Establishment and characterization of the MSKR inbred strain originated from Japanese wild mice (Mus musculus molossinus)

A new inbred strain, MSKR, originated from Japanese wild mice was established in April, 1998. The MSKR mice were 60% of the C57BL/6N inbred mice in the 60-day body weight. Tail length/head-body length and hind-foot length/head-body length of the MSKR mice were significantly smaller than those of the C57BL/6N mice (0.896 vs 1.061, 0.189 vs 0.204), but ear length/head-body length of the MSKR mice was significantly larger than that of the C57BL/6N mice (0.143 vs 0.137). The age of the first parturition and size of the first litter were 63.20 +/- 2.71 days and 6.20 +/- 0.37, respectively, at the 20th and 22nd inbreeding generations. Genetic characterization of the MSKR strain was performed using 34 microsatellite markers, 29 biochemical markers, 9 immunogenetic markers, 3 coat color markers, and mitochondrial DNA RFLP-haplotypes. The result indicated that this newly established inbred strain has some different gene constitution from already known molossinus and common laboratory strains.     

Susceptibility of transgenic mice expressing chimeric sheep,bovine and human PrP genes to sheep scrapie

The use of Transgenic (Tg) mice expressing chimeric sheep/mouse (Sh/Mo) prion protein (PrP) and chimeric bovine/mouse (Bo/Mo) PrP genes was evaluated as a sheep scrapie model. We also investigated the potential for the transmission of sheep scrapie to a human/mouse (Hu/Mo) PrP Tg mouse line. The Sh/Mo PrP and Bo/Mo PrP Tg Prnp(+/+) or Prnp(0/0) mouse lines were inoculated intracerebrally with brain homogenates from three sheep with natural scrapie (KU, Y5 or S2). Incubation periods were slightly shorter in Sh/Mo PrP Tg Prnp(+/+), than in non-Tg mice inoculated with KU brain homogenate. In contrast, the incubation period was significantly prolonged (p<0.05) in Bo/Mo PrP Tg Prnp(+/+) mice inoculated with KU brain homogenate. The incubation period was significantly longer in all Tg Prnp(+/+) and Prnp(0/0), than in non-Tg mice (p<0.01) inoculated withY5 brain homogenate. None of the Tg Prnp(0/0) mice inoculated with S2 brain homogenate developed clinical signs and PrP(Sc) was undetectable in their brains. These results suggested that expression of the Sh/Mo PrP or Bo/Mo PrP transgenes does not confer susceptibility to sheep prions upon mice, and thus none of the Tg mouse lines could be a suitable model of sheep scrapie. Hu/Mo PrP Tg Prnp(0/0) mice inoculated with natural and experimental scrapie or mouse prions did not develop clinical signs of scrapie and PrP(Sc) was undetectable. These results suggested that neither sheep nor mouse strains of scrapie are highly transmissible to humans.