Smad泛素化调节因子2siRNA的筛选及转染原代肾小管上皮细胞条件的优化简

试验旨在优化Smad泛素化调节因子2（Smurf2） siRNA最佳转染条件,筛选最佳siRNA干扰片段,进而实现对Smurf2基因的瞬时沉默,为研究Smurf2基因在马兜铃酸肾病（aristolochic acid nephrohathy,AAN）中的作用奠定基础。本研究通过培养小鼠原代肾小管上皮细胞（renal tubular epithelial cells,RTECs）,并以脂质体Lipofectamine^TM 2000为转染介质,将Smurf2 siRNA转染入RTECs;通过观察绿色荧光的表达量及Real-time PCR反应优化转染条件,转染后Real-time PCR检测mRNA表达抑制率。CCK-8法检测Smurf2 siRNA复合物对RTECs活性的影响;同时,用Real-time PCR和Western blotting检测不同位点Smurf2 siRNA对Smurf2 表达的影响。结果显示,当Lipofectamine ^TM 2000与siRNA比例为1.5 μL∶30 pmol时,转染效率最高,为70%～80%;Smurf2-619 siRNA干扰效果最明显;与正常组相比,转染siRNA组的Smurf2蛋白表达水平显著下降（P<0.05）。最终确定了Smurf2 siRNA最佳转染条件,其中Smurf2-619 siRNA对Smurf2 表达的抑制率最高。     

BHBA对HT22细胞中ACE表达的影响

生酮饮食(KD)已被证实具有神经保护作用,但其机制尚不明确。本试验以小鼠海马神经元HT22为细胞模型,利用2 mmol/Lβ-羟基丁酸酯(BHBA)对HT22细胞进行处理,通过荧光定量PCR,免疫蛋白印迹和信号通路阻断试验,观察BHBA对HT22细胞ACE表达的影响,并探讨参与调控的信号通路。结果表明,2 mmol/L BHBA可通过激活HT22细胞内GPR109A受体和ERK1/2信号通路进而增加HT22细胞中ACE的表达。     

Studies on the mechanism of vaccinal immunity to Marek''s disease

Current knowledge of the nature of the antigens and of the host immune responses in vaccinal immunity to Marek's disease is reviewed. It is suggested that a two-step mechanism of resistance operates. The first step involves humoral and cell-mediated responses directed against viral antigens; the second step occurs after challenge with Marek's disease virus and consists of cellmediated responses directed against tumour cells.     

口服ND弱毒苗对鸡十二指肠中2种神经多肽阳性细胞的影响

应用新城疫弱毒苗经消化道黏膜2次免疫鸡后,采用免疫组化的方法检测鸡十二指肠中生长抑素(Somatostatin,SS)阳性细胞和P物质(Substance P,SP)阳性细胞的分布及其数量变化。结果表明,动物首免后第3周,新城疫免疫组SS阳性细胞数量极显著增加(P<0.01),而SP阳性细胞数量显著减少(P<0.05);首免后第5周和第7周,新城疫免疫组和对照组中的2种阳性细胞数差异不显著。     

凋亡素基因真核表达载体的构建及在人肿瘤细胞中的表达

为探讨凋亡素对肿瘤的基因治疗效果,构建了凋亡素基因的真核表达载体。将凋亡素基因重组入 ｐｃ Ｄ Ｎ Ａ３ 载体,并通过脂质体介导凋亡素在人喉癌、人肺癌细胞系中表达;通过 Ｒ Ｔ Ｐ Ｃ Ｒ 在转染细胞中检测到了凋亡素的 ｍ Ｒ Ｎ Ａ,这为应用凋亡素进行肿瘤的基因治疗奠定了基础。     

利用柞蚕核型多角体病毒表达载体系统在柞蚕蛹中表达增强型绿色荧光蛋白

为了探讨外源蛋白在柞蚕蛹-柞蚕核型多角体病毒(ApNPV)表达系统的表达水平和稳定性,将增强型绿色荧光蛋白(EGFP)基因克隆到柞蚕核型多角体病毒转移表达载体pApM748BE中,获得重组质粒DNA,与ApNPV DNA共转染樗蚕(Phi-losamia cynthiam)培养细胞Pc-01后,用末端稀释法筛选获得重组病毒ApNPV-Δph/egfp+。将该重组病毒感染柞蚕蛹,采用SDS-PAGE和Western blot方法检测EGFP在柞蚕蛹中的表达,结果显示:在柞蚕蛹感染重组病毒ApNPV-Δph/egfp+后6 d,EG-FP就有明显的表达;感染后12～18 d,蛹体液中的EGFP含量保持较高水平,以EGFP标准样品定量分析EGFP在柞蚕蛹体液中的表达水平高于1 mg/mL;感染后30～39 d仍可以检测到蛹体液中EGFP的表达,而且蛋白稳定。研究结果表明,利用柞蚕核型多角体病毒作表达载体,可在柞蚕蛹中高效稳定地表达EGFP,并且EGFP在雌雄蛹间的表达水平无明显差异。     

Mastitis caused by Mycobacterium kansasii infection in a dog

A 2‐year, 7‐month‐old female Chihuahua was admitted for a mammary mass measuring one cm in diameter. The dog had a history of demodicosis for 4 months and showed signs of pseudopregnancy at the time of the visit. Cytologic examination of an aspirate of the mass revealed a large number of macrophages containing nonstaining bacterial rods, which were acid‐fast in a Ziehl–Neelsen stain, suggesting mycobacterial infection. Histologic examination of the mass revealed a pyogranulomatous mastitis characterized by an infiltration with macrophages containing acid‐fast bacteria. Mycobacterium kansasii was subsequently cultured and identified by PCR. Surgical excision of the mass resulted in the growth of other dermal masses, but antimycobacterial treatment with rifampin and clarithromycin resolved these masses within 1 month. Three months after discontinuation of the treatment, similar organisms were found in aspirates of the enlarged bilateral inguinal lymph nodes by cytologic examination. Despite antimycobacterial treatment for another 4 months, there was no improvement and demodicosis also recurred. The dog eventually died of lymphoma 5 months after the relapse of mycobacterial infection. Although M kansasii is considered an important pathogen for pulmonary and cutaneous disease in people, there is only one report in a dog with an infection in a pleural effusion. As both adult‐onset demodicosis in dogs as well as mycobacterial infection in people have been associated with T‐lymphocyte deficiency, the M kansasii infection in this dog may have been associated with a condition of immune compromise.     

CnAβ基因对大鼠RBL-2H3细胞脱颗粒的影响

过敏是人和养殖动物常见的疾病,而且发生的频率呈现逐渐增加的趋势.为了深入了解过敏反应的机制,本研究利用CRISPR/Cas9基因编辑技术,构建钙调磷酸酶A beta(calcineurin A beta,CnAβ)基因敲除的大鼠RBL-2H3细胞株,并探讨CnAβ基因对RBL-2H3细胞生长及脱颗粒的影响.选取大鼠Cn...     

Identification of CD3+ T lymphocytes in the green turtle Chelonia mydas

To understand the role of the immune system with respect to disease in reptiles, there is the need to develop tools to assess the host's immune response. An important tool is the development of molecular markers to identify immune cells, and these are limited for reptiles. We developed a technique for the cryopreservation of peripheral blood mononuclear cells and showed that a commercially available anti-CD3 epsilon chain antibody detects a subpopulation of CD3 positive peripheral blood lymphocytes in the marine turtle Chelonia mydas. In the thymus and in skin inoculated with phytohemagglutinin, the same antibody showed the classical staining pattern observed in mammals and birds. For Western blot, the anti-CD3 antibodies identified a 17.6 kDa band in membrane proteins of peripheral blood mononuclear cell compatible in weight to previously described CD3 molecules. This is the first demostration of CD3+ cells in reptiles using specific antibodies.