线虫fat-1基因密码子优化设计

本研究旨在优化线虫fat-1基因,使其在牛肌肉组织中高效表达,为进一步生产fat-1转基因肉牛提供目的基因。试验在不改变fat-1基因编码氨基酸序列的基础上,参考了11个牛肌肉蛋白相关基因的密码子使用频率,对fat-1基因进行了密码子优化。结果发现,共改变了65个碱基,涉及65个密码子、7个氨基酸,约占fat-1基因总碱基数的5.38%,G+C含量从45.08%上升到50.04%,且分布均匀利于基因的表达。结构预测结果显示,该优化后的基因能够在牛肌肉中高效表达。     

蒲公英内生真菌PG23抗癌活性的初步研究

研究药用植物蒲公英内生真菌PG23多糖对肺癌A549细胞的抑制作用。蒲公英内生真菌PG23通过培养,乙醇沉淀获得粗多糖、苯酚-硫酸法测定多糖含量,采用MTT法测定PG23多糖对肺癌A549细胞的抑制率,并计算半抑制浓度(IC50)。结果表明,PG23发酵液中多糖得率为2.754%,多糖含量为26.82%;对A549肺癌细胞的抑制率以第1天为最高;抑制率不与多糖浓度成比例,最佳作用浓度为14.42μg/mL,抑制率为87.63%(P<0.01),IC50为1.073mg/mL(24h)。PG23发酵液多糖对肺癌A549细胞具有较强的抑制能力。     

Cytologically atypical anal sac adenocarcinoma in a dog

A 10-year-old intact female Shetland Sheepdog with tenesmus had a subcutaneous mass at the left ventral aspect of the anus. On cytologic examination, 2 types of cells were observed. Most of the cells were oval to polygonal and had elliptical or elongate nuclei and a moderate amount of pale to basophilic cytoplasm. The remaining cells had round to oval nuclei and pale to basophilic cytoplasm. Cells of both types were loosely adhered to each other and were arranged in rosette-like structures. Both neoplastic cell types had fine homogenous chromatin and either a small indistinct nucleolus or no visible nucleolus. Mild anisokaryosis and anisocytosis were observed. Histologically, the mass consists of glandular structures formed by cuboidal cells admixed with bundles of spindle cells. Eosinophilic PAS- and Alcian blue-positive secretory material was found in the center of some glandular structures. Both neoplastic cell types had positive staining with paradoxical concanavalin A and expressed cytokeratin, but not vimentin, S-100, α-smooth muscle actin, or desmin. Based on location and histologic and immunohistochemical features, the final diagnosis was adenocarcinoma of the apocrine gland of the anal sac, which should be included as a cytologic differential diagnosis when spindle cells and typical epithelial cells are observed in masses in the region of the anal sac of dogs.     

OPG/RANKL/RANK系统的生理功能

骨是一个代谢活跃的器官,在整个生命周期中都在进行不断的更新与重建。在骨形态发生和重建的过程中主要受到细胞因子和激素的调节,同时骨代谢也受到OPG/RANKL/RANK系统的调控。论文主要对骨骼生理结构与骨组织细胞之间的关系,骨代谢和骨的形成过程,OPG/RANKL/RANK系统的生理功能,OPG/RANKL/RANK系统的影响因子以及骨代谢紊乱所引起的疾病几个方面作一综述。     

猪SENP1基因克隆、生物信息学分析与表达研究

试验旨在对猪SUMO特异性蛋白酶1(sentrin-specific protease 1,SENP1)基因CDS区进行克隆和生物信息学分析,并探讨SENP1基因在乙脑病毒(Japanese encephalitis virus,JEV)感染PK15细胞后的表达情况。根据GenBank中公布的猪SENP1基因预测序列(登录号:XM_013997974.2)设计特异性引物,通过RT-PCR和T/A克隆技术对猪SENP1基因CDS区序列进行克隆测序,并进行生物信息学分析,利用实时荧光定量PCR分析猪SENP1基因在JEV感染PK15细胞不同时间点(0、12、24、36、48 h)的表达情况。结果显示,猪SENP1基因CDS序列全长2 034 bp,共编码677个氨基酸。序列比对和系统进化树分析结果表明,猪SENP1蛋白序列和山羊、犬、人、牛的相似性分别为94.8%、94.2%、93.9%和93.8%,说明在这些物种中SENP1的保守性较强;猪与犬的SENP1分子亲缘关系较近。生物信息学分析结果显示,猪SENP1蛋白相对分子质量为76 825.76,等电点为8.53,体外半衰期为30 h,不稳定系数为53.59,总平均亲水指数为－0.622。SENP1蛋白不含信号肽序列,无跨膜结构域。二级结构分析结果显示,SENP1蛋白中α-螺旋、无规则卷曲、延伸链和β-转角分别占31.31%、46.68%、16.25%和5.76%。三级结构分析结果显示,猪SENP1蛋白中存在8个α-螺旋区和7个β-转角区。实时荧光定量PCR结果显示,猪SENP1基因在JEV感染的PK15细胞中呈现上调表达趋势,其中在感染后48 h SENP1基因表达量显著高于对照组(P<0.05)。本试验结果为后续深入研究SENP1基因功能提供了参考。     

上皮间质转化促乳腺癌机制及药物干预治疗研究进展

乳腺癌是犬、猫等伴侣动物与人类常发疾病,作为人类及动物常患恶性肿瘤和主要致死肿瘤之一,其疾病负担仍呈逐步加重趋势,乳腺癌的预防及治疗形势愈加严峻。上皮间质转化(EMT)是乳腺癌发生发展中重要的生物学过程。EMT还可促进恶性肿瘤的侵袭、扩散及耐药,因此它在肿瘤的研究中日益受到关注,靶向于EMT是治疗乳腺癌的重要研究方向与热点。文章就EMT发生过程中细胞形态功能及标志物的变化、EMT分类及EMT与乳腺癌的关系分别展开论述,详细解析了EMT相关TGF-β/Smad、NF-κB及Wnt信号通路转导途径;随后对乳腺癌治疗药物研究进展,包括TGF-β/Smad通路抑制剂开发,相关药物、基因及细胞因子治疗前景、NF-κB通路与Wnt通路抑制剂的动物试验研究结果进行了详细论述;最后对乳腺癌的治疗发展与趋势进行了展望。深入认识信号通路调控乳腺癌EMT的生物学过程,明确其发生发展机制,寻找关键靶点及开发靶向药物,将为乳腺癌的精准治疗带来曙光。     

m6A甲基化在鸡肌肉生长发育中的表达研究

试验旨在研究RNA m6A修饰相关基因去甲基化酶Alk B同源蛋白5（Alk B homologue 5,ALKBH5）、去甲基化酶肥胖相关蛋白（fat mass and obesity-associated protein,FTO）、甲基转移酶样蛋白3（methyltransferase like 3,METTL3）、甲基转移酶样蛋白14（methyltransferase like 14,METTL14）和成肾细胞瘤1-结合蛋白（Wilms’tumor 1-associating protein,WTAP）在鸡骨骼肌发育过程中的表达,分析其与骨骼肌m6A甲基化水平的相关性。首先,利用实时荧光定量PCR技术检测m6A甲基化相关基因在金茅花鸡12（E12）、14（E14）、16（E16）、18（E18）胚龄和1日龄腿肌和胸肌组织中mRNA表达水平,以及其在鸡成肌细胞50%、100%增殖期和1、2、3、4、5 d分化期的mRNA表达水平;随后,利用m6A甲基化试剂盒检测金茅花鸡E12和1日龄腿肌和胸肌组织中m6A甲基化修饰水平,与m6A甲基化相关基因表达水平进行相关性分析。结果显示,m6A去甲基化基因ALKBH5和FTO mRNA表达水平在骨骼肌发育过程中显著上调（P<0.05）,即在E12、E14低表达,E16、E18逐渐上调,1日龄达到最高。m6A甲基化写入基因METTL14、METTL3和WTAP mRNA表达水平在E12、E14、E16逐渐上升,E18下降,随后至1日龄表达量回升。在细胞增殖过程中,ALKBH5、FTO、METTL14、METTL3和WTAP基因表达均上调;在细胞分化过程中ALKBH5和FTO基因表达水平显著上调（P<0.05）,在分化第5天达到最高。METTL14、METTL3和WTAP基因mRNA表达水平在细胞诱导分化的1、2、3、4 d表达量呈下降趋势,而在诱导分化的第5天有所回升。甲基化水平检测结果显示,腿肌和胸肌m6A甲基化水平变化趋势一致,均在胚胎发育过程中显著下降（P<0.05）,至1日龄达到最低。相关性分析结果显示,鸡骨骼肌RNA m6A甲基化水平与m6A去甲基化修饰基因ALKBH5、FTO mRNA表达水平呈显著负相关（P<0.05）。综合以上试验结果,推测m6A甲基化修饰与鸡骨骼肌发育相关,而去甲基化基因ALKBH5、FTO可能通过调控RNA m6A甲基化水平,影响鸡骨骼肌发育。本研究结果为进一步研究m6A甲基化修饰调控鸡骨骼肌生长发育的功能和分子机制提供理论依据。     

Characteristics of 18F-FDG and 18F-FDOPA PET in an 8-year-old neutered male Yorkshire Terrier dog with glioma: long-term chemotherapy using hydroxyurea plus imatinib with prednisolone and immunoreactivity for PDGFR-β and LAT1

An 8-year-old neutered male Yorkshire Terrier dog presented with head pressing, vestibular ataxia, neck tenderness, and no oculocephalic reflex. A demarcated lesion in the pons was identified on MRI. The patient was tentatively diagnosed with a glioma and was treated with hydroxyurea plus imatinib and prednisolone. After 30 days of therapeutic treatment, the patient showed a clear improvement in neurological signs, which lasted for 1117 days. On day 569 after the initiation of treatment, ¹⁸F-fluorodeoxyglucose (FDG)-positron emission tomography (PET) was performed with no significant findings on visual analysis. The average and maximal standardized uptake values (SUVs) were 1.92 and 2.29, respectively. The tumor-to-normal-tissue (T/N) ratio was 0.97. The first evidence of clinical deterioration was noticed on day 1147. On day 1155, 3,4-dihydroxy-6-[¹⁸F]-fluoro-l-phenylalanine (¹⁸F-FDOPA)-PET was performed. High uptake of ¹⁸F-FDOPA was observed in the intracranial lesion. The mean and maximal SUVs of the tumor were 1.59 and 2.29, respectively. The T/N ratio was 2.22. The patient was euthanized on day 1155 and histopathologic evaluations confirmed glioma (astrocytoma). This case shows that chemotherapy with hydroxyurea plus imatinib may be considered in the treatment of canine glioma. Furthermore, this is the first case describing the application of ¹⁸F-FDG and ¹⁸F-FDOPA in a dog with glioma.