心脏特异表达Calponin 1转基因小鼠的心脏功能分析

目的建立心脏特异表达Calponin 1转基因小鼠,研究Calponin 1对心脏发育及心肌病的调节作用。方法利用心脏特异启动子α-MHC构建转基因表达载体,显微注射法建立Calponin 1转基因小鼠,PCR法鉴定转基因小鼠的基因型,Western Blot检测Calponin 1在心脏组织中的表达,心脏超声检测转基因小鼠的心脏结构和功能,HE染色和Masson染色检测转基因小鼠心脏的病理改变。结果 Calponin 1在野生型小鼠心脏中有表达,在扩张型心肌病小鼠的心脏组织表达降低。通过显微注射法,建立了2个心脏组织Calponin 1基因高表达的转基因小鼠系。与野生型小鼠相比,Calponin 1转基因小鼠收缩期左室内径（LVID,systolic）增加28%（P〈0.01,n=12）,舒张期左室内径（LVID,diastolic）增加16.2%（P〈0.01,n=12）,收缩期左室后壁厚度（LVPW,systolic）减小15.7%（P〈0.01,n=12）,舒张期左室后壁厚度（LVPW,diastolic）减小21%（P〈0.01,n=12）,射血分数（ejection fraction,EF）降低11.5%（P〈0.01,n=12）,短轴内径缩短率（fraction shortening,FS）降低14.6%（P〈0.05,n=12）。转基因小鼠心脏组织病理H＆E染色和Masson染色显示,转基因小鼠心室扩张,心肌细胞不均匀肥大,细胞间隙变大,心肌间质纤维增多。结论 Calponin 1在心脏特异过表达引起转基因小鼠心脏左室内径增加,收缩期容积和舒张期容积显著增大,心室壁变薄,射血分数及短轴缩短率降低等扩张性心肌病表型,推测Calponin 1是参与心肌病病理发生的基因之一。     

独脚金内酯调控侧枝发育的研究进展

植物通过内源激素或环境信号调控叶腋内腋芽的形成和发育,从而控制其分枝特性。独脚金内酯(strigolactones,SLs),一种产生于植物根部的类胡萝卜素衍生物,具有刺激寄生植物种子的萌发和促进丛枝菌根真菌菌丝分枝的作用,最近的研究表明,它还可以沿茎干向上运输,与生长素和细胞分裂素一起直接或间接抑制植物分枝,目前已经作为一种新的植物激素受到广泛认可。本文综述了独脚金内酯的结构、合成途径和生物活性,以及调控植物分枝的分子机理,并展望了其在抑制杂草或新型除草剂的研发、促进植物和有益真菌的共生,以及调控作物的分枝和株型等方面的应用前景。     

甘蓝型油菜BnCOP1基因编码区全长cDNA的克隆与功能研究

通过分析拟南芥、豌豆、番茄和水稻的COP1 (constitutively photomorphogenic 1) 的cDNA序列, 运用RT-PCR和改进的基因组步行 (genome walking) 技术相结合的方法, 首次从甘蓝型油菜中克隆到油菜 BnCOP1编码区cDNA的全长序列, 其全长2 034 bp, 编码677个氨基酸. 同源 性分析表明, 其编码的氨基酸序列与拟南芥的同源性高达94%. 对BnCOP1编 码序列（cDNA）演绎出的氨基酸序列分析表明, 其编码的蛋白包含有N端的 环形锌指结合域(ring finger zinc binding domain, RING)、中间的卷曲 螺旋形结构域(coiled-coil domain, coiled-coil ), 7个C端的WD-40重复 序列(WD-40 repeats, WD-40)的功能域. 半定量RT-PCR和实时荧光定量PCR 分析该基因在油菜中的表达模式,结果显示, BnCOP1在甘蓝型油菜的各个组 织器官中均有表达,其中在花中的表达明显高于在根、叶、茎、果荚及子叶 和胚轴中,暗示该蛋白可能与开花途径相关. 过表达BnCOP1的转基因拟南芥 植株在高度、主茎的直径和叶片大小上都呈现出比野生型弱小的表型, 表 明BnCOP1抑制了拟南芥光形态建成从而影响了植物的生长发育.     

影响农杆菌介导的植物转基因的因素

农杆菌介导法是植物转基因研究中应用最多的方法,具有转化成本低、效率高、可转移较长基因片段等特点.多种因素影响此遗传转化过程中的转化率,包括外植体类型、植物基因型、外植体的处理、菌株和质粒载体类型、预培养以及培养基成份等方面.     

Effects of a synthetic PEG-ylated Tie-2 agonist peptide on endotoxemic lung injury and mortality

A synthetic 7-mer, HHHRHSF, was recently identified by screening a phage display library for binding to the Tie-2 receptor. A polyethylene-oxide clustered version of this peptide, termed vasculotide (VT), was reported to activate Tie-2 and promote angiogenesis in a mouse model of diabetic ulcer. We hypothesized that VT administration would defend endothelial barrier function against sepsis-associated mediators of permeability, prevent lung vascular leakage arising in endotoxemia, and improve mortality in endotoxemic mice. In confluent human microvascular endothelial cells, VT prevented endotoxin-induced (lipopolysaccharides, LPS O111:B4) gap formation, loss of monolayer resistance, and translocation of labeled albumin. In 8-wk-old male C57Bl6/J mice given a ～70% lethal dose of endotoxin (15 mg/kg ip), VT prevented lung vascular leakage and reversed the attenuation of lung vascular endothelial cadherin induced by endotoxemia. These protective effects of VT were associated with activation of Tie-2 and its downstream mediator, Akt. Echocardiographic studies showed only a nonsignificant trend toward improved myocardial performance associated with VT. Finally, we evaluated survival in this mouse model. Pretreatment with VT improved survival by 41.4% (n = 15/group, P = 0.02) and post-LPS administration of VT improved survival by 33.3% (n = 15/group, P = 0.051). VT-mediated protection from LPS lethality was lost in Tie-2 heterozygous mice, in agreement with VT's proposed receptor specificity. We conclude that this synthetic Tie-2 agonist, completely unrelated to endogenous Tie-2 ligands, is sufficient to activate the receptor and its downstream pathways in vivo and that the Tie-2 receptor may be an important target for therapeutic evaluation in conditions of pathological vascular leakage.     

Biosimilars clinical development program: confirmatory clinical trials: a virtual/simulated case study comparing equivalence and non-inferiority approaches

As part of long term commitment of the Biologicals and Vaccines Committee (B&V) of the International Federation of Pharmaceutical Manufacturers and Association (IFPMA) to provide expert input to the WHO on their recently finalized GUIDELINES ON EVALUATION OF SIMILAR BIOTHERAPEUTIC PRODUCTS (SBPs), and in response to WHO's request, the IFPMA B&V prepared a clinical case study at a recent WHO workshop in Seoul, Korea. The case study, presented by Mark Fletcher on behalf of B&V, involved a model scenario for a clinical efficacy trial to support the approval of a Similar Biotherapeutic Product (SBP) as part of the required comparative clinical program against a Reference Biotherapeutic Product (RBP). A key goal was to understand and illustrate key clinical and statistical principles, and considerations described in the WHO Guidance for regulatory authorities when designing and implementing WHO guidelines and post-approval regulatory oversight for SBPs. Using this model SBP/RBP pair, an interactive discussion was carried out among the workshop participants on the pros and cons of using equivalence vs. non-inferiority designs to assess the two products' similarity. Through discussion of the case, the complexity of demonstrating similar efficacy and safety of a SBP vs. RBP for biotherapeutic products is outlined and discussed in the context of the key principles laid out in the recently published WHO GUIDELINES ON EVALUATION OF SIMILAR BIOTHERAPEUTIC PRODUCTS (SBPs). The exercise illustrates the need for a case-by-case approach when interpreting clinical data from SBP dossiers to adequately assure similar efficacy and safety of SBPs for any studied indication.     

酪酸梭菌活菌散在防治早产儿喂养不耐受方面的应用

目的观察和评价酪酸梭菌活菌散防治早产儿喂养不耐受的临床疗效。方法将56例早产儿随机分成观察组和对照组,观察组30例,对照组26例。对照组给予早产儿配方奶及部分肠外营养等常规治疗,观察组在常规治疗基础上同时添加酪酸梭菌活菌散。观察2组早产儿恢复出生体重、达到全胃肠喂养时间及发生喂养不耐受等情况。结果观察组早产儿恢复出生体重、达到全胃肠喂养时间明显短于对照组(P<0.01或P<0.05),喂养不耐受的发生率也显著少于对照组(P<0.05)。结论早产儿服用酪酸梭菌活菌散,对防治喂养不耐受具有积极的作用,能减少早产儿喂养过程中呕吐、胃潴留、腹胀的发生,促进患儿早期的生长发育,缩短达到全胃肠喂养的时间。     

拮抗菌株Kc-t99的鉴定及其抑菌活性研究

通过传统分类与分子生物学技术相结合的方法对从京郊菜园土壤中分离筛选的拮抗菌株Kc-t99进行鉴定,并采用生物测定的方法评价其抑菌活性.结果表明,菌株Kc-t99的形态学特征和生理生化特性与枯草芽孢杆菌基本一致,其16S rDNA序列与GenBank中已鉴定的枯草芽孢杆菌的16S rDNA序列同源性高达98.06%,据此初步确定其为枯草芽孢杆菌.抑菌试验表明该菌株对供试的5种病原真菌和4种细菌均具抑菌活性,其中对甘蓝枯萎病菌、黄瓜角斑病菌和桃褐腐病菌抑制作用显著.     

水稻蛋白质组双向电泳优化流程及方法

双向电泳是分析蛋白质混合物的一种有力手段,已在蛋白质组研究中得到广泛应用。水稻（Oryzasativa）作为重要的粮食作物,对其蛋白质组学研究开展较早。但由于技术复杂,对实验操作要求高,初学的研究者很难在较短的时间内掌握该实验技术。该文介绍了水稻研究中适合多个组织的双向电泳实验方法和优化流程。该优化流程能使新的研究者逐步优化实验条件,更快更好地完成双向电泳实验。同时详细介绍了实验关键环节的操作方法。     

Functions of lipopeptides bacillomycin D and fengycin in antagonism of Bacillus amyloliquefaciens C06 towards Monilinia fructicola

In previous studies, Bacillus amyloliquefaciens C06 has been proven to be effective in controlling brown rot of stone fruit caused by Monilinia fructicola. When tested in vitro, cell-free filtrate of B. amyloliquefaciens C06 significantly inhibited mycelial growth and conidial germination of the fungal pathogen. This study aimed to determine the role of the antifungal compound(s) in the cell-free filtrate of B. amyloliquefaciens C06 by an approach combining a DNA-based suppression subtractive hybridization (SSH) method with MALDI-TOF-MS analysis. It was demonstrated that B. amyloliquefaciens C06 harbored two genes, bmyC and fenD, involved in biosynthesis of bacillomycin D and fengycin, two lipopeptides belonging to the iturin and fengycin family, respectively. To determine the roles of bacillomycin D and fengycin of B. amyloliquefaciens C06 in suppressing M. fructicola, the mutants of B. amyloliquefaciens C06 deficient in producing bacillomy- cin D, fengycin or both were constructed, and evaluated in vitro together with the wild-type B. amyloliquefaciens C06. The results indicated that bacillomycin D and fengycin jointly contributed to the inhibition of conidial germination of M. fructicola, and fengycin played a major role in suppressing mycelial growth of the fungal pathogen.