GLP-1-IgG4-Fc融合蛋白的表达与鉴定

将合成的人胰高血糖素样肽-1(glucagon-like peptide-1,GLP-1)突变体基因与IgG4抗体的Fc部分进行融合获得GLP-1-IgG4-Fc片段,获得的基因片段与pXC17.4载体进行连接,用电转化方法将线性化质粒稳定转染CHO-K1细胞,通过Clone Pix 2筛选出高表达细胞株,产量达1.5g/L。收集培养上清并经Protein A和Source 30Q纯化,得到的GLP-1-IgG4-Fc融合蛋白,SDS-PAGE纯度高于95%,高效液相色谱(high performance liquid chromatography,HPLC)纯度和毛细管区域凝胶电泳(capillary zone electrophoresis,CZE)纯度均不低于80%,尺寸排阻层析(size-exclusion chromatography,SEC)纯度高于99%。经质谱和肽图谱测定,分子量与理论值一致,肽图谱序列与对照品高度一致。生物学活性分析表明,GLP-1-IgG4-Fc融合蛋白具有促进表达有GLP-1受体的HEK293细胞分泌环磷酸腺苷(cyclic adenosine monophosphate,c AMP)的活性,并且该活性与对照品高度相似。     

重金属累积对泽兰实蝇幼虫生长发育和保护酶的影响

为了弄清重金属通过寄主植物紫茎泽兰累积后对专性寄生天敌泽兰实蝇的生理影响,采用模拟Cd~(2+)、Pb~(2+)或Zn~(2+)污染的土壤培养法胁迫紫茎泽兰,让泽兰实蝇低龄幼虫寄生取食,利用原子吸收分光光度法测定泽兰实蝇老熟幼虫中Cd~(2+)、Pb~(2+)或Zn~(2+)的含量,同时测定分析泽兰实蝇老熟幼虫的体长、鲜重、能量物质和保护酶的变化。结果表明,实验处理的Cd~(2+)、Pb~(2+)或Zn~(2+)均可通过紫茎泽兰传递并累积到泽兰实蝇老熟幼虫体内,引起老熟幼虫一系列生理变化:长度和鲜重降低;总糖、总蛋白和总脂的含量降低,相应的热量值也降低;超氧化物歧化酶(SOD)、过氧化物酶(POD)和过氧化氢酶(CAT)等保护酶的活性降低。本研究为进一步阐明重金属污染对食物链各环节的影响提供了重要的理论依据。     

Expression of Androgen Receptor and Cancer Stem Cell Markers (CD44+/CD24− and ALDH1+): Prognostic Implications in Invasive Breast Cancer

BACKGROUND: Androgen receptor (AR) has emerged as a significant prognostic marker in early breast cancer (BCa). Association of AR with cancer stem cell (CSC) markers in BCa is unknown. Aim of the present study was to evaluate the immunohistochemical expression of AR, CD44, CD24 and ALDH1 in a cohort of Pakistani patients diagnosed with invasive BCa and to correlate the expression with 5- year disease free survival. PATIENTS AND METHODS: We evaluated immunohistochemical expression AR, CD44, CD24 and ALDH1 in formalin fixed paraffin embedded archival blocks of 166 cases of primary invasive BCa (stage I-III) and correlated the expression with clinicopathological variables and outcome using univariable and multivariable analysis. Survival data was computed by Kaplan Meier curves. RESULTS: Expression of AR was observed in 62.7% tumors whereas CD44, CD24 and ALDH1 were expressed in 61.4%, 44% and 30.1% tumors, respectively. AR expression was significantly associated with T1-T2 tumors, lower grade, estrogen and progesterone receptor expression (P < .05) and remained an independent prognostic indicator in multivariable analysis (adjusted HR 0.33, 95% CI 0.13–0.81; P = .016). Significant association was observed between concordant expression of AR and CD24 (P = .001) with a favorable impact on survival (P = .007) whereas expression of CSC phenotypes (CD44⁺, CD44⁺/CD24^? and ALDH1⁺) did not correlate with adverse outcome (P > .05). However, AR expression retained the association with better prognosis even in patients whose tumors exhibited a CSC phenotype. CONCLUSIONS: Expression of AR and CD24 in stage I-III invasive BCa correlates with favorable clinicopathological features and delineates a subgroup of patients with better disease-free survival.     

Nucleotide polymorphism in colicin E2 gene clusters: evidence for nonneutral evolution

To explore the molecular mechanisms behind the diversification of colicin gene clusters, we examined DNA sequence polymorphism for the colicin gene clusters of 14 colicin E2 (ColE2) plasmids obtained from natural isolates of Escherichia coli. Two types of ColE2 plasmids are revealed, with type II gene clusters generated by recombination between type I ColE2 and ColE7 gene clusters. The levels and patterns of DNA polymorphism are different between the two types. Type I polymorphism is distributed evenly along the gene cluster, while type II accumulates polymorphism at an elevated rate in the 5' end of the colicin gene. These differences may be explained by recombinational origins of type II gene clusters. The pattern of divergence between the ColE2 gene cluster and its close relative ColE9 is not correlated with the pattern of polymorphism within ColE2, suggesting that this gene cluster is not evolving in a neutral fashion. A statistical test confirms significant departures from the predictions of neutrality. These data lend further support to the hypothesis that colicin gene clusters may evolve under the influence of nonneutral forces.      

浙江潜水蜂属一新种（膜翅目：姬蜂总科：潜水蜂科）

本文报道采用浙江庆元百山祖自然保护区的潜水蜂属Ａｇｒｉｏｔｙｐｕｓ Ｃｕｒｔｉｓ１新种;浙江潜水蜂Ａ;ｚｈｅｊｉａｎｇｅｎｓｉｓ Ｈｅ ｅｔ Ｃｈｅｎ,ｓｐ;．ｎｏｖ。模式标本存浙江农业大学。     

细小病毒H-1对人肝癌移植瘤的抑制及其组织学与组织化学的研究

本文用人肝癌裸小鼠QGY-9204移植模型为材料,进行了细小病毒H-1抑瘤的组织学、组织化学及分子生物学研究。以两种不同剂量(5×10~7PFU和5×10~8PFU)的H-1进行瘤内注射,发现注入H-1后的肿瘤生长速率明显缓于对照组,且5×10~7PFU的H-1注入即可产生这一效应。以1×10~8PFU H-1注入瘤内,不同时间进行组织切片观察,发现从感染后第3天起瘤内开始出现坏死,且随感染时间的延长坏死范围也逐渐扩大,而对照组在该期间均未见此变化。电镜结果也表明,在受损的肿瘤细胞内有H-1病毒颗粒存在。PCR反应和ABC免疫染色显示在感染后的肿瘤组织内也有H-1 DNA扩增和NS-1蛋白表达,并且两者显示的时间与组织内坏死的过程相一致。这些结果提示,细小病毒H-1通过它的DNA复制和NS-1蛋白的表达以促进肿瘤的坏死,从而达到肿瘤抑制和裂解。     

Quox-1基因同源序列在人胚早期发育中的表达

以Quox-1基因的特异性片段b_2为探针,与人基因组DNA作Southern杂交,结果显示,人基因组中存在Quox-1基因的同源序列。以抗Quox-1蛋白的抗体与早期人胚胎组织的切片作免疫组化反应研究了Quox-1基因同源序列在人胚早期发育过程中的表达,结果表明其表达有明显的时间和空间特异性。胚龄30天以前,Quox-1基因的同源序列在人胚包括神经管等许多部位表达,30天以后表达部位局限于脊索、心肌细胞、生肌节、消化道上皮及周皮等处。文中讨论了Quox-1基因同源顺序对人类胚胎早期发育过程可能的调控作用。     

活性氧参与一氧化氮诱导的神经细胞凋亡

采用激光共聚焦成像技术，用氧化还原敏感的特异性荧光探针(DCFH-DA和DHR123)直接研究了一氧化氮供体S-亚硝基-N-乙酰基青霉胺(SNAP)诱导未成熟大鼠小脑颗粒神经元凋亡过程中的细胞胞浆、线粒体中活性氧水平的变化，发现神经细胞经0.5 mmol/L SNAP处理1 h后，细胞胞浆及线粒体中活性氧水平大大增加.一氧化氮清除剂血红蛋白能够有效抑制细胞胞浆、线粒体中活性氧的产生，防止细胞凋亡.外源性谷胱甘肽对细胞也具有良好的保护作用，而当细胞中谷胱甘肽的合成被抑制后，一氧化氮的神经毒性大大增强.实验结果表明一氧化氮通过促进神经细胞产生内源性活性氧而启动细胞凋亡程序，而谷胱甘肽可能是重要的防止一氧化氮引发神经损伤的内源性抗氧化剂.