HBV pre-c-Fc融合蛋白在杆状病毒表达系统中的表达及其生物学活性研究

目的:在杆状病毒表达系统中表达融合蛋白乙型肝炎病毒前C蛋白-小鼠IgG Fc蛋白(HBV precore protein-mouse IgG Fc,HBV pre-c-Fc),并鉴定其免疫原性。方法:目的基因HBV prec-Fc连接到pFastBac1载体,获得的pFastBac1-HBV pre-c-Fc质粒转化DH10Bac感受态,通过Tn7转座子将目的基因转座到Bacmid中,得到Bacmid-HBV pre-c-Fc穿梭载体,脂质体包被后转染Sf9昆虫细胞获得P1代病毒,重复转染Sf9获得高滴度病毒。收集细胞上清超滤后通过Protein G亲和层析柱纯化得到目的蛋白HBV pre-c-Fc。纯化的蛋白大腿内侧肌肉注射免疫BALB/c小鼠并检测血清中乙型肝炎病毒核心蛋白抗体产生量。结果:HBV pre-c-Fc在昆虫细胞中成功表达,纯化后蛋白纯度达90%以上,蛋白产量约为3.03mg/L,纯化蛋白能有效刺激BALB/c小鼠产生特异抗体。结论:成功地在杆状病毒表达系统中表达了具有免疫原性的HBV pre-c-Fc蛋白,为生产乙肝治疗性疫苗奠定了基础。     

Quantitation of mercapturic acid conjugates of 4-hydroxy-2-nonenal and 4-oxo-2-nonenal metabolites in a smoking cessation study

The breakdown of polyunsaturated fatty acids (PUFAs) under conditions of oxidative stress results in the formation of lipid peroxidation (LPO) products. These LPO products such as 4-hydroxy-2-nonenal (HNE) and 4-oxo-2-nonenal (ONE) can contribute to the development of cardiovascular and neurodegenerative diseases and cancer. Conjugation with glutathione, followed by further metabolism to mercapturic acid (MA) conjugates, can mitigate the effects of these LPO products in disease development by facilitating their excretion from the body. We have developed a quantitative method to simultaneously assess levels of 4-oxo-2-nonen-1-ol (ONO)-MA, HNE-MA, and 1,4-dihydroxy-2-nonene (DHN)-MA in human urine samples utilizing isotope-dilution mass spectrometry. We are also able to detect 4-hydroxy-2-nonenoic acid (HNA)-MA, 4-hydroxy-2-nonenoic acid lactone (HNAL)-MA, and 4-oxo-2-nonenoic acid (ONA)-MA with this method. The detection of ONO-MA and ONA-MA in humans is significant because it demonstrates that HNE/ONE branching occurs in the breakdown of PUFAs and suggests that ONO may contribute to the harmful effects currently associated with HNE. We were able to show significant decreases in HNE-MA, DHN-MA, and total LPO-MA in a group of seven smokers upon smoking cessation. These data demonstrate the value of HNE and ONE metabolites as in vivo markers of oxidative stress.     

酪氨酸激酶抑制剂类抗肿瘤药物研究方法进展

酪氨酸激酶(protein tyrosine kinases,PTKs)在肿瘤细胞的增殖、分化、迁移、侵袭等相关信号通路中起到了关键的调控作用,已经成为肿瘤靶向性治疗的重要靶点.本文对靶向酪氨酸激酶的小分子抑制剂的筛选和评价方法进行综述,以期促进酪氨酸激酶抑制剂类抗肿瘤药物的研究.     

卡托普利对高血压大鼠动脉血管平滑肌细胞内质网相关因子葡萄糖调节蛋白78和C/EBP同源蛋白表达的影响

目的:研究卡托普利对早期高血压大鼠动脉血管平滑肌细胞内质网相关因子葡萄糖调节蛋白78(glucose-regulated protein of 78kd,GRPT8)和C/EBP同源蛋白(CAAT/enhancer binding protein homologous protein,CHOP)表达的影响.方法:将18只成年雄性Sprague-Dawley(SD)大鼠随机分为对照组、模型组和卡托普利组(n=6),模型组和卡托普利组均采用大鼠腹主动脉结扎建立高血压大鼠模型.4周后测量血压,应用免疫组化方法检测主动脉血管平滑肌细胞GRP78和CHOP的表达;缺口末端标记法(TUNEL)检测细胞凋亡.结果:(1)模型组平均动脉压(mean arterial blood pressure,MAP)明显增加,卡托普利组血压低于模型组,高于对照组,差异具有统计学意义(P<0.05);(2)模型组内质网因子GRP78、CHOP表达均增加,卡托普利组GRP78、CHOP表达低于模型组,高于对照组,差异具有统计学意义(P<0.01);(3)模型组血管平滑肌细胞凋亡率减少,卡托普利组血管平滑肌细胞凋亡率高于模型组,低于对照组,差异具有统计学意义(P<0.01).结论:卡托普利可降低高血压大鼠动脉血管平滑肌细胞GRP78和CHOP的表达,增加细胞凋亡.可能与其减弱高血压所致内质网反应,维持血管平滑肌细胞的增殖/凋亡平衡有关.     

藏酋猴外周血细胞及血清生化指标测定与分析

目的 通过对原代藏酋猴进行外周血细胞及血液生化指标测定,了解不同年龄段藏酋猴的生理指标差异.方法 在清醒状态下采集30只藏酋猴后肢静脉血,检测血液学和血液生化指标.结果 与同属灵长类动物检测结果比较,具有较高的一致性,但也存在种属特点.结论 藏酋猴随着体重和年龄增大,血糖有增高的趋势.     

浙江省国家重点风景名胜区楠溪江两栖动物资源调查与区系分析

为了解浙江省永嘉县国家重点风景名胜区楠溪江野生动物资源现状,促进风景区的有效管理和建设,于2008年5月、8月、9月、10月和2009年5月对楠溪江流域的两栖动物进行野外调查.调查结果表明,楠溪江流域共计有两栖类24种,隶属于2目7科,中国特有种13种,其中福建大头蛙和天台粗皮蛙是温州地区的新纪录种.区系组成以东洋界华中华南区共有种为主,东洋界占明显优势,达22种,占物种总数的91.7%.     

Newly discovered penicillin acylase activity of aculeacin A acylase from Actinoplanes utahensis

Aculeacin A acylase from Actinoplanes utahensis produced by Streptomyces lividans revealed acylase activities that are able to hydrolyze penicillin V and several natural aliphatic penicillins. Penicillin K was the best substrate, showing a catalytic efficiency of 34.79 mM(-1) s(-1). Furthermore, aculeacin A acylase was highly thermostable, with a midpoint transition temperature of 81.5 degrees C.     

Natural products with calmodulin inhibitor properties

This review summarizes properties of various naturally occurring compounds with reported calmodulin (CaM)-inhibitory properties which include about 159 natural products belonging to different structural classes. Most inhibitors are alkaloid and peptide type of compounds and have been isolated from a wide variety of natural sources, including many plant species. Among the most potent natural anti-CaM substances, however, are several animal venoms and the antibiotic polymixin. The largest number of compounds described were discovered by means of enzymatic functional assays.     

大黄鱼对几种饲料蛋白原料消化率的研究

以白鱼粉为主要蛋白源制成参考饲料,以0.1%的Cr2O3为外源性指示剂,按参考饲料和实验原料70%∶30%的比例配制成实验饲料,测定了大黄鱼（Pseudosciaena crocea）对白鱼粉、红鱼粉、肉骨粉、豆粕、花生粕、菜籽粕和棉籽粕的表观消化率。实验大黄鱼（平均初始体重15.0±1.6g）养殖于海水浮式网箱（1.5m×1.5m×2.0m）中,分别以各实验饲料投喂实验鱼5周,然后采用挤压法收集粪便。实验结果表明各原料表观干物质消化率在43.6%至70.0%之间,能量消化率在42.9%到82.6%的范围内。实验原料的蛋白质表观消化率在70.7%到92.4%之间,其中白鱼粉和红鱼粉的蛋白消化率最高（分别为92.4%和89.3%）,而棉籽粕则最低（70.7%）。脂肪的表观消化率在61.3%到90.5%之间,其中棉籽粕最低（61.3%）。磷的表观消化率相对较低,仅白鱼粉和红鱼粉在53.2%以上,其余皆低于37.5%。总之,大黄鱼对这几种原料的表观消化率存在较大差异,但蛋白消化率均较高,因此,可作为大黄鱼的饲料蛋白源在实际饲料配制中使用。