Targeted Gene Transfer for Adenocarcinoma Using a Combination of Tumor-specific Antibody and Tissue-specific Promoter

We have developed a highly specific gene transfer method for adenocarcinoma using a monoclonal antibody against tumor-specific antigen coupled with a plasmid containing the carcinoembryonic antigen (CEA)-specific promoter. The chimeric CEA promoter (CC promoter), which contained an enhancer from the immediate early gene of cytomegalovirus and the CEA promoter, achieved 4- to 5-fold higher transgene expression in CEA-producing cells than the original CEA promoter while maintaining CEA specificity. Furthermore, a complex of a monoclonal antibody against Lewis Y antigen (LYA), the CC promoter-containing plasmid and cationic liposomes (DOTAP) achieved specific gene expression in CEA-producing and LYA-positive adenocarcinoma cell lines that was 200-fold more efficient than in CEA-non-producing and LYA-negative cell lines during a short in vitro incubation. This strategy may be applicable for clinical gene therapy.     

用荧光探针JC-1检测氧化应激对精子线粒体膜电位的影响

目的：利用荧光探针JC-1观察氧化应激对精子线粒体膜电位的变化。方法：用FeSO4/H2O2体系诱导精子氧化损伤模型，利用新型荧光探针JC 1标记精子线粒体,在FACS Calibur流式细胞仪上检测JC 1荧光强度的变化。结果：正常精子线粒体维持较高的膜电位,JC 1在线粒体内形成聚合物, 红色荧光占主导地位。FeSO4/H2O2体系导致线粒体膜电位下降, JC 1聚合物分解成单体,红色荧光强度减弱。结论：氧化应激导致精子线粒体膜电位下降; JC 1结合流式细胞仪是一种理想的检测线粒体膜电位的手段。     

Antimicrobial activity of myotoxic phospholipases A2 from crotalid snake venoms and synthetic peptide variants derived from their C-terminal region.

A short peptide derived from the C-terminal region of Bothrops asper myotoxin II, a Lys49 phospholipase A(2) (PLA(2)), was previously found to reproduce the bactericidal activity of its parent molecule. In this study, a panel of eight PLA(2) myotoxins purified from crotalid snake venoms, including both Lys49 and Asp49-type isoforms, were all found to express bactericidal activity, indicating that this may be a common action of the group IIA PLA(2) protein family. A series of 10 synthetic peptide variants, based on the original C-terminal sequence 115-129 of myotoxin II and its triple Tyr-->Trp substituted peptide p115-W3, were characterized. In vitro assays for bactericidal, cytolytic and anti-endotoxic activities of these peptides suggest a general correlation between the number of tryptophan substitutions introduced and microbicidal potency, both against Gram-negative (Salmonella typhimurium) and Gram-positive (Staphylococcus aureus) bacteria. Peptide variants with high bactericidal activity also tended to be more cytolytic towards skeletal muscle C2C12 myoblasts, thus limiting their potential in vivo use. However, the peptide variant pEM-2 (KKWRWWLKALAKK) showed reduced toxicity towards muscle cells, while retaining high bactericidal potency. This peptide also showed the highest endotoxin-neutralizing activity in vitro, and was shown to functionally interact with lipopolysaccharide (LPS) using a chimeric bacteria model. The bactericidal and anti-endotoxic properties of pEM-2, combined with its relatively low toxicity towards eukaryotic cells, highlight it as a promising candidate for further evaluation of its antimicrobial potential in vivo.     

细胞凋亡的检测技术与方法

细胞凋亡在保证多细胞生物的健康生存过程中扮演着关键的角色。根据死亡细胞的形态学、生物化学和分子生物学特征,可以将凋亡细胞与坏死细胞区别开来。本文综述了常用的细胞凋亡检测技术与方法的基本原理,如形态学观察、荧光素染色、DNA阶梯、半胱氨酸天冬氨酸蛋白酶活性的检测等,并对各种方法的优点及局限性做一简要比较。用透射电镜进行超微结构观察仍是鉴定细胞凋亡的金标准。在选择细胞凋亡检测方法时,要充分了解各种方法的原理和应用范围,进行综合考虑。多种检测方法的联合应用常可获得准确的结果。     

不同商品规格川贝母总生物碱含量测定的方法研究

目的 建立酸性染色法用于测定川贝母中总生物碱的含量.方法 采用三氯甲烷-甲醇(4∶1)混合溶液提取川贝母总生物碱;采用酸性染料染色法,以西贝母碱为指标成分,测定总生物碱的含量.结果 4个规格川贝母样品中总生物碱含量均符合2010年版《中国药典(一部)》的规定.结论 该方法不需要复杂仪器、操作简便,可作为川贝母的质量检测方法.     

嗜酸细胞活化趋化因子在变应性鼻炎中的作用

目的 通过建立变应性鼻炎(allergjcrhinitis,AR)豚鼠模型,探讨嗜酸细胞活化趋化因子(eotaxin)对鼻黏膜中嗜酸细胞(eosinophils,Eos)局部浸润及活化的作用,进一步阐明Eos在鼻黏膜的摹集及其活化的机制.方法 卵清蛋白致敏法制备AR豚鼠模型,分别采用免疫组化、逆转录聚合酶链反应技术检测eotaxin蛋白及mRNA在鼻黏膜中的表达,免疫荧光法测定鼻腔冲洗液中嗜酸细胞阳离子蛋白(eosinophilcationic protein,ECP)含量标示Eos活化程度,鼻黏膜HE切片每高倍视野下Eos平均个数标示Eos浸润程度.结果与对照组相比,AR模型组鼻黏膜中eotaxin蛋白阳性表达率、eotaxin mRNA表达、Eos个数以及鼻腔灌洗液中ECP含量均显著增加(P<0.001,P=0.006,P<0.001,P<0.001);相关分析表明eotaxin表达与Eos浸润程度及ECP含量呈正相关(r分别=0.849、0.691、0.656、0.81 9).地塞米松干预后以上指标一致性下降.结论 AR豚鼠鼻黏膜中eotaxin表达增强,是Eos局部浸润和活化的重要影响因素;地塞米松可以通过降低AR豚鼠鼻黏膜中eotaxin的表达减少鼻黏膜中Eos的局部浸润和活化.     

脂质体-IP10复合物对小鼠4T1乳腺癌的治疗作用

目的 用1,2-二油酰-3-三甲胺基丙烷(DOTAP)+胆固醇(CHOL)脂质体包裹pcDNA3.1-γ干扰素诱导蛋白-10(IP10)制备脂质体质粒DNA复合体即Lip-IP10,观察其对抗小鼠4T1乳腺癌的原位瘤及肺转移瘤的治疗作用,并探讨其作用机制.方法 建立4T1乳腺癌模型,将36只荷瘤小鼠随机分为生理盐水(NS)组、脂质体-pcDNA3.1复合物(Lip-null)组和脂质体-IP10复合物(Lip-IP10)组,尾静脉给药,观察各组肿瘤体积、肺转移结节数及小鼠生存期.原位瘤进行CD31染色,测定微血管密度(MVD).结果 与两对照组比较,Lip-IP10组的肿瘤体积明显较小(P<0.05),肺转移结节显著减少(各组中位数为NS组45个,Lip-null组40个,Lip-IP10组 3个,P<0.05),生存期延长(P<0.05),CD31染色结果 显示,Lip-IP10组的肿瘤MVD值较低(NS组44.25±5.51,Lip-null组 43.45±4.21,Lip-IP10组21.50±5.41,P<0.05). 结论 静脉给予Lip-IP10复合体可抑制小鼠4T1乳腺癌的原位瘤及肺转移瘤,其作用机制与IP10抑制肿瘤血管生成有关.     

对苯二胺氧化产物对小鼠经皮肤染毒的亚慢性毒性

[目的 ]研究氧化型染发剂主要成分对苯二胺 (PPD)氧化产物经皮吸收后对小鼠的亚慢性毒作用。 [方法 ]对苯二胺经双氧水氧化后的氧化产物为受试物 ,每隔 3d在小鼠背部皮肤染毒 1次 ,周期 1个月。整个过程中观察小鼠的一般生长情况 ,并于染毒 1个月后观察受试动物血生化指标、血常规、肝肾脾组织病理学改变和遗传毒性。 [结果 ]PPD氧化产物可影响小鼠的肝肾功能 ,随着PPD染毒剂量增加 ,ALT和BUN值逐渐增高 ,与对照组比较差异有显著性 (P<0 0 5或P <0 0 1)。PPD具有遗传毒性 ,能造成骨髓细胞染色体和DNA的损伤 ,并在受试剂量范围内存在剂量 效应关系。 [结论 ]频繁使用氧化型染发剂可能对机体具有潜在的危害     

大鼠海马结构至额前皮质和外侧隔核的侧支投射——逆行荧光双标记法

本实验用大鼠24只,采用逆行荧光染料示踪法进行双标记研究。将荧光示踪物二盐酸4′-6-双脒基-2-苯吲哚(DAPI)、碘化丙啶(PI)和依文思蓝(EB)注射入额前皮质的边缘下区以及外侧隔核。结果表明用 DAPI 和 PI 配对注射入上述区破时,可见到海马结构的腹侧下托及其 CA_1区颞侧部有明显的双标记细胞,双标记细胞多位于锥体层,偶见于放射层和起层。提示腹侧下托和 CA_1区颞侧部有侧支投射到额前皮质的边缘下区和外侧隔核。     

染料木素对维甲酸模型大鼠血生化指标和骨密度的作用

目的 :探讨染料木素对维甲酸所致骨质疏松模型大鼠的影响 .方法 :取SD雄性大鼠 2 5 6只 ,随机分为正常对照组、模型组、骨疏康组 (5g·kg-1)、染料木素大、中、小剂量组(18,9,4 .5mg·kg-1) .给予维甲酸 70mg·kg-1·d-1灌胃 2wk(正常对照组除外 ) ,分别于不同时间从各组取 8只大鼠对体质量、血生化指标和骨密度等进行测定 .结果 :染料木素①可增加维甲酸模型大鼠的左股骨质量、左胫骨质量 (P <0 .0 5或P <0 .0 1) ;②使模型大鼠的血清Ca ,P ,Mg,CT水平明显降低 ,BGP ,ALP水平升高 ,与模型组有显著差异 (P <0 .0 5或P <0 .0 1) ;③在 4 .5mg·kg-1,9mg·kg-1时可明显增强模型大鼠的股骨、胫骨和腰椎L2 -4的骨密度 (P <0 .0 5或P <0 .0 1) .结论 :染料木素可明显改善维甲酸模型大鼠的骨密度和血生化指标 ,对骨质疏松有较好的疗效