细胞培养中支原体污染的去除

在运用细胞培养手段的生物学研究和生物工程产品中，支原体污染仍是一个颇为棘手的问题。１１株人的细胞ＳＰＣ－Ａ１、Ｋ５６２、ＨＵＴ－１０２、ＢＴ－３２５、６Ｔ－ＣＥＭ、ＮＫＭ－４５、ＣＮＥ、ＨＬ－６０、ＱＧＹ－７７０１、ＱＺＧ、人羊膜细胞；３株小鼠细胞Ｐ３８８－１、ＹＡＣ－１、Ｐ８１５；１株绒猴细胞Ｂ９５－８；和一株杂交瘤ＱＫＴ３等，已污染了支原体的细胞经ＭＣ－２１０ １μｇ／ｍｌ处理７天，支原体污染     

人瘦素基因克隆与序列测定

【目的】构建人瘦素基因cDNA克隆 ,并进行序列分析。【方法】用逆转录聚合酶链反应扩增人瘦素基因cDNA ,获得片段 (6 40bp)连接至 pUC19载体 ,并转化大肠杆菌DH5α。以末端终止法进行序列测定。【结果】所克隆的人瘦素cDNA含全长的人瘦素基因编码区 ,仅 2 87位的腺苷酸被鸟苷酸代替 ,导致 96位编码谷氨酰胺的密码子CAA改变为编码精氨酸的CGA ,其意义尚待阐明。【结论】以逆转录聚合酶链反应方法成功地构建了人瘦素基因cDNA克隆。     

DNA探针板杂交法检测临床标本中结核杆菌的应用

目的：在酶标板上进行ＤＮＡ探针杂交,评价其在检测临床标本中结核杆菌的应用价值。方法：收集７１份结核性标本和３２份非结核性标本,进行ＤＮＡ扩增,把扩增产物分别进行电泳和ＤＮＡ探针杂交,比较其结果。结果：７１份结核性标本电泳法阳性率为４５１％,ＤＮＡ探针法阳性率为６９０％;３２份非结核性标本二者均为阴性。结论：ＤＮＡ探针杂交可提高临床标本中结核杆菌的阳性检出率,是结核病早期诊断的一种新方法。     

大剂量干扰素治疗慢性丙型肝炎50例临床观察

目的：探讨使用大剂量干扰素（ＩＦＮα）治疗慢性丙型肝炎（ＣＡＨ）的效果。方法：使用大剂量ＩＦＮα,每天６００万Ｕ,连续投药３周,以后每周３次间断投药２１周,共计２４周,治疗ＣＡＨ５０例。观察治疗前后血清ＨＣＶＲＮＡ与ＡＬＴ的变化,以此作为ＩＦＮ疗效的判定指标。结果：ＩＦＮ治疗ＣＡＨ有效率为６８％,停药６个月后有３４％复发。结论：ＩＦＮ的疗效与ＣＡＨ的基因亚型有关（Ⅲ型疗效显著,Ⅱ型较差）,与ＩＦＮ的剂量无明显关系。     

Increased reactive oxygen species in familial amyotrophic lateral sclerosis with mutations in SOD1

Amyotrophic lateral sclerosis (ALS) is a paralytic disorder characterized by degeneration of large motor neurons of the brain and spinal cord. A subset of ALS is inherited (familial ALS, FALS) and is associated with more than 70 different mutations in the SOD1 gene. Here we report that lymphoblast cell lines derived from FALS patients with 16 different mutations in SOD1 gene exhibit significant increase of intracellular reactive oxygen species (ROS) compared with sporadic ALS (SALS) and normal controls (spouses of ALS patients). The ROS generation did not correlate with SOD1 activity. Further, cells incubated with vitamin C, catalase or the flavinoid quercetin significantly reduced ROS in all groups. The catalase inhibitor 3-amino-1,2,4-triazole resulted in a ten-fold increase of ROS in all groups. Neither L-nitroarginine, a nitric oxide synthase inhibitor or vitamin E altered the ROS levels. Thus, these studies suggest that hydrogen peroxide (H(2)O(2)) is a major ROS elevated in FALS lymphoblasts and it may contribute to the degeneration of susceptible cells. Further, we postulate a mechanism by which increased H(2)O(2) could be generated by mutant SOD1.     

氯沙坦对自发性高血压大鼠脑组织肾素血管紧张素系统的影响

目的　探讨氯沙坦对自发性高血压大鼠 (SHR)脑组织血管紧张素Ⅰ型受体 (AT1 R)mRNA的表达及脑组织局部肾素 血管紧张素系统 (RAS)活性的影响。方法　 6wk龄♂WKY和SHR各 1 6只分别随机分为氯沙坦用药组和生理盐水对照组 ,喂养 1 8wk后运用RT PCR法检测脑组织AT1 RmRNA的表达 ;放免法测定肾素活性 (RA)和血管紧张素Ⅱ(AngⅡ )的水平 ;紫外分光光度法测定血管紧张素转换酶活性 (ACEA)。结果　SHR组各部位脑组织AT1 RmRNA和血浆、下丘脑RA、AngⅡ、血清ACEA表达均高于WKY组(P <0 0 5) ;氯沙坦可降低各部位脑组织AT1 RmRNA的表达但升高血浆RA、AngⅡ ,对下丘脑RA、AngⅡ、ACEA则无明显影响。结论　SHR脑组织AT1 RmRNA的表达及循环和组织RAS活性增加可能与高血压的发生有关 ,氯沙坦可能通过降低脑组织AT1 RmRNA表达发挥脑保护作用     

华北大黄中非噁类成分的研究

目的　对华北大黄中非类成分进行研究。方法　色谱和光谱分析法分离和鉴定化学成分。结果　从 95 %乙醇提取物中分得 13个非类化合物 ,分别鉴定为 :大黄酚、大黄素甲醚、β-谷甾醇、大黄素、胡萝卜苷、大黄酸、大黄素甲醚 -8-O-β-D-葡萄糖苷、rheumin、没食子酸、d-儿茶素、蔗糖、1,6-二没食子酰 -O-β-D-吡喃葡萄糖、1-O-β-D-没食子酰吡喃葡萄糖。结论　首次证明华北大黄含有少量大黄酸 ;除大黄酚、大黄素甲醚、大黄素外 ,其他成分均为首次从该植物中分得     

聚乙二醇修饰水蛭素的分离纯化与活性分析

用活化的聚乙二醇(PEG)对水蛭素进行化学修饰，质谱分析表明，修饰产物是修饰度不同的分子质量相差5000u的水蛭素的混合物。用离子交换层析和凝胶过滤层析对反应产物进行分离纯化，离子交换层析难以达到完全分离；而采用凝胶过滤层析方法可较好地分离修饰度不同的各组份。生物学活性分析表明，不同的PEG修饰度对水蛭素的活性具有显著影响，连接1～2个PEG的水蛭素保持了原有活性，而连接3个PEG的水蛭素活性显著下降。     

Mg, Mn, and Co ions enhance the formation of Entamoeba histolytica cyst-like structures resistant to sodium dodecyl sulfate

BACKGROUND: Entamoeba histolytica forms cyst-like structures (CLS) in PEHPS but not in TYS-33 medium. Sodium dodecyl sulfate [(SDS (0.1%)] dissolves most of them in 10 min, but not natural cysts. Chitin is responsible mainly for cyst wall resistance. Its synthesis depends on Mg(2)+, Mn(2)+, or Co(2)+, whose action is interactive. With the aid of the Simplex method, we analyzed the effect of 20 blends of these cations to find the one that, when added to PEHPS, produced the highest proportion of CLS resistant to 1% SDS (RCLS). METHODS: The concentration of Mg(2)+, Mn(2)+, and Co(2)+ was determined in PEHPS and TYI-S-33 with a flame atomic absorption spectrometer. The proportion of RCLS produced in PEHPS with each ion blend was tested. The CLS and RCLS affinity to fluorescein wheat germ agglutinin (WGA/FITC), which binds chitin, was determined. RESULTS: PEHPS contained a similar concentration of Co(2)+ (0.52 microM) and 3.4 and 1.6 times more Mg(2)+ (798 microM) and Mn(2)+ (3.15 microM) than TYI-S-33, respectively. The proportion of RCLS increased gradually in PEHPS until reaching 3.6 +/- 1.43% with MgCl(2) 1.22 mM, MnCl(2) 14.44 mM, and CoCl(2) 19.44 mM (ion blend No. 20). Both CLS and RCLS bound WGA/FITC. The RCLS formed in the presence of ion blend No. 20 appeared wrinkled. CONCLUSIONS: Mg(2)+, Mn(2)+, and Co(2)+ enhanced the ability of PEHPS to form RCLS, possibly because these ions stimulated their chitin synthesis. Although ion blend No. 20 produced the highest proportion of RCLS, this high ion concentration may be toxic for encysting amebas.     

Inhibitory Effect of Pulmonary Carcinoma by Adenovirus-Mediated CD/UPRT Gene

The cell killing effects and bystander effects of double suicide gene on pulmonary carcinoma cells were explored. Lung adenocarcinoma cells (A549) were transfected with different liters of adenovirus vector and followed with different concentrations of 5-FC after a recombinant adenovirus vector carrying CD/UPRT gene (Ad-CD/UPRT) was constructed. The cell viability was measured by MTT assay 4 days later. The cell viability was dropped to 30.57 %-8.62 % after 10 MOI of Ad-CD/UPRT transfected and 5-FC (10-1000μg/mL) administration. Furthermore, Ad-CD/UPRT-infected A549 cells showed a profound neighbor cell killing effect in the same methods. These results suggested that Ad-CD/UPRT/5-FC system can effectively suppress growth of lung adenocarcinoma cells, which may provide a novel and powerful candidate for lung cancer gene therapy strategies.