Antisense Smad2 inhibits high glucose-stimulated production of extracellular matrix in cultured rat glomerular mesangial cells

Transforminggrowthfactor-β1(TGF-β1)is amultifunctionalpolypeptidethatregulatesanum-berofcellularprocesses,includingcellprolifera-tion,differentiation,apoptosis,migration,matrix synthesis,andtheimmuneresponse[1,2].Inchron-icrenaldiseases,TGF-β1isakeymediatorofex-tracellularmatrix(ECM)accumulation[3].Oneof thetargetrenalcellsforTGF-β1isglomerular mesangialcellsthatarecapableofproducingcom-ponentsofECM,suchascollagens,lamininand fibronectin[4,5].Recentstudiesindicatedthatinhi-bitionofT…     

Lines of Murine Oligodendroglial Precursor Cells Immortalized by an Activated neu Tyrosine Kinase Show Distinct Degrees of Interaction with Axons In Vitro and In Vivo

Replication-defective retroviruses expressing the t- neu oncogene, or a hybrid protein with the neu tyrosine kinase linked to the external region of the human epidermal growth factor receptor ( egfr-neu ), were used to establish lines of murine oligodendroglial precursor cells. Differentiation of the t- neu lines into myelin-associated glycoprotein (MAG)-positive oligodendrocytes was induced by dibutyryl cAMP, and the egfr-neu line showed limited differentiation in vitro upon withdrawal of epidermal growth factor. Cerebellar granule cell neurons expressed mitogens for the cell lines. Upon transplantation into demyelinated lesions, t- neu line cells engaged with the demyelinated axons whereas the egfr-neu line cells differentiated further and ensheathed the axons. These cell lines thus interact with neurons in vitro and in vivo and can be used as tools to define the molecules involved in different stages of neuron-glia interaction.     

转人GM—CSF基因人膀胱癌细胞株的建立及体外生物学特性

经逆转录病毒载体将人GM-CSF基因导入人膀胱癌细胞株BIU-87细胞中,建立了转基因细胞株BIU/GM。经流式细胞仪行细胞DNA周期分析表明GM-CSF基因的导人及表达对BIU-87细胞的增长无影响。免疫荧光测定发现转GM-CSF基因及表达不能促进BIU-87细胞表面HLA-ABC、DR、DQ抗原的表达。转基因瘤细胞株经6000rad X射线照射灭活后,丧失增殖能力,逐步死亡,但能维持一定水平的GM-CSF分泌活性达两周以上。从而为制备灭活的转基因瘤苗提供了初步经验。     

人IL-10基因重组复制缺陷型腺病毒DNA的构建

目的 构建人IL-10基因的重组腺病毒并进行体外表达和检测。方法从质粒pcDNA3IL-10的CMV启动子下游完整切下IL-10 cDNA的片段,将其定向插入Gateway载体,在克隆酶的作用下将阳性克隆质粒转入目的腺病毒载体。PacI酶线性化IL-10腺病毒DNA后阳离子质脂体转染293A细胞,获得人IL-10的复制缺陷型重组腺病毒。体外感染人胰腺癌细胞株Bxpc-3和大鼠胰腺细胞株AR-42J;Western blot检测人IL-10的表达。结果 成功地构建人IL-10重组腺病毒,病毒滴度达2×109PFU/mL。体外感染的细胞株均检测到IL-10的表达。结论 构建复制缺陷型重组腺病毒能够介导IL-10的基因表达,为细胞因子的抗炎冶疗奠定实验基础。     

Inverting a Laplacian topography map

Summary There are some occasions when it is better to use the Laplacian of the scalp voltages (e.g., for topographic displays and dipole source localization) and other occasions when it is better to use the original voltages (e.g., for comparison to non-Laplacian topography maps). This paper presents a simple algorithm for going between these two representations of the data. The inverse Laplacian involves iterative feedback. An Excel spreadsheet implementation of the algorithm is presented.This research was supported by grant AFOSR 89-0238 from the Air Force Office of Sponsored Research.     

基质金属蛋白酶-2C端片段pex的克隆、测序及真核表达载体的构建

目的 :克隆小鼠基质金属蛋白酶 - 2 ( MMP- 2 ) C端片断 pex编码区的 c DNA序列 ,并构建其真核表达载体 ,为进一步研究其抑制肿瘤作用奠定基础。方法 :根据基因 Gen Bank中 MMP- 2基因的碱基序列 ,利用 RT- PCR的方法从 NIH3T3细胞中分别扩增信号肽及目的基因 pex,然后用多重 PCR的方法将两个片段拼接起来 ,再将拼接起来的目的片段 sig- pex与 p MD1 8T载体连接作全自动测序 ,利用亚克隆的方法将 sig- pex c DNA片段克隆到 pc DNA3.1载体中。结果 :DNA测序证实该片断序列与文献报道完全一致。结论 :成功构建出 pc DNA3.1 - sig- pex真核表达载体     

Construction of Shuttle Expression Plasmid and Stable Expression of Foreign Gene in Mycobacteria and E．Coli

ＣｏｎｓｔｒｕｃｔｉｏｎｏｆＳｈｕｔｔｌｅＥｘｐｒｅｓｓｉｏｎＰｌａｓｍｉｄａｎｄＳｔａｂｌｅＥｘｐｒｅｓｓｉｏｎｏｆ　ＦｏｒｅｉｇｎＧｅｎｅｉｎＭｙｃｏｂａｃｔｅｒｉａａｎｄＥ．ＣｏｌｉＨＵＡＮＧＦＵＹｏｎｇ－ｍｕ（皇甫永修）；ＺＨＡＮＧＤａ－ｊｕ...     

赖型钩体flaB2与VR1012中的CpG基序分析

目的：对问号赖型钩端螺旋体（赖型钩体）DNA疫苗[包括内鞭毛蛋白基因（flaB2)和质粒DNA表达载体（VR1012）]的CpG基序（CpG motifs)进行分析，为DNA疫苗免疫机制的阐明和提高DNA疫苗的效能奠定基础。方法：以flaB2与VR1012构建重组DNA的免疫原，对flaB2及VR1012全核苷酸序列进行计算机分析（分类、计数和定位）。结果：CpG的“C”的侧翼为两个嘌呤，“G”的侧翼为两个嘧啶，在flaB2中共3个，分别为GACGCT，GACGTC和GACGCC；在VR1012中共11个，分别为GACGTC1个，GACGCT2个，GACGCC1个，GACGTT1个，GGCGTT2个，GGCGCT2个，GGCGCC1个，AACGCT1个，其中特别重要的TGACGTCA4个和TAACGCCA有1个，位于5'端456－463；509－516；592－599；778－785和486－493；4个TGACGTCA和1个TAACGCCA均位于5'端且相对集中。结论：赖型钩体flaB2与VR1012构成的DNA疫苗含有TGACGTCA等CpG，这些基序又称免疫刺激序列，构成了DNA疫苗中的佐剂。     

腺病毒载体介导CTLA4Ig基因治疗小鼠变应性鼻炎

目的 检测CTLA4Ig-重组腺病毒载体（Ad-CTLA4Ig）在变应性鼻炎治疗中的作用。方法 采用卵清蛋白（OVA）致敏和激发诱导小鼠变应性鼻炎。实验组在OVA激发前30min予以Ad-CTA4Ig腹腔注射。未转CTLA4IgcDNA的腺病毒载体（Adv）作对照。比较各组动物鼻部症状和鼻粘膜形态学改变，并采用ELISA法测定血清中OVA－特异性IgE水平。结果 CTLA4Ig重组腺病毒实验组小鼠鼻部症状和鼻粘膜病理改变不明显，并且血清中OVA－特异性IgE水平明显低于对照组（P＜0.05）。结论 Ad-CTLA4Ig可防治小鼠应变性鼻炎的发生，提示CTLA4Ig-重组腺病毒载体有可能运用于临床变应性鼻炎的治疗。     

人白细胞介素7基因克隆及其真核表达载体的构建

目的：构建人白细胞介素7（hIL-7）的真核表达载体。方法：从人脾脏组织的总RNA中，用RT－PCR方法扩增出编码成熟hIL-7的cDNA，将其克隆于pMD18-T质粒中，并进行序列测定。再将hIL-7cDNA以正义及反义插入真核、原核双重表达载体pBK-CMV质粒，转化至大肠杆菌DH5α。最后将表达的lacz-hIL-7重组蛋白行SDS－PAGE电泳，并作考马斯亮蓝染色分析，western blot鉴定。结果：hIL-7序列测定结果与预期一致。pBK-CMV-hIL-7（正义插入）的无隆表达重组蛋白，western blot证实此蛋白为IL－7。结论：成功构建了hIL-7的真核表达载体，为进一步研究hIL-7的抗肿瘤作用创造了条件。