Role of viral antigens in destructive cellular immune responses to adenovirus vector-transduced cells in mouse lungs.

Adenoviruses missing E1 have been used as gene delivery vectors to the lungs for the treatment of cystic fibrosis. Transient expression of the recombinant gene and the development of inflammation have been two major limitations to the application of first-generation recombinant adenoviruses for gene therapy. Studies with mouse models of liver- and lung-directed gene therapy suggested that CD8+ cytotoxic T lymphocytes (CTLs) are effectors that contribute to extinction of transgene expression. The precise antigens responsible for activation of CTLs have not been identified. In this study, we examine the relative contributions of viral proteins versus the transgene product to the activation of CTLs which eliminate transgene-containing cells in mouse lungs. Instillation of a lacZ-expressing virus into the lungs of C57BL/6 mice elicited CTL responses to both viral proteins and the transgene product, beta-galactosidase, which collectively contribute to loss of trans-gene expression in mouse airways. Similar results were obtained in two experimental models in which the animals should be tolerant to the transgene, i.e., lacZ virus delivered to an animal transgenic for lacZ and a virus expressing the liver-specific enzyme ornithine transcarbamylase administered to the lungs of various strains of immune-competent mice. These data confirm the hypothesis that CTLs specific for viral antigens contribute to the problem of transgene instability in mouse lungs and indicate that CTLs specific for transgene product alone cannot account for the observed problem.     

Identification and characterization of three genes and two pseudogenes on chromosome 13

A study was conducted on the feasibility of isolating genes and pseudogenes that map to chromosome 13 by a hybridization-based approach using a 13-specific library and pools of repeat-free cDNA clones. Five pairs of cDNA and chromosome 13 genomic clones were identified and characterized. Partial or full-length sequence was derived from all cDNAs, and database searches were performed for putative gene identification. Partial sequence was also obtained from the chromosome 13 genomic clones for comparison with those of the hybridizing cDNAs. As a result of these analyses we identified three genes, a putative homologue of a porcine mRNA encoding an unidentified hepatic protein, a putative homologue of a yeast integral membrane protein, and a gene for a translationally controlled tumor protein, and two processed pseudogenes, ribosomal proteins L23a and S3a. The latter was formerly identified as the v-fos transformation effector gene, Fte-1, and recently cited as a possible candidate for the BRCA2 gene on chromosome 13. All genes and pseudogenes were localized to cytogenetic bands by in situ hybridization of metaphase chromosomes with probes derived from the chromosome 13 genomic clones.     

应用MUCAP试剂快速检测沙门氏菌

报告了用4-甲基伞形酮辛酯(4-Methylumbelliferyl-caprylate, MUCAP)快速检测沙门氏菌的特异性、敏感性和实用性。经HE,DHL,SS和麦康凯琼脂平板分离的65株沙门氏菌标准菌株和48株从食品中分离的沙门氏菌,用MUCAP测试均呈阳性反应;394株非沙门氏菌中呈阳性反应的假单胞菌、气单胞菌、邻单胞菌可通过氧化酶试验与沙门氏菌区分开;与粘质沙雷氏菌的交叉反应改用加1％蔗糖的分离平板也可排除。此方法的敏感性和特异性均达到97％以上,而且操作简便、快速,数分钟内即可完成。     

Isolation and expression in Escherichia coli of hepB and hepC, genes coding for the glycosaminoglycan-degrading enzymes heparinase II and heparinase III, respectively, from Flavobacterium heparinum.

Upon induction with heparin, Flavobacterium heparinum synthesizes and secretes into its periplasmic space heparinase I (EC 4.2.2.7), heparinase II, and heparinase III (heparitinase; EC 4.2.2.8). Heparinase I degrades heparin, and heparinase II degrades both heparin and heparan sulfate, while heparinase III degrades heparan sulfate predominantly. We isolated the genes encoding heparinases II and III (designated hepB and hepC, respectively). These genes are not contiguous with each other or with the heparinase I gene (designated hepA). hepB and hepC were found to contain open reading frames of 2,316 and 1,980 bp, respectively. Enzymatic removal of pyroglutamate groups permitted sequence analysis of the amino termini of both mature proteins. It was determined that the mature forms of heparinases II and III contain 746 and 635 amino acids, respectively, and have calculated molecular weights of 84,545 and 73,135, respectively. The preproteins have signal sequences consisting of 26 and 25 amino acids. Truncated hepB and hepC genes were used to produce active, mature heparinases II and III in the cytoplasm of Escherichia coli. When these enzymes were expressed at 37 degrees C, most of each recombinant enzyme was insoluble, and most of the heparinase III protein was degraded. When the two enzymes were expressed at 25 degrees C, they were both present predominantly in a soluble, active form.     

无色素腺棉籽蛋白粉与棉子油的制备

低棉酚棉籽为高蛋白含油的植物资源,适宜于水溶法制备油及蛋白粉,出油率一般可达９５％。     

以融合蛋白形式在大肠杆菌中表达人MCP－1

将编码人单核细胞趋化蛋白－１（ＭＣＰ－１）的基因亚克隆到大肠杆菌表达载体ｐＥＸ３１Ａ中,在大肠杆菌中表达出ＭＳ２／ＭＣＰ－１融合蛋０白,该表达产物约占菌体总蛋白的１５％左右,Ｗｅｓｔｅｒｎｂｌｏｔ检测表明,表达产物可与ＭＣＰ－１抗体特异反应。采用琼脂糖平板法进行活性测定表明,表达产物具有明显的单核细胞趋化活性,说明Ｎ端融合一段细菌蛋白对ＭＣＰ－１有无趋化活性可能没有影响。     

RGDS肽对大鼠主动脉球囊内膜剥脱后血管壁增殖的影响

在大鼠主动脉球囊内膜剥脱术后血管壁细胞过度增殖模型上,用合成的血小板膜纤维蛋白原受体（ｇｌｙｃｏｐｒｏｔｅｉｎⅡｂ／Ⅲａｃｏｍｐｌｅｘ,ＧＰⅡｂ／Ⅲａ）拮抗剂ＲＧＤＳ（Ａｒｇ－Ｇｌｙ－Ａｓｐ－Ｓｅｒ,５０μｍｏｌ·ｋｇ－１·ｄ－１）治疗可有效地抑制损伤血管壁的细胞计数增加和内膜增厚以及血管平滑肌细胞增殖,显著降低其血管组织３Ｈ－ＴｄＲ和３Ｈ－Ｌｅｕ的参入增加程度。实验结果提示ＲＧＤＳ肽作为血管成型术的辅佐剂,对于防治血管再狭窄可能具有潜在的临床应用前景。