In vivo gene transfer: focus on the kidney

Renal gene transfer techniques are being developed as a novelexperimental approach to understand the pathogenesis of renaldisease and to potentially develop new therapeutic tools. Wereview the currently available technology to introduce foreigngenetic material into renal tissue, i.e., retroviral, adenoviral,and liposomal transfer systems with their respective advantagesand caveats. Today, the transfer efficiency of these methodsappears to be sufficiently high to study the effects of transducedgenes on renal function and morphology in rat kidney. This willallow (i) the elucidation of the function of genes on the courseof renal disease in experimental animal models and (ii) themodulation of local expression of endogenous genes which presumptivelycontribute to renal pathology in these models. One strategyto accomplish this aim is the use of recombinant DNA technologyto design antisense DNA constructs or oligonucleotides, whichinterfere with the renal expression of target genes. We willalso discuss some of the shortcomings of the currently usedtechniques with respect to potential therapeutic use of genetransfer systems and gene modulation.     

Comparison of hprt and lacI mutant frequency with DNA adduct formation in N-hydroxy-2-acetylaminofluorene-treated Big Blue rats

N-Hydroxy-2-acetylaminofluorene (N-OH-AAF) is the proximate carcinogenic metabolite of the powerful rat liver carcinogen 2-acetylaminofluorene. In this study, transgenic Big Blue(R) rats were used to examine the relationship between in vivo mutagenicity and DNA adduct formation by N-OH-AAF in the target liver compared with that in nontarget tissues. Male rats were given one, two, or four doses of 25 mg N-OH-AAF/kg body weight by i.p. injection at 4-day intervals, and groups of treated and control rats were euthanized up to 10 weeks after beginning the dosing. Mutant frequencies were measured in the spleen lymphocyte hprt gene, and lacI mutant frequencies were determined in the liver and spleen lymphocytes. At 6 weeks after beginning the dosing, the hprt mutant frequency in spleen lymphocytes from the four-dose group was 16.5 x 10(-6) compared with 3.2 x 10(-6) in control animals. Also at 6 weeks, rats given one, two, or four doses of N-OH-AAF had lacI mutant frequencies in the liver of 97.6, 155.6, and 406.8 x 10(-6), respectively, compared with a control frequency of 25.7 x 10(-6); rats given four doses had lacI mutant frequencies in spleen lymphocytes of 55.8 x 10(-6) compared with a control frequency of 20.4 x 10(-6). Additional rats were evaluated for DNA adduct formation in the liver, spleen lymphocytes, and bone marrow by (32)P-postlabeling. Adduct analysis was conducted 1 day after one, two, and four treatments with N-OH-AAF, 5 days after one treatment, and 9 days after two treatments. N-(Deoxyguanosin-8-yl)-2-aminofluorene was the major DNA adduct identified in all the tissues examined. Adduct concentrations increased with total dose to maximum values in samples taken 1 day after two doses, and remained essentially the same after four doses. In samples taken after four doses, adduct levels were 103, 28, and 7 fmol/microg of DNA in liver, spleen lymphocytes, and bone marrow, respectively. The results indicate that the extent of both DNA adduct formation and mutant induction correlates with the organ specificity for N-OH-AAF carcinogenesis in the rat. Environ. Mol. Mutagen. 37:195-202, 2001. Published 2001 Wiley-Liss, Inc.     

APP转基因小鼠脑内单胺类递质的变化

用HPLC－ECD法对APP695,751V717Ⅰ转基因模型小鼠大脑皮层神经递质进行检测,结果显示APP751转基因鼠大脑皮层中的5－HIAA一与对照组比较有显著性差异。APP695转基因鼠大脑上以层中DA和5－HT平均含量分别比对照鼠升高38.9％和45.8%。提示Aβ的形成可能影响了多巴胺系统和5－羟色胺系统。     

口服转内皮抑素基因双歧杆菌制剂对肺癌的疗效

目的：探讨转人内皮抑素基因双歧杆菌口服制剂对肺癌的治疗效果。方法：选择肺癌患者118例,比较服用转内皮抑素基因双歧杆菌口服制剂前后患者症状和体征,影像学检查（X线、CT,MRI）,细胞学检查,纤维支气管镜检查及病理活检、免疫学检查（CEA）和生活质量等方面的变化。结果：服用转内皮抑素基因双歧杆菌口服制剂后患者症状和体征及生活质量等较服用前均有好转或明显好转,肺癌细胞生长被明显抑制。结论：转内皮抑素基因双歧杆菌口服制剂对肺癌有较好的治疗效果。     

Spontaneous mutation of the lacI transgene in rodents: absence of species, strain, and insertion-site influence

Comparison of spontaneous mutation spectra derived from different transgenic constructs can provide valuable insights for interpreting the mechanisms of spontaneous mutation. In this study, spontaneous mutation frequencies and spectra of the lacI transgene are compared in the liver of C57BL/6, B6C3F1, and BC-1 mice and F344 rats. Before correction for clonal expansion, the mutant frequency varied from 2.6 +/- 0.45 to 5.0 +/- 2.4 x 10(-5). Correction for potential clonal expansion reduced the range in mutation frequency to between 2.3 +/- 0.45 and 3.5 +/- 2.0 x 10(-5). There is thus no statistical difference in spontaneous mutation frequency between the different strains and species. G:C --> A:T transitions and to a lesser extent, G:C --> T:A transversions dominate the mutational spectra in all of these animals. In three strains of mice, G:C --> A:T transitions account for 50.7-53.3% of mutation, 81.7-83.8% of which involve CpG sites, whereas G:C --> T:A transversions account for 17.8-32.9% of mutations with 43.2-50.0% found at CpG sites. In rats, G:C --> A:T transitions account for 38.0% of the spectra, 70.0% of which involve CpG sites, whereas G:C --> T:A transversions account for 23.0% of the spectra, 70.0% of which involve CpG sites. The distribution of other classes of mutations is also very similar. We conclude that, despite reports about species and strain differences in induced mutation, spontaneous mutations in the lacI transgene appear to be similar, regardless of genomic location, rodent strain, or species. In addition to insights into spontaneous mutation, this study also provides essential data for comparison with and interpretation of induced mutations.     

A novel brain receptor is expressed in a distinct population of olfactory sensory neurons

Three novel G-protein-coupled receptor genes related to the previously described RA1c gene have been isolated from the mouse genome. Expression of these genes has been detected in distinct areas of the brain and also in the olfactory epithelium of the nose. Developmental studies revealed a differential onset of expression: in the brain at embryonic stage 17, in the olfactory system at stage E12. In order to determine which cell type in the olfactory epithelium expresses this unique receptor type, a transgenic approach was employed which allowed a coexpression of histological markers together with the receptor and thus visualization of the appropriate cell population. It was found that the receptor-expressing cells were located very close to the basal membrane of the epithelium; however, the cells extended a dendritic process to the epithelial surface and their axons projected into the main olfactory bulb where they converged onto two or three glomeruli in the dorsal and posterior region of the bulb. Thus, these data provide evidence that this unique type of receptor is expressed in mature olfactory neurons and suggests that it may be involved in the detection of special odour molecules.     

胆固醇酯转移蛋白转基因家兔制作及生物学特性分析

摘要: 目的 制作肝脏特异性高表达人胆固醇酯转移蛋白（CETP）转基因家兔并对其生物学特性分析进行分析。方法 利用显微注射方法制作CETP转基因家兔,利用Western Blot、Real-time PCR和CETP活性鉴定试剂盒鉴定模型家兔中人CETP转基因的表达和活性。结果 PCR检验结果显示我们成功获得CETP转基因家兔,转基因主要在肝脏特异性表达,其血浆CETP活性相对于非转基因家兔明显升高。结论 本研究首次成功建立了高表达人CETP的转基因家兔,为研究CETP功能和其参与心血管疾病的机制提供了良好模型。