An inbred 129SvEv GFPCre transgenic mouse that deletes loxP-flanked genes in all tissues

A common method for generating mice with subtle genetic manipulations uses homologous recombination (HR) in embryonic stem (ES) cells to replace a wild-type gene with a slightly modified one. Generally, a drug resistance gene is inserted with the modified gene to select correctly targeted clones. Often, however, the presence of this drug resistance gene interferes with the normal locus and creates a null or hypomorphic allele. Flanking of the selectable marker by loxP sites followed by Cre-mediated deletion after drug selection can overcome this problem. The simplest method used to remove a loxP-flanked selectable marker is to breed an animal carrying a loxP-flanked drug resistance gene to an animal that expresses Cre recombinase in the germline. To date only outbred transgenic mice are available for this purpose. This can be problematic for phenotypic analysis in many organ systems, including the brain, and for the analysis of behavior. While attempting to make 129S6/SvEvTac inbred background (isogenic to our ES cells) mice that express Cre under the control of several tissue-specific promoters, we serendipitously generated a line that excises loxP-flanked drug resistance genes in all tissues, including the germline. This reagent allows deletion of loxP-flanked sequences while maintaining the mutation on an inbred background.     

Cardiomyocyte-specific endothelin A receptor knockout mice have normal cardiac function and an unaltered hypertrophic response to angiotensin II and isoproterenol

Even though endothelin is recognized as an important vasoregulatory molecule, the roles of endothelin receptors in specific cell types are not yet fully understood. Mice with a null mutation in endothelin A receptor gene (ET(A)) or in the gene of its ligand (endothelin 1) die neonatally due to craniofacial and cardiac abnormalities. This early lethality has in the past hindered studies on the role of endothelin in cardiovascular physiology and pathophysiology. To overcome this obstacle, we utilized the cre/loxP technology to generate mice in which the ET(A) gene could be deleted specifically in cardiomyocytes. The cre recombinase transgene driven by the alpha-myosin heavy-chain promoter deleted the floxed ET(A) allele specifically in the hearts of these mice, resulting in a 78% reduction in cardiac ET(A) mRNA level compared to wild-type controls. Cardiomyocyte-specific ET(A) knockout animals are viable and exhibit normal growth, cardiac anatomy, and cardiac contractility, as assessed by echocardiography. In addition, these animals exhibit hypertrophic and contractile responses to 10-day infusion of angiotensin II or isoproterenol similar to those observed in control animals. These results indicate that in adult mice cardiac ET(A) receptors are not necessary for either baseline cardiac function or stress-induced response to angiotensin II or isoproterenol.     

Exploration of structural features of monomeric helical peptides designed with a genetic algorithm

A genetic algorithm (GA)-based strategy to dissect the determinants of peptide folding into alpha-helix was developed. The structural information of helical peptides was obtained with respect to patterns of sequence variability. In many previously reported studies the intrinsic alpha-helical propensities of amino acids although sequence-dependent are apparently independent of the amino acid position. In this research, monomeric helical peptides selected from possible sequences produced by a GA-chemical synthesis were analyzed to identify possible influential structural features. These hexadeca-peptides were obtained after four successive generations. A total of 128 synthetic peptides were evaluated via circular dichroism (CD) measurements in aqueous solution, while the mean ellipticity at 222 nm confirmed the monomeric state of the peptides. The results presented here show that our GA-based strategy may be useful in the design of proteins with increased alpha-helix content.     

Discovery and biological evaluation of potent dual ErbB-2/EGFR tyrosine kinase inhibitors: 6-thiazolylquinazolines

We have identified a novel class of 6-thiazolylquinazolines as potent and selective inhibitors of both ErbB-2 and EGFR tyrosine kinase activity, with IC(50) values in the nanomolar range. These compounds inhibited the growth of both EGFR (HN5) and ErbB-2 (BT474) over-expressing human tumor cell lines in vitro. Using xenograft models of the same cell lines, we found that the compounds given orally inhibited in vivo tumor growth significantly compared with control animals.     

Differential regulation of glutamate receptors in Alzheimer's disease

Selective neurodegeneration is a prominent feature in Alzheimer's disease; however, the mechanism of neuronal death is still unclear. Nonetheless, the topographical distribution of different types of receptors is thought to contribute to the regional selective nature of neuronal degeneration. Specifically, since glutamatergic transmission is severely altered by the early degeneration of cortico-cortical connections and hippocampal projections in Alzheimer's disease, we suspect that glutamate receptors may play a new role in the pathophysiology of disease. Here we review the salient aspects of glutamate receptor expression in Alzheimer's disease and how their differential regulation can contribute to the selective neurodegeneration seen in the disease. Additionally, we assess the potential therapeutic value of glutamate receptors as a target for drug intervention in Alzheimer's disease.     

活细胞染色体切割（光刀）和光捕捉（光钳）的研究

本文报道了光捕捉活细胞染色体的最新实验结果。对ＰＴＫ＿２有丝分裂细胞的染色体先围激光刀切割,再用光钳捕捉使该切割的染色体片断的行为发生改变。光捕捉中期切割的染色体片断有可能使它们整合到同一个子细胞中或丢失在分裂沟中。光捕捉后期切割的染色体可使该切割片断或掺入相反的细胞中或丢失在分裂沟中或回到原有的相应子细胞中。光捕捉操纵染色体去水螈肺上支子细胞中不仅同样有效,还可以在纺缍体的边缘,即纺缍体和间丝笼之间的细胞质清澈区域内用光钳操纵染色体片断移动,旋转。根据细胞和染色体形态和行为,对７００－８４０ｎｍ波长范围内的各种波长的光捕捉进行了比较,结果表明,７００ｎｍ或８００－８２０ｎｍ波长操纵的细胞,出现最少的异常细胞百分率,７６０ｎｍ则诱发百分之百的异常细胞率。根据各方面的综合比较,７００ｎｍ为最佳波长,共次为１０６０和８００ｎｍ。７６０ｎｍ损伤细胞最严重,应避免使用。文中并讨论了光捕捉染色体的应用前景。     

Controlled comparison of cimetidine and carbenoxolone sodium in gastric ulcer.

Fifty-four outpatients with endoscopically diagnosed benign gastric ulcer were allocated at random to treatment with either cimetidine 800 mg daily for six weeks or carbenoxolone sodium 300 mg daily for one week then 150 mg daily for five weeks. Ulcers were reassessed by endoscopy at the end of the trial. The endoscopist was unaware of the treatment and did not take part in the clinical care of the patients. Twenty-one of the 27 patients (78%) given cimetidine and 14 of the 27 (52%) given carbenoxolone had healed ulcers. Symptomatic response occurred earlier with cimetidine but was not significantly better. Unwanted effects were more common in the carbenoxolone group: 12 patients developed hypokalaemia, four of whom needed oral potassium supplements. The results suggest that histamine H2-receptor blockade is at least as effective as carbenoxolone sodium for benign gastric ulcer and produces fewer side effects.     

Noncompetitive Inhibition of N-Methyl-D-Aspartate by Conantokin-G: Evidence for an Allosteric Interaction at Polyamine Sites

Conantokins T and G are polypeptide toxins present in snails of the genus Conus. These substances were recently reported to act as N-methyl-D-aspartate (NMDA) antagonists. In the present study, we examined the possible mechanisms producing this antagonism. Conantokin-G inhibited spermine- and spermidine-stimulated [3H]MK-801 binding to extensively washed rat forebrain membranes in a noncompetitive manner with IC50 values of approximately 507 and approximately 946 nM, respectively. In contrast, glutamate-enhanced [3H]MK-801 binding was unaffected by conantokin-G concentrations of less than or equal to 20 microM. At concentrations greater than or equal to 5 microM, conantokin-G effected a modest, noncompetitive inhibition of glycine-stimulated [3H]MK-801 binding and also produced a small enhancement of basal [3H]MK-801 binding. Conantokin-G reduced (IC50 approximately 1.08 microM) the NMDA-stimulated accumulation of cyclic GMP in cerebellar granule cell cultures to basal values, but did not affect kainate-mediated increases in cyclic GMP. These findings indicate that conantokin-G acts as a noncompetitive NMDA antagonist through an allosteric inhibition of polyamine responses. The neurochemical profile of this polypeptide is distinct from previously described noncompetitive NMDA antagonists.     

The genetic mapping of the OXI3 mitochondrial region

Sixteen mit- mutations in the OXI3 region which specifies in Saccharomyces cerevisiae the subunit I of cytochrome oxidase, were ordered by means of deletion mapping and recombination frequency procedures. These results allowed to distinguish a group of mutants with large overlapping deletions. In one of the analyzed mutants the whole investigated segments is deleted.