Transgenic rat model of Huntington's disease

Huntington's disease (HD) is a late manifesting neurodegenerative disorder in humans caused by an expansion of a CAG trinucleotide repeat of more than 39 units in a gene of unknown function. Several mouse models have been reported which show rapid progression of a phenotype leading to death within 3-5 months (transgenic models) resembling the rare juvenile course of HD (Westphal variant) or which do not present with any symptoms (knock-in mice). Owing to the small size of the brain, mice are not suitable for repetitive in vivo imaging studies. Also, rapid progression of the disease in the transgenic models limits their usefulness for neurotransplantation. We therefore generated a rat model transgenic of HD, which carries a truncated huntingtin cDNA fragment with 51 CAG repeats under control of the native rat huntingtin promoter. This is the first transgenic rat model of a neurodegenerative disorder of the brain. These rats exhibit adult-onset neurological phenotypes with reduced anxiety, cognitive impairments, and slowly progressive motor dysfunction as well as typical histopathological alterations in the form of neuronal nuclear inclusions in the brain. As in HD patients, in vivo imaging demonstrates striatal shrinkage in magnetic resonance images and a reduced brain glucose metabolism in high-resolution fluor-deoxy-glucose positron emission tomography studies. This model allows longitudinal in vivo imaging studies and is therefore ideally suited for the evaluation of novel therapeutic approaches such as neurotransplantation.     

Tumor necrosis factor alpha-induced interleukin-8 production via NF-kappaB and phosphatidylinositol 3-kinase/Akt pathways inhibits cell apoptosis in human hepatocytes

Tumor necrosis factor alpha (TNF-alpha) not only induces apoptotic signals but also causes antiapoptotic and regenerative responses in the liver. However, the molecular mechanism(s) of the latter events remains unclear. In the present study, we examined TNF-alpha-induced genes in Hc human normal (unsensitized) hepatocytes by cDNA microarray analysis. Interleukin-8 (IL-8) induction was the most pronounced of the upregulated genes. The IL-8 protein level was also increased. IL-8 belongs to the ELR-CXC chemokine family and appears to exert mitogenic and antiapoptotic functions in other cell systems. IL-8 expression by TNF-alpha was inhibited when two survival signals, nuclear factor kappaB (NF-kappaB) and phosphatidylinositol 3-kinase (PI3K)/Akt, were inhibited by a mutant form of inhibitor of NF-kappaB (IkappaB); by dominant negative (kinase-dead) Akt; or by treatment with LY 294002, an inhibitor of PI3K. TNF-alpha induced apoptosis in Hc cells that were sensitized by inhibition of NF-kappaB and PI3K activation. IL-8 administration protected mice against concanavalin A-induced hepatitis in vivo. IL-8 also rescued the sensitized Hc cells, at least in part, from TNF-alpha-induced apoptosis in vitro. TNF-alpha inhibited DNA synthesis in unsensitized Hc cells in the absence of serum. Exogenous IL-8 reversed, though anti-IL-8 neutralization antibody enhanced, growth inhibition by TNF-alpha. These results indicate that IL-8, the production of which is stimulated by TNF-alpha, inhibits apoptosis of sensitized hepatocytes and releases normal (unsensitized) hepatocytes from growth inhibition induced by TNF-alpha.     

Roseomonas, a new genus associated with bacteremia and other human infections.

In the 1980s, a pink bacterium different from species of the genus Methylobacterium was implicated in human infection. Using biochemical tests and DNA hybridization, we examined 42 strains of pink-pigmented, gram-negative bacteria that were not members of the genus Methylobacterium. The isolates included 6 strains each of CDC "pink coccoid" groups I, II, III, and IV; 10 isolates from Gilardi's "unnamed taxon"; and 8 blood isolates from ill, debilitated, or immunosuppressed patients. The DNA hybridization studies supported the creation of six genomospecies encompassing the 42 strains. Reactions for esculin hydrolysis, glycerol oxidation, and D-mannose oxidation enabled separation of genomospecies 1 through 4. These tests, as well as motility, nitrate reduction, citrate utilization, and oxidation of L-arabinose, D-galactose, and D-xylose, differentiated genomospecies 5 and 6 from each other and from genomospecies 1 through 4. These organisms were susceptible in vitro to the aminoglycosides, tetracycline, and imipenem and generally susceptible to the quinolones. We propose the new genus, Roseomonas, for these bacteria to include three named species, Roseomonas gilardii sp. nov., Roseomonas cervicalis sp. nov., and Roseomonas fauriae sp. nov., and three unnamed genomospecies.     

Providencia rustigianii: a new species in the family Enterobacteriaceae formerly known as Providencia alcalifaciens biogroup 3.

The name Providencia rustigianii sp. nov. is proposed for a group of organisms previously known as Providencia alcalifaciens biogroup 3. By DNA hybridization, strains of P. rustigianii were 81 to 99% related to each other at 60 degrees C, but only 44 to 49% related to P. alcalifaciens biogroups 1 and 2 and 26 to 33% related to Providencia stuartii. P. rustigianii could be differentiated from P. alcalifaciens and P. stuartii by simple biochemical tests. P. rustigianii produced acid from D-galactose but not from trehalose; P. stuartii produced acid from both; and P. alcalifaciens produced acid from neither. P. rustigianii could be distinguished from Providencia rettgeri (formerly Proteus rettgeri) by urea hydrolysis and acid production from D-arabitol; P. rustigianii was negative for these two tests, but P. rettgeri was positive. Strains of P. rustiganii were 32 to 34% related to strains of P. rettgeri. Three of the 11 strains of P. rustigianii were isolated from stools, but the sources of the other isolates are unknown. Three strains (27%) were sensitive to colistin, and 82 to 100% were sensitive to ampicillin, carbenicillin, cephalothin, gentamicin, kanamycin, nalidixic acid, streptomycin, and tetracycline. Strain ATCC 33673 (CDC no. 0132-68) is the type strain for this species.     

Artificial-infection protocols allow immunodetection of novel Borrelia burgdorferi antigens suitable as vaccine candidates against Lyme disease

Vaccination with recombinant outer surface protein A (OspA) from Borrelia burgdorferi provides excellent antibody-mediated protection against challenge with the pathogen in animal models and in humans. However, the bactericidal antibodies are ineffective in the reservoir host, since OspA is expressed by spirochetes only in the vector, but rarely, if at all, in mammals. Using an artificially generated immune serum (anti-10(8) spirochetes) with high protective potential for prophylactic and therapeutic treatment, we have now isolated from an expression library of B. burgdorferi (strain ZS7) three novel genes, zs7.a36, zs7.a66 and zs7.a68. All three genes are located, together with ospA/B, on the linear plasmid lp54, and are expressed in vitro and in ticks. At least temporarily two of them, ZS7.A36 and ZS7.A66, are also expressed during infection. The respective natural antigens are poorly immunogenic ininfected normal mice but elicited antibodies in Lyme disease patients. We show that recombinant preparations of ZS7.A36, ZS7.A66 and ZS7.A68 induce functional antibodies in rabbits capable of protecting immunodeficient mice against subsequent experimental infection. These findings suggest that all three recombinant antigens represent potential candidates for a "second generation" vaccine to prevent and/or cure Lyme disease.     

The Bioperl toolkit: Perl modules for the life sciences

The Bioperl project is an international open-source collaboration of biologists, bioinformaticians, and computer scientists that has evolved over the past 7 yr into the most comprehensive library of Perl modules available for managing and manipulating life-science information. Bioperl provides an easy-to-use, stable, and consistent programming interface for bioinformatics application programmers. The Bioperl modules have been successfully and repeatedly used to reduce otherwise complex tasks to only a few lines of code. The Bioperl object model has been proven to be flexible enough to support enterprise-level applications such as EnsEMBL, while maintaining an easy learning curve for novice Perl programmers. Bioperl is capable of executing analyses and processing results from programs such as BLAST, ClustalW, or the EMBOSS suite. Interoperation with modules written in Python and Java is supported through the evolving BioCORBA bridge. Bioperl provides access to data stores such as GenBank and SwissProt via a flexible series of sequence input/output modules, and to the emerging common sequence data storage format of the Open Bioinformatics Database Access project. This study describes the overall architecture of the toolkit, the problem domains that it addresses, and gives specific examples of how the toolkit can be used to solve common life-sciences problems. We conclude with a discussion of how the open-source nature of the project has contributed to the development effort.     

Influence of scaffold thickness and scaffold composition on bioartificial graft survival

Biological scaffolds exhibit advantageous properties for tissue engineering of small diameter vessels. The influence of their extracellular matrix (ECM) components during in vivo repopulation is unknown. We implanted different xenogenic vascular matrices in a rat model to determine the influence of scaffold-thickness and ECM composition on in vivo repopulation. Decellularized ovine jugular vein (JV, n=42), carotid artery (CA, n=42) and aorta (AO, n=42) were implanted subcutaneously in the neck of adult male rats. Animals were sacrificed 2, 4 and 8 weeks after implantation. Cell and matrix morphology of explanted scaffolds were characterized by hematoxylin-eosin and pentachrome staining. Monoclonal anti-rat-CD31 was used to identify revascularization. Quantification of cell density was done by DNA-isolation.THICKNESS OF IMPLANTED XENOGENIC SCAFFOLDS VARIED ACCORDING TO THE MATERIAL USED (AO: 3.0-3.8mm; CA: 0.7-0.88mm; JV: 0.35-0.61mm). Immunohistology revealed complete repopulation of AO, CA, and JV scaffolds with endothelial cells and myofibroblasts within 2 weeks. After 8 weeks of implantation, AO scaffolds were completely covered by an endothelial monolayer and showed signs of a central matrix degeneration. JV scaffolds were completely degenerated at this stage. In contrast, CA scaffolds showed preserved ECM with a normal myofibroblast population and endothelial cell coverage.     

Further observations on use of atropine in the treatment of myopia

In order to further our observations on the effects of atropine eyedrops for the management of myopia, we conducted a retrospective study of seventy-nine (79) patients, followed over a ten-year period (1971 to 1980). The atropine sulfate drops were used daily in most cases, tapering the frequency in the later teenage years. In general, those children who showed a good initial response during their first year of treatment, continued to use them for several years. Bifocal or reading glasses were used and family acceptance was good. Those children who showed less favorable results in the first year or who had unconcerned parents, stopped the drops within a year or two and went back to glasses or later, contact lenses. The data support the fact that children with low refractive errors may well have "functional myopia," as opposed to the "axial myopia," that characterizes the higher levels of myopia. These low degree myopes are the best candidates for using atropine to reduce or diminish myopia changes.     

Improved high-performance liquid chromatography assay of doxorubicin: Detection of circulating aglycones in human plasma and comparison with thin-layer chromatography

Summary We compared doxorubicin and metabolite pharmacokinetic data obtained from thin-layer chromatography (TLC) and high-performance liquid chromatography (HPLC) assay of plasma samples from six patients who had been treated with doxorubicin. Duplicate 1-ml samples were extracted with chloroform: isopropanol (1:1) and assayed using a sensitive HPLC system incorporating a dual pump gradient with tetrahydrofuran as the mobile phase and fluorescence detection. Duplicate 1-ml samples from the same specimens were assayed using a modification of a previously described TLC assay. Areas under the curve for doxorubicin by HPLC (3.36±2.30 M · h) and TLC (4.16±2.50 M · h) were not significantly different (P=0.5). Terminal half-life of doxorubicin by HPLC (28.0±6.98 h) and TLC (23.2±7.8) (P=0.29) and the calculated total-body clearances by HPLC (0.55±0.29 l/min) and TLC (0.45±0.23) (P=0.55) were not significantly different. Areas under the curve for doxorubicinol by HPLC (2.75±1.4 M · h) and TLC (2.53±7.1 M · h) (P=0.73) showed no significant differences. HPLC detected a mixed 7-deoxydoxorubicinol aglycone-doxorubicin aglycone peak, 7-deoxydoxorubicin aglycone, and two nonpolar, unidentified metabolites. TLC detected the following aglycone metabolites: doxorubicin aglycone, doxorubicinol aglycone, 7-deoxydoxorubicinol aglycone, an unidentified polar metabolite, and several unidentified nonpolar metabolites. From these data we conclude that HPLC and TLC detect concentrations of doxorubicin and doxorubicinol from human plasma equally well to concentrations of 7.0 nM (4 pmol injected doxorubicin). Aglycones do circulate in human plasma at concentrations above the detection limits of both assays. Doxorubicinol aglycone, which is detected by TLC but not by HPLC, may be formed from artifactual breakdown of doxorubicinol during TLC development. Unidentified nonpolar compounds seen on HPLC and TLC may represent further doxorubicin metabolism than previously described.     

Differential diagnosis of acute dactylitis psoriatica

A bulbous or "sausage-shaped" finger may represent a special type of psoriatic arthritis without any rheumatoid factor activity. Dactylitis psoriatica must be suspected in cases of arthralgias progressing along the phalanges of a finger associated with global soft tissue swelling. Acute dactylitis psoriatica is characterized by an intermittent course combined with psoriatic and partly extremely mild or masked skin and nail changes. X-rays findings are spicular protuberances at diaphyses in a "cloudy collar" image. Symptomatic treatment of dactylitis psoriatica consists of non-steroidal antirheumatic drugs or basic gold salt therapy. In the interval additional surgical treatment using arthro-plasties and arthrodeses is indicated in cases of joint destruction. Amputation of the affected finger is not to be considered as a mandatory causal therapy of dactylitis psoriatica.