Regulatory mechanisms for 3′-end alternative splicing and polyadenylation of the Glial Fibrillary Acidic Protein, GFAP, transcript

The glial fibrillary acidic protein, GFAP, forms the intermediate cytoskeleton in cells of the glial lineage. Besides the common GFAPα transcript, the GFAPε and GFAPκ transcripts are generated by alternative mRNA 3′-end processing. Here we use a GFAP minigene to characterize molecular mechanisms participating in alternative GFAP expression. Usage of a polyadenylation signal within the alternatively spliced exon 7a is essential to generate the GFAPκ and GFAPκ transcripts. The GFAPκ mRNA is distinct from GFAPε mRNA given that it also includes intron 7a. Polyadenylation at the exon 7a site is stimulated by the upstream splice site. Moreover, exon 7a splice enhancer motifs supported both exon 7a splicing and polyadenylation. SR proteins increased the usage of the exon 7a polyadenylation signal but not the exon 7a splicing, whereas the polypyrimidine tract binding (PTB) protein enhanced both exon 7a polyadenylation and exon 7a splicing. Finally, increasing transcription by the VP16 trans-activator did not affect the frequency of use of the exon 7a polyadenylation signal whereas the exon 7a splicing frequency was decreased. Our data suggest a model with the selection of the exon 7a polyadenylation site being the essential and primary event for regulating GFAP alternative processing

High rate of per oral mecillinam treatment failure in community-acquired urinary tract infections caused by ESBL-producing escherichia coli

A population-based study was performed to investigate the efficacy of mecillinam treatment of community-acquired urinary tract infections (CA-UTI) caused by extended-spectrum ß-lactamase (ESBL) producing Escherichia coli. The study was conducted in South-Eastern Norway. Data from patients with CA-UTI caused by ESBL-producing and non-producing (random controls) E. coli were collected through interviews, questionnaires, medical records and the Norwegian Prescription Database. Treatment failure was defined as a new antibiotic prescription appropriate for UTI prescribed within two weeks after the initial antimicrobial therapy. Multivariable logistic regression analysis was performed to identify treatment agents and patient- or bacterial traits associated with treatment failure. A total of 343 patients (mean age 59) were included, of which 158 (46%) were treated with mecillinam. Eighty-one patients (24%, mean age 54) had infections caused by ESBL producing E. coli, and 41 of these patients (51%) received mecillinam as the primary treatment. Mecillinam treatment failure was observed in 18 (44%) of patients infected by ESBL-producing strains and in 16 (14%) of patients with a CA-UTI caused by ESBL non-producing strains. Multivariable analysis showed that ESBL status (odds ratio (OR) 3.2, 95% confidence interval (CI) 1.3–7.8, p = 0.009) and increased MIC of mecillinam (OR 2.0 for each doubling value of MIC, CI 1.4–3.0, p<0.001) were independently associated with mecillinam treatment failure. This study showed a high rate of mecillinam treatment failure in CA-UTIs caused by ESBL producing E. coli. The high failure rate could not be explained by the increased MIC of mecillinam alone. Further studies addressing the use of mecillinam against ESBL-producing E. coli, with emphasis on optimal dosing and combination therapy with ß-lactamase inhibitors, are warranted

Examining the homeostatic distribution of metals and Zn isotopes in Göttingen minipigs

International audienceThe role of metals in biologic systems is manifold, and understanding their behaviour in bodily processes, especially those relating to neurodegenerative diseases, is at the forefront of medical science. The function(s) of metals - such as the transition metals - and their utility in both the diagnosis and treatment of diseases in human beings, is often examined via the characterization of their distribution in animal models, with porcine models considered exceptional proxies for human physiology. To this end, we have investigated the homeostatic distribution of numerous metals (Mg, K, Ca, Mn, Fe, Cu, Zn, Rb and Mo), the non-metal P, and Zn isotopes in the organs and blood (red blood cells, plasma) of Gottingen minipigs. These results represent the first set of data outlining the homeostatic distribution of metals and Zn isotopes in Gottingen minipigs, and indicate a relatively homogeneous distribution of alkali/alkaline earth metals and P among the organs, with generally lower levels in the blood, while indicating more heterogeneous and systematic abundance patterns for transition metals. In general, the distribution of all elements analysed is similar to that found in humans. Our elemental abundance data, together with data reported for humans in the literature, suggest that element-to-element ratios, e.g. Cu/Mg, show potential as simple diagnostics for diseases such as Alzheimer's. Isotopic data indicate a heterogeneous distribution of Zn isotopes among the organs and blood, with the liver, heart and brain being the most depleted in heavy Zn isotopes, and the blood the most enriched, consistent with observations in other animal models and humans. The Zn isotopic composition of Gottingen minipigs displays a systematic offset towards lighter delta Zn-66 values relative to mice and sheep models, suggesting physiology that is more closely aligned with that of humans. Cumulatively, these observations strongly suggest that Gottingen minipigs are an excellent animal model for translational research involving metals, and these data provide a strong foundation for future research

Fibrillary Acidic Protein, GFAP, transcript

Glia

Longitudinal biometal accumulation and Ca isotope composition of the Göttingen minipig brain

Biometals play a critical role in both the healthy and diseased brain's functioning. They accumulate in the normal aging brain, and are inherent to neurodegenerative disorders and their associated pathologies. A prominent example of this is the brain accumulation of metals such as Ca, Fe and Cu (and more ambiguously, Zn) associated with Alzheimer's disease (AD). The natural stable isotope compositions of such metals have also shown utility in constraining biological mechanisms, and in differentiating between healthy and diseased states, sometimes prior to conventional methods. Here we have detailed the distribution of the biologically relevant elements Mg, P, K, Ca, Fe, Cu and Zn in brain regions of Göttingen minipigs ranging in age from three months to nearly six years, including control animals and both a single- and double-transgenic model of AD (PS1, APP/PS1). Moreover, we have characterized the Ca isotope composition of the brain for the first time. Concentration data track rises in brain biometals with age, namely for Fe and Cu, as observed in the normal ageing brain and in AD, and biometal data point to increased soluble amyloid beta (Aβ) load prior to AD plaque identification via brain imaging. Calcium isotope results define the brain as the isotopically lightest permanent reservoir in the body, indicating that brain Ca dyshomeostasis may induce measurable isotopic disturbances in accessible downstream reservoirs such as biofluids