Correlation between patterns of DNase I-hypersensitive sites and upstream promoter activity of the human epsilon-globin gene at different stages of erythroid development.

DNA 5' to the human epsilon-globin gene exhibits unique patterns of DNase I-hypersensitive sites (DHS) in three human erythroleukemic cell lines which represent the embryonic (K562), fetal (HEL), and adult (KMOE) stages of erythroid development. We have mapped 10 epsilon-globin DHS in K562 cells, in which the epsilon-globin gene is maximally active. Major sites are located -11.7, -10.5, -6.5, -2.2 kilobase pairs (kbp) and -200 base pairs (bp) upstream of the gene and directly over the major cap site. Minor sites are located -5.5, -4.5, and -1.48 kbp and -900 bp upstream of the cap site. In HEL cells, in which the epsilon-globin gene is expressed at extremely low levels, the -11.7-, -10.5-, -5.5-, -4.5-, and -2.2-kbp DHS are no longer detectable; the -200-bp site is approximately 300-fold less sensitive to DNase I; and the -1.48-kbp, -900-bp, and major cap site DHS are 3- to 4-fold less sensitive. Only the DHS located -6.5 kbp relative to the major cap site is detectable at all three stages of erythroid development, including KMOE cells in which epsilon-globin synthesis is undetectable. We suggest that this site may be implicated in maintaining the entire beta-globin cluster in an active chromatin conformation. The five DHS downstream of the -6.5-kbp element possess associated promoters. Thus two distinct types of DHS exist--promoter positive and promoter negative. In HEL cells, all the upstream promoters are inactivated, although the -1.48-kbp and -900- and -200-bp DHS are still present. This suggests that the maintenance of DHS and regulation of their associated promoters occur by independent mechanisms. The inactivation of the upstream promoters in HEL cells while the major cap site remains active represents a unique pattern of expression and suggests that HEL cells possess regulatory factors which specifically down regulate the epsilon-globin upstream promoters.     

Controlled surface modification of cellulose fibers by amino derivatives using N,N′-carbonyldiimidazole as activator

Surface grafting of different amino derivatives was carried on under mild condition using N,N′-carbonyldiimidazole (CDI) as an activator. The action of a diamine or a triamine on previously activated cellulose fibres proceeds by the reaction of one amine function giving rise to a carbamate derivative. The other terminal amino groups remained available for further reaction. In particular, their activation with CDI generates a reactive carboxamide able to condense with an aliphatic amine through a urea linkage. Evidence for the occurrence of the reaction at each modification step was confirmed by Fourier Transform Infrared Spectroscopy (FTIRS) and X-ray photoelectron spectroscopy (XPS). The contact angle measurement, using water as a probe, was used to explore the evolution of the surface wettability for the different modification sequences. It was shown that the contact angle value is determined by the ratio between polar and methylene groups and by the spatial arrangement of the molecule on the surface.     

Genetics in Osteoarthritis

Osteoarthritis is a degenerative articular disease with complex pathogeny because diverse factors interact causing a process of deterioration of the cartilage. Despite the multifactorial nature of this pathology, from the 50’s it´s known that certain forms of osteoarthritis are related to a strong genetic component. The genetic bases of this disease do not follow the typical patterns of mendelian inheritance and probably they are related to alterations in multiple genes. The identification of a high number of candidate genes to confer susceptibility to the development of the osteoarthritis shows the complex nature of this disease. At the moment, the genetic mechanisms of this disease are not known, however, which seems clear is that expression levels of several genes are altered, and that the inheritance will become a substantial factor in future considerations of diagnosis and treatment of the osteoarthritis.     

Plant species richness in the Cape Verde Islands—eco-geographical determinants

Plant species richness in the Cape Verde archipelago is examined relative to island eco-geographical factors. Species-area and species-area-habitat relationships are analysed using the classical species-area model and the recently proposed species-choros model. The number of floristic zones (used to estimate the choros parameter) provides an adequate estimate of the potential habitat diversity, and the species-choros model achieved a better fit with both total flora and endemic species. In addition to area and habitat diversity, longitude also emerges as an important determinant of species diversity, whereas latitude, minimum distance to the nearest island, and total rural population do not display any correlation. As in other insular ecosystems, the species richness (about 140 per 100 km²) is lower than in nearby mainland regions; the proximity to the desert areas of the Sahel can also be seen as related with this low value. The floristic heterogeneity in Cape Verde is high, as is usual in island ecosystems. In a comparative analysis of the species richness on the different islands (using α-values), Brava stands out as having the highest total flora species densities, while for endemic flora Brava and São Nicolau jointly occupy the leading position. The high diversity for both total and endemic species on Santo Antão, São Vicente, São Nicolau, Fogo and Brava reinforces their importance in conservation terms - in the case of most of them, something that is already recognized in the established network of protected areas.     

Dengue viruses activity in Piauí, Brazil

The present paper reports a laboratory investigation performed between the years of 2000 and 2002 to study a virological surveillance program introduced in the state of Piauí to support an epidemiological survey of the disease. Dengue virus type 3 (DENV-3) existence in the state was detected in May 2002 when a high number of dengue cases due to DENV-1 and DENV-2 were reported. An assessment on the population knowledge about the disease and its transmission showed that almost 50% of the population were still unaware of the epidemiological features of dengue.     

Mitochondrial Dysfunction and Reactive Oxygen Species in Excitotoxicity and Apoptosis: Implications for the Pathogenesis of Neurodegenerative Diseases

In recent years we have witnessed a major interest in the study of the role of mitochondria, not only as ATP producers through oxidative phosphorylation but also as regulators of intracellular Ca²⁺ homeostasis and endogenous producers of reactive oxygen species (ROS). Interestingly, the mitochondria have been also implicated as central executioners of cell death. Increased mitochondrial Ca²⁺ overload as a result of excitotoxicity has been associated with the generation of superoxide and may induce the release of proapoptotic mitochondrial proteins, proceeding through DNA fragmentation/condensation and culminating in cell demise by apoptosis and/or necrosis. In addition, these processes have been implicated in the pathogenesis of many neurodegenerative diseases, which share several features of cell death: selective brain areas undergo neurodegeneration, involving mitochondrial dysfunction (mitochondrial complexes are affected), loss of intracellular Ca²⁺ homeostasis, excitotoxicity, and the extracellular or intracellular accumulation of insoluble protein aggregates in the brain.     

Amyloid beta peptide 1-42 disturbs intracellular calcium homeostasis through activation of GluN2B-containing N-methyl-d-aspartate receptors in cortical cultures

Alzheimer's disease (AD) is a progressive neurodegenerative disorder that leads to debilitating cognitive deficits. Recent evidence demonstrates that glutamate receptors are dysregulated by amyloid beta peptide (Aβ) oligomers, resulting in disruption of glutamatergic synaptic transmission which parallels early cognitive deficits. Although it is well accepted that neuronal death in AD is related to disturbed intracellular Ca(2+) (Ca(2+)(i)) homeostasis, little is known about the contribution of NMDARs containing GluN2A or GluN2B subunits on Aβ-induced Ca(2+)(i) rise and neuronal dysfunction. Thus, the main goal of this work was to evaluate the role of NMDAR subunits in dysregulation of Ca(2+)(i) homeostasis induced by Aβ 1-42 preparation containing both oligomers (in higher percentage) and monomers in rat cerebral cortical neurons. The involvement of NMDARs was evaluated by pharmacological inhibition with MK-801 or the selective GluN2A and GLUN2B subunit antagonists NVP-AAM077 and ifenprodil, respectively. We show that Aβ, like NMDA, increase Ca(2+)(i) levels mainly through activation of NMDARs containing GluN2B subunits. Conversely, GluN2A-NMDARs antagonism potentiates Ca(2+)(i) rise induced by a high concentration of Aβ (1μM), suggesting that GluN2A and GluN2B subunits have opposite roles in regulating Ca(2+)(i) homeostasis. Moreover, Aβ modulate NMDA-induced responses and vice versa. Indeed, pre-exposure to Aβ (1μM) decrease NMDA-evoked Ca(2+)(I) rise and pre-exposure to NMDA decrease Aβ response. Interestingly, simultaneous addition of Aβ and NMDA potentiate Ca(2+)(I) levels, this effect being regulated by GluN2A and GluN2B subunits in opposite manners. This study contributes to the understanding of the molecular basis of early AD pathogenesis, by exploring the role of GluN2A and GluN2B subunits in the mechanism of Aβ toxicity in AD.     

Valosin-Containing Protein (VCP/p97) Is an Activator of Wild-Type Ataxin-3

Alterations in the ubiquitin-proteasome system (UPS) have been reported in several neurodegenerative disorders characterized by protein misfolding and aggregation, including the polylgutamine diseases. Machado-Joseph disease (MJD) or Spinocerebellar Ataxia type 3 is caused by a polyglutamine-encoding CAG expansion in the ATXN3 gene, which encodes a 42 kDa deubiquitinating enzyme (DUB), ataxin-3. We investigated ataxin-3 deubiquitinating activity and the functional relevance of ataxin-3 interactions with two proteins previously described to interact with ataxin-3, hHR23A and valosin-containing protein (VCP/p97). We confirmed ataxin-3 affinity for both hHR23A and VCP/p97. hHR23A and ataxin-3 were shown to co-localize in discrete nuclear foci, while VCP/p97 was primarily cytoplasmic. hHR23A and VCP/p97 recombinant proteins were added, separately or together, to normal and expanded ataxin-3 in in vitro deubiquitination assays to evaluate their influence on ataxin-3 activity. VCP/p97 was shown to be an activator specifically of wild-type ataxin-3, exhibiting no effect on expanded ataxin-3, In contrast, we observed no significant alterations in ataxin-3 enzyme kinetics or substrate preference in the presence of hHR23A alone or in combination with VCP. Based on our results we propose a model where ataxin-3 normally functions with its interactors to specify the cellular fate of ubiquitinated proteins.     

Compromised mitochondrial complex II in models of Machado-Joseph disease

Machado-Joseph disease (MJD), also known as Spinocerebellar Ataxia type 3, is an inherited dominant autosomal neurodegenerative disorder. An expansion of Cytosine-Adenine-Guanine (CAG) repeats in the ATXN3 gene is translated as an expanded polyglutamine domain in the disease protein, ataxin-3. Selective neurodegeneration in MJD is evident in several subcortical brain regions including the cerebellum. Mitochondrial dysfunction has been proposed as a mechanism of neurodegeneration in polyglutamine disorders. In this study, we used different cell models and transgenic mice to assess the importance of mitochondria on cytotoxicity observed in MJD. Transiently transfected HEK cell lines with expanded (Q84) ataxin-3 exhibited a higher susceptibility to 3-nitropropionic acid (3-NP), an irreversible inhibitor of mitochondrial complex II. Increased susceptibility to 3-NP was also detected in stably transfected PC6-3 cells that inducibly express expanded (Q108) ataxin-3 in a tetracycline-regulated manner. Moreover, cerebellar granule cells from MJD transgenic mice were more sensitive to 3-NP inhibition than wild-type cerebellar neurons. PC6-3 (Q108) cells differentiated into a neuronal-like phenotype with nerve growth factor (NGF) exhibited a significant decrease in mitochondrial complex II activity. Mitochondria from MJD transgenic mouse model and lymphoblast cell lines derived from MJD patients also showed a trend toward reduced complex II activity. Our results suggest that mitochondrial complex II activity is moderately compromised in MJD, which may designate a common feature in polyglutamine toxicity.