Cell surface sphingolipid glycohydrolases in neuronal differentiation and aging in culture

Qualitative and quantitative changes in glycosphingolipids, together with changes in the expression of the corresponding glycosyltransferases, have been reported along neuronal differentiation and aging. Plasma membrane (PM) glycosphingolipid pattern and content are the result of a complex network of metabolic pathways, including those potentially involving the activity of PM glycohydrolases. We analyzed the total cell activities of sialyltransferase I, II and IV, sialidase, β-galactosidase and β-glucosidase, and the PM-associated activities of sialidase Neu3, β-galactosidase, Conduritol B Epoxide-sensitive β-glucosidase and β-glucosidase GBA2 in rat cerebellar granule cells along differentiation and aging in culture. Sialyltransferase activities increased during cell differentiation, in agreement with the known increase of the total ganglioside content during neuronal maturation. The remodeling of ganglioside pattern could be because of the augmented activities of total sialidase and, within PM, to the action of the cell surface associated sialidase Neu3. Sialidase activities remained high during aging, in agreement with the known progressive ganglioside reduction in brain senescence. As PM β-galactosidase and β-glucosidase activities and parallely ceramide levels markedly increased along in vitro aging, PM ceramide production in neurons might be because of local catabolism of glycosphingolipids and not only to that of sphingomyelin, as already reported in human fibroblasts.     

Evidence for Sub-Haplogroup H5 of Mitochondrial DNA as a Risk Factor for Late Onset Alzheimer's Disease

Background

Alzheimer''s Disease (AD) is the most common neurodegenerative disease and the leading cause of dementia among senile subjects. It has been proposed that AD can be caused by defects in mitochondrial oxidative phosphorylation. Given the fundamental contribution of the mitochondrial genome (mtDNA) for the respiratory chain, there have been a number of studies investigating the association between mtDNA inherited variants and multifactorial diseases, however no general consensus has been reached yet on the correlation between mtDNA haplogroups and AD.

Methodology/Principal Findings

We applied for the first time a high resolution analysis (sequencing of displacement loop and restriction analysis of specific markers in the coding region of mtDNA) to investigate the possible association between mtDNA-inherited sequence variation and AD in 936 AD patients and 776 cognitively assessed normal controls from central and northern Italy. Among over 40 mtDNA sub-haplogroups analysed, we found that sub-haplogroup H5 is a risk factor for AD (OR = 1.85, 95% CI:1.04–3.23) in particular for females (OR = 2.19, 95% CI:1.06–4.51) and independently from the APOE genotype. Multivariate logistic regression revealed an interaction between H5 and age. When the whole sample is considered, the H5a subgroup of molecules, harboring the 4336 transition in the tRNA^Gln gene, already associated to AD in early studies, was about threefold more represented in AD patients than in controls (2.0% vs 0.8%; p = 0.031), and it might account for the increased frequency of H5 in AD patients (4.2% vs 2.3%). The complete re-sequencing of the 56 mtDNAs belonging to H5 revealed that AD patients showed a trend towards a higher number (p = 0.052) of sporadic mutations in tRNA and rRNA genes when compared with controls.

Conclusions

Our results indicate that high resolution analysis of inherited mtDNA sequence variation can help in identifying both ancient polymorphisms defining sub-haplogroups and the accumulation of sporadic mutations associated with complex traits such as AD.     

Nuclear Overhauser effect investigation on GM1 ganglioside containingN-glycolyl-neuraminic acid (II3Neu5GcGgOse4Cer)

The conformational properties of the oligosaccharide chain of GM1 ganglioside containingN-glycolyl-neuraminic acid, -Gal-(1-3)--GalNAc-(1-4)-[-Neu5Gc-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer, were studied through NMR nuclear Overhauser effect investigations on the monomeric ganglioside in dimethylsulfoxide, and on mixed micelles of ganglioside and dodecylphosphocholine in water. Several interresidual contacts for the trisaccharide core--GalNAc-(1-4)-[-Neu5Gc-(2-3)]--Gal-were found to fix the relative orientitation of the three saccharides, while the glycosidic linkage of the terminal -Gal-was found to be quite mobile as the -Gal-(1-3)--GalNAc-disaccharide exists in different conformations. These results are similar to those found for two GM1 gangliosides containingN-acetyl-neuraminic acid and neuraminic acid [1].Abbreviations Ganglioside nomenclature is in accordance with Svennerholm [23] and the IUPAC-IUB Recommendations [24] GM3(Neu5Ac) II³Neu5AcLacCer, -Neu5Ac-(2-3)--Gal-(1-4)--Glc-(1-1)-Cer - GM3(Neu5Gc) II³Neu5GcLacCer, -Neu5Gc-(2-3)--Gal-(1-4)--Glc-(1-1)-Cer - GM1(Neu5Ac) II³Neu5AcGgOse₄Cer, -Gal-(1-3)--GalNAc-(1-4)-[-Neu5Ac-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer - GM1(Neu5Gc) II³Neu5GcGgOse₄Cer, -Gal-(1-3)--GalNAc-(1-4)-[-Neu5Gc-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer - GM1(Neu) II³NeuGgOse₄Cer, -Gal-(1-3)--GalNAc-(1-4)-[-Neu-(2-3)]--Glc-(1-1)-Cer - GD1a IV³Neu5AcII³Neu5AcGgOse₄Cer, -Neu5Ac-(2-3)--Gal-(1-3)--GalNAc-(1-4)-[-Neu5Ac-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer - GalNAc-GD1a IV⁴GalNAcIV³Neu5AcII³Neu5AcGgOse₄Cer, -GalNAc-(1-4)-[-Neu5Ac-(2-3)]--Gal-(1-3)--GalNAc-(1-4)-[-Neu5Ac-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer - Neu neuraminic acid - Neu5Ac N-acetyl-neuraminic acid - Neu5Gc N-glycolyl-neuraminic acid - Cer ceramide     

Place of Residence Moderates the Risk of Infant Death in Kenya: Evidence from the Most Recent Census 2009

Background

Substantial progress has been made in reducing childhood mortality worldwide from 1990–2015 (Millennium Development Goal, target 4). Achieving target goals on this however remains a challenge in Sub-Saharan Africa. Kenya’s infant mortality rates are higher than the global average and are more pronounced in urban areas as compared to rural areas. Only limited knowledge exists about the differences in individual level risk factors for infant death among rural, non-slum urban, and slum areas in Kenya. Therefore, this paper aims at 1) assess individual and socio-ecological risk factors for infant death in Kenya, and at 2) identify whether living in rural, non-slum urban, or slum areas moderated individual or socio-ecological risk factors for infant death in Kenya.

Methodology

We used a cross-sectional study design based on the most recent Kenya Population and Housing Census of 2009 and extracted the records of all females who had their last child born in 12 months preceding the survey (N = 1,120,960). Multivariable regression analyses were used to identify risk factors that accounted for the risk of dying before the age of one at the individual level in Kenya. Place of residence (rural, non-slum urban, slum) was used as an interaction term to account for moderating effects in individual and socio-ecological risk factors.

Results

Individual characteristics of mothers and children (older age, less previously born children that died, better education, girl infants) and household contexts (better structural quality of housing, improved water and sanitation, married household head) were associated with lower risk for infant death in Kenya. Living in non-slum urban areas was associated with significantly lower infant death as compared to living in rural or slum areas, when all predictors were held at their reference levels. Moreover, place of residence was significantly moderating individual level predictors: As compared to rural areas, living in urban areas was a protective factor for mothers who had previous born children who died, and who were better educated. However, living in urban areas also reduced the health promoting effects of better structural quality of housing (i.e. poor or good versus non-durable). Furthermore, durable housing quality in urban areas turned out to be a risk factor for infant death as compared to rural areas. Living in slum areas was also a protective factor for mothers with previous child death, however it also reduced the promoting effects of older ages in mothers.

Conclusions

While urbanization and slum development continues in Kenya, public health interventions should invest in healthy environments that ideally would include improvements to access to safe water and sanitation, better structural quality of housing, and to access to education, health care, and family planning services, especially in urban slums and rural areas. In non-slum urban areas however, health education programs that target healthy diets and promote physical exercise may be an important adjunct to these structural interventions.     

Identification and characterization of a novel non-structural protein of bluetongue virus

Bluetongue virus (BTV) is the causative agent of a major disease of livestock (bluetongue). For over two decades, it has been widely accepted that the 10 segments of the dsRNA genome of BTV encode for 7 structural and 3 non-structural proteins. The non-structural proteins (NS1, NS2, NS3/NS3a) play different key roles during the viral replication cycle. In this study we show that BTV expresses a fourth non-structural protein (that we designated NS4) encoded by an open reading frame in segment 9 overlapping the open reading frame encoding VP6. NS4 is 77-79 amino acid residues in length and highly conserved among several BTV serotypes/strains. NS4 was expressed early post-infection and localized in the nucleoli of BTV infected cells. By reverse genetics, we showed that NS4 is dispensable for BTV replication in vitro, both in mammalian and insect cells, and does not affect viral virulence in murine models of bluetongue infection. Interestingly, NS4 conferred a replication advantage to BTV-8, but not to BTV-1, in cells in an interferon (IFN)-induced antiviral state. However, the BTV-1 NS4 conferred a replication advantage both to a BTV-8 reassortant containing the entire segment 9 of BTV-1 and to a BTV-8 mutant with the NS4 identical to the homologous BTV-1 protein. Collectively, this study suggests that NS4 plays an important role in virus-host interaction and is one of the mechanisms played, at least by BTV-8, to counteract the antiviral response of the host. In addition, the distinct nucleolar localization of NS4, being expressed by a virus that replicates exclusively in the cytoplasm, offers new avenues to investigate the multiple roles played by the nucleolus in the biology of the cell.     

BRAF V600E mutations are common in pleomorphic xanthoastrocytoma: diagnostic and therapeutic implications

Pleomorphic xanthoastrocytoma (PXA) is low-grade glial neoplasm principally affecting children and young adults. Approximately 40% of PXA are reported to recur within 10 years of primary resection. Upon recurrence, patients receive radiation therapy and conventional chemotherapeutics designed for high-grade gliomas. Genetic changes that can be targeted by selective therapeutics have not been extensively evaluated in PXA and ancillary diagnostic tests to help discriminate PXA from other pleomorphic and often more aggressive astrocytic malignancies are limited. In this study, we apply the SNaPshot multiplexed targeted sequencing platform in the analysis of brain tumors to interrogate 60 genetic loci that are frequently mutated in 15 cancer genes. In our analysis we detect BRAF V600E mutations in 12 of 20 (60%) WHO grade II PXA, in 1 of 6 (17%) PXA with anaplasia and in 1 glioblastoma arising in a PXA. Phospho-ERK was detected in all tumors independent of the BRAF mutation status. BRAF duplication was not detected in any of the PXA cases. BRAF V600E mutations were identified in only 2 of 71 (2.8%) glioblastoma (GBM) analyzed, including 1 of 9 (11.1%) giant cell GBM (gcGBM). The finding that BRAF V600E mutations are common in the majority of PXA has important therapeutic implications and may help in differentiating less aggressive PXAs from lethal gcGBMs and GBMs.