Circadian Rhythms in 6-Sulphatoxymelatonin and Nocturnal Sleep in Blind Children

This article describes the relationship between melatonin secretion and sleep quality and subjective complaints about sleep in totally blind children. Eleven boarding-school children (mean age 15.2 years) participated. The major urinary melatonin metabolite 6-sulphatoxymelatonin (aMT6s) was measured five times a day for 48 h. Sleep-wake cycles were recorded by continuous actigraphic recordings during the same time period. Results showed that delayed secretory peaks in aMT6s were significantly associated with disturbed nocturnal sleep and with complaints about morning fatigue.     

CaV1.2 I-II linker structure and Timothy syndrome

Ca_V channels are multi-subunit protein complexes that enable inward cellular Ca²⁺ currents in response to membrane depolarization. We recently described structure-function studies of the intracellular α1 subunit domain I-II linker, directly downstream of domain IS6. The results show the extent of the linker’s helical structure to be subfamily dependent, as dictated by highly conserved primary sequence differences. Moreover, the difference in structure confers different biophysical properties, particularly the extent and kinetics of voltage and calcium-dependent inactivation. Timothy syndrome is a human genetic disorder due to mutations in the Ca_V1.2 gene. Here, we explored whether perturbation of the I-II linker helical structure might provide a mechanistic explanation for a Timothy syndrome mutant’s (human Ca_V1.2 G406R equivalent) biophysical effects on inactivation and activation. The results are equivocal, suggesting that a full mechanistic explanation for this Timothy syndrome mutation requires further investigation.     

Taxonomic Classification of Bacterial 16S rRNA Genes Using Short Sequencing Reads: Evaluation of Effective Study Designs

Massively parallel high throughput sequencing technologies allow us to interrogate the microbial composition of biological samples at unprecedented resolution. The typical approach is to perform high-throughout sequencing of 16S rRNA genes, which are then taxonomically classified based on similarity to known sequences in existing databases. Current technologies cause a predicament though, because although they enable deep coverage of samples, they are limited in the length of sequence they can produce. As a result, high-throughout studies of microbial communities often do not sequence the entire 16S rRNA gene. The challenge is to obtain reliable representation of bacterial communities through taxonomic classification of short 16S rRNA gene sequences. In this study we explored properties of different study designs and developed specific recommendations for effective use of short-read sequencing technologies for the purpose of interrogating bacterial communities, with a focus on classification using naïve Bayesian classifiers. To assess precision and coverage of each design, we used a collection of ∼8,500 manually curated 16S rRNA gene sequences from cultured bacteria and a set of over one million bacterial 16S rRNA gene sequences retrieved from environmental samples, respectively. We also tested different configurations of taxonomic classification approaches using short read sequencing data, and provide recommendations for optimal choice of the relevant parameters. We conclude that with a judicious selection of the sequenced region and the corresponding choice of a suitable training set for taxonomic classification, it is possible to explore bacterial communities at great depth using current technologies, with only a minimal loss of taxonomic resolution.     

Removal of adult cyathostomins alters faecal microbiota and promotes an inflammatory phenotype in horses

The interactions between parasitic helminths and gut microbiota are considered to be an important, although as yet incompletely understood, factor in the regulation of immunity, inflammation and a range of diseases. Infection with intestinal helminths is ubiquitous in grazing horses, with cyathostomins (about 50 species of which are recorded) predominating. Consequences of infection include both chronic effects, and an acute inflammatory syndrome, acute larval cyathostominosis, which sometimes follows removal of adult helminths by administration of anthelmintic drugs. The presence of cyathostomins as a resident helminth population of the equine gut (the “helminthome”) provides an opportunity to investigate the effect helminth infection, and its perturbation, has on both the immune system and bacterial microbiome of the gut, as well as to determine the specific mechanisms of pathophysiology involved in equine acute larval cyathostominosis. We studied changes in the faecal microbiota of two groups of horses following treatment with anthelmintics (fenbendazole or moxidectin). We found decreases in both alpha diversity and beta diversity of the faecal microbiota at Day 7 post-treatment, which were reversed by Day 14. These changes were accompanied by increases in inflammatory biomarkers. The general pattern of faecal microbiota detected was similar to that seen in the relatively few equine gut microbiome studies reported to date. We conclude that interplay between resident cyathostomin populations and the bacterial microbiota of the equine large intestine is important in maintaining homeostasis and that disturbance of this ecology can lead to gut dysbiosis and play a role in the aetiology of inflammatory conditions in the horse, including acute larval cyathostominosis.     

Cognitive State following Stroke: The Predominant Role of Preexisting White Matter Lesions

Background and purpose

Stroke is a major cause of cognitive impairment and dementia in adults, however the role of the ischemic lesions themselves, on top of other risk factors known in the elderly, remains controversial. This study used structural equation modeling to determine the respective impact of the new ischemic lesions'' volume, preexisting white matter lesions and white matter integrity on post stroke cognitive state.

Methods

Consecutive first ever mild to moderate stroke or transient ischemic attack patients recruited into the ongoing prospective TABASCO study underwent magnetic resonance imaging scans within seven days of stroke onset and were cognitively assessed one year after the event using a computerized neuropsychological battery. The volumes of both ischemic lesions and preexisting white matter lesions and the integrity of the normal appearing white matter tissue were measured and their contribution to cognitive state was assessed using structural equation modeling path analysis taking into account demographic parameters. Two models were hypothesized, differing by the role of ischemic lesions'' volume.

Results

Structural equation modeling analysis of 142 patients confirmed the predominant role of white matter lesion volume (standardized path coefficient β = −0.231) and normal appearing white matter integrity (β = −0.176) on the global cognitive score, while ischemic lesions'' volume showed no such effect (β = 0.038). The model excluding the ischemic lesion presented better fit to the data (comparative fit index 0.9 versus 0.092).

Conclusions

Mild to moderate stroke patients with preexisting white matter lesions are more vulnerable to cognitive impairment regardless of their new ischemic lesions. Thus, these patients can serve as a target group for studies on cognitive rehabilitation and neuro-protective therapies which may, in turn, slow their cognitive deterioration.     

Mass Spectrometry-based Structural Analysis and Systems Immunoproteomics Strategies for Deciphering the Host Response to Endotoxin

One cause of sepsis is systemic maladaptive immune response of the host to bacteria and specifically, to Gram-negative bacterial outer-membrane glycolipid lipopolysaccharide (LPS). On the host myeloid cell surface, proinflammatory LPS activates the innate immune system via Toll-like receptor-4/myeloid differentiation factor-2 complex. Intracellularly, LPS is also sensed by the noncanonical inflammasome through caspase-11 in mice and 4/5 in humans. The minimal functional determinant for innate immune activation is the membrane anchor of LPS called lipid A. Even subtle modifications to the lipid A scaffold can enable, diminish, or abolish immune activation. Bacteria are known to modify their LPS structure during environmental stress and infection of hosts to alter cellular immune phenotypes. In this review, we describe how mass spectrometry-based structural analysis of endotoxin helped uncover major determinations of molecular pathogenesis. Through characterization of LPS modifications, we now better understand resistance to antibiotics and cationic antimicrobial peptides, as well as how the environment impacts overall endotoxin structure. In addition, mass spectrometry-based systems immunoproteomics approaches can assist in elucidating the immune response against LPS. Many regulatory proteins have been characterized through proteomics and global/targeted analysis of protein modifications, enabling the discovery and characterization of novel endotoxin-mediated protein translational modifications.