More than 50 genomic regions have now been shown to influence the risk of multiple sclerosis (MS). However, the mechanisms of action, and the cell types in which these associated variants act at the ...molecular level remain largely unknown. This is especially true for associated regions containing no known genes. Given the evidence for a role for B cells in MS, we hypothesized that MS associated genomic regions co-localized with regions which are functionally active in B cells. We used publicly available data on 1) MS associated regions and single nucleotide polymorphisms (SNPs) and 2) chromatin profiling in B cells as well as three additional cell types thought to be unrelated to MS (hepatocytes, fibroblasts and keratinocytes). Genomic intervals and SNPs were tested for overlap using the Genomic Hyperbrowser. We found that MS associated regions are significantly enriched in strong enhancer, active promoter and strong transcribed regions (p = 0.00005) and that this overlap is significantly higher in B cells than control cells. In addition, MS associated SNPs also land in active promoter (p = 0.00005) and enhancer regions more than expected by chance (strong enhancer p = 0.0006; weak enhancer p = 0.00005). These results confirm the important role of the immune system and specifically B cells in MS and suggest that MS risk variants exert a gene regulatory role. Previous studies assessing MS risk variants in T cells may be missing important effects in B cells. Similar analyses in other immunological cell types relevant to MS and functional studies are necessary to fully elucidate how genes contribute to MS pathogenesis.
Attenuated Total Reflection Fourier Transform Infrared (ATR-FTIR) and Raman spectroscopy were used to compare chloroquine (CQ)-treated and untreated cultured Plasmodium falciparum-infected human red ...blood cells (iRBCs). The studies were carried out in parallel from the same starting cultures using both spectroscopic techniques, in duplicate. ATR FTIR spectra showed modifications in the heme vibrational bands as well as increases in the CH2/CH3 stretching bands in the 3100-2800 cm(-1) region of CQ-treated iRBCs consistent with an increase in lipid content. Other changes consisted of secondary structural variations including shifts in the amide I and II modes, along with changes in RNA and carbohydrate bands. Raman microspectroscopy of single red blood cells using 532 nm revealed subtle changes in the positions and intensity of ν37 of the core size region marker band and ν4 in the pyrrole ring-stretching region between untreated and CQ-treated iRBCs. Similar patterns in the corresponding relations were also observed in the non-fundamental (overtone region) between the control and treated cells. These differences were consistent with higher levels of oxygenated hemoglobin (oxyHb) in the treated cells as shown in a Principle Component Analysis (PCA) loadings plot. The results obtained demonstrate that vibrational spectroscopic techniques can provide insight into the effect of quinolines on iRBCs and thus may assist understanding the sensitivity and resistance of new and existing anti-malarial drugs.
Resident peritoneal macrophages (M phi) were labeled in situ by intraperitoneal (i.p.) injection of the green fluorescent cell tracking dye PKH-1. After immunofluorescence staining with M phi ...specific monoclonal antibodies (Mabs) and phycoerythrin (PE) second antibody, the resident M phi were labeled with both the green dye and red Mab label, while recruited M phi were labeled only with the red Mab tag. These populations were distinguished by two-color flow cytometry. PKH-1 labeled resident peritoneal M phi were followed for 1-49 days in mice that received no further treatment (steady state). Dye labeled M phi were still detectable after 49 days in vivo, although their green fluorescence intensity had decreased steadily over time. The decrease in dye intensity was limited to M phi, as the fluorescence intensity of PKH-1 labeled peritoneal lymphocytes did not change. Resident M phi populations were clearly separated from recruited M phi by the intensity of their staining with PKH-1 for up to 28 days. No decrease in the number of resident (dye labeled) peritoneal M phi was observed over 1-28 days. These data indicate that resident peritoneal M phi were not replaced by recruited blood monocytes in the steady state.
The initial characterization of the first single-photon emission computed tomography (SPECT) insert for simultaneous human brain imaging inside magnetic resonance imaging (MRI) is presented. The ...instrument features a novel multimini-slit-slat collimator and a static ring of 20 detectors. The scanner architecture is modular and represents the scale-up of a preclinical version whose MRI-compatibility has been already successfully demonstrated. The detectors are based on CsI(Tl) crystals (50 mm <inline-formula> <tex-math notation="LaTeX">\times </tex-math></inline-formula> 100 mm <inline-formula> <tex-math notation="LaTeX">\times </tex-math></inline-formula> 8 mm) coupled to arrays of silicon photomultipliers and ASIC readout for a total of 1440 channels. The transaxial field of view (FoV) is 20 cm by 9 cm in the axial direction. The experimental results show a sensitivity of 362 cps/MBq, an energy resolution of ~17% (at 140 keV), and a tomographic extrinsic resolution in the transaxial plane of ~8 mm across the whole FoV.
A loss of function mutation in the TRESK K2P potassium channel (KCNK18), has recently been linked with typical familial migraine with aura. We now report the functional characterisation of additional ...TRESK channel missense variants identified in unrelated patients. Several variants either had no apparent functional effect, or they caused a reduction in channel activity. However, the C110R variant was found to cause a complete loss of TRESK function, yet is present in both sporadic migraine and control cohorts, and no variation in KCNK18 copy number was found. Thus despite the previously identified association between loss of TRESK channel activity and migraine in a large multigenerational pedigree, this finding indicates that a single non-functional TRESK variant is not alone sufficient to cause typical migraine and highlights the genetic complexity of this disorder.
The gut microbiome changes in response to a range of environmental conditions, life events and disease states. Pregnancy is a natural life event that involves major physiological adaptation yet ...studies of the microbiome in pregnancy are limited and their findings inconsistent. Pregnancy with type 1 diabetes (T1D) is associated with increased maternal and fetal risks but the gut microbiome in this context has not been characterized. By whole metagenome sequencing (WMS), we defined the taxonomic composition and function of the gut bacterial microbiome across 70 pregnancies, 36 in women with T1D.
Women with and without T1D exhibited compositional and functional changes in the gut microbiome across pregnancy. Profiles in women with T1D were distinct, with an increase in bacteria that produce lipopolysaccharides and a decrease in those that produce short-chain fatty acids, especially in the third trimester. In addition, women with T1D had elevated concentrations of fecal calprotectin, a marker of intestinal inflammation, and serum intestinal fatty acid-binding protein (I-FABP), a marker of intestinal epithelial damage.
Women with T1D exhibit a shift towards a more pro-inflammatory gut microbiome during pregnancy, associated with evidence of intestinal inflammation. These changes could contribute to the increased risk of pregnancy complications in women with T1D and are potentially modifiable by dietary means. Video abstract.
The purpose of the present study was to determine whether exposure to stress or elevated corticosterone concentrations in the days preceding cerebral ischemia exacerbates ischemic injury as assessed ...by histological and behavioral outcomes.
For 7 consecutive days, male C57/BL6 mice were exposed to social stress for 45 minutes or injected with 1 mg/kg corticosterone or vehicle. The animals exposed to social stress were injected with either 1 mg/kg mifepristone, a glucocorticoid receptor antagonist, or the vehicle 30 minutes before stress. On the seventh day, all animals were trained in a passive avoidance task. Twenty-four hours after training, the animals were subjected to 60 minutes of intraluminal middle cerebral artery occlusion (MCAO) or sham surgery. At 72 hours of reperfusion, the animals were tested for retention of the passive avoidance task, and infarction size was determined.
Animals subjected to chronic social stress or treated with exogenous corticosterone before MCAO exhibited larger infarcts and reduced retention of passive avoidance compared with the nonstressed MCAO control. The effects of social stress on infarct volume and passive avoidance were reversed by pretreatment with mifepristone. There was no difference between stressed and control groups in physiological parameters or reduction of laser-Doppler flow signal during MCAO or reperfusion.
Prior exposure to social stress increases infarction volume and exacerbates cognitive deficits associated with transient cerebral ischemia. The mechanism underlying the effects of stress on stroke outcome likely involves corticosterone acting through glucocorticoid receptors to increase subsequent ischemia-induced neuronal death.
ABSTRACT
The genomes of malaria parasites (Plasmodium spp.) contain a family of genes encoding proteins with a Plasmodium helical interspersed subtelomeric (PHIST) domain, most of which are predicted ...to be exported into the parasite‐infected human red blood cell (iRBC). Here, using transgenic parasites and a combination of cellular, biochemical, and biophysical assays, we have characterized and determined the function of a novel member of the PHIST protein family in Plasmodium falciparum, termed lysine‐rich membrane‐associated PHISTb (LyMP). LyMP was shown to associate directly with the cytoskeleton of iRBCs where it plays a role in their abnormal ability to adhere to a protein expressed on vascular endothelial cells, resulting in sequestration. Deletion of LyMP dramatically reduced adhesion of iRBCs to CD36 by 55%, which was completely restored to wild‐type levels on complementation. Intriguingly, in the absence of LyMP, formation of RBC membrane knobs and the level of surface exposure of the parasites' major cytoadhesive ligand, PfEMP1, were identical to those for the parental parasite line, demonstrating for the first time an additional mechanism that enhances cytoadherence of iRBCs beyond those already recognized. Our findings identify LyMP as a previously unknown RBC cytoskeletal‐binding protein that is likely to be of major significance in the complex pathophysiology of falciparum malaria.—Proellocks, N. I., Herrmann, S., Buckingham, D. W., Hanssen, E., Hodges, E. K., Elsworth, B., Morahan, B. J., Coppel, R. L., Cooke, B. M. A lysine‐rich membrane‐associated PHISTb protein involved in alteration of the cytoadhesive properties of Plasmodium falciparum infected red blood cells. FASEB J. 28, 3103–3113 (2014). www.fasebj.org
The kinetics of macrophage (M phi) recruitment to the peritoneum following the induction of acute inflammation by thioglycollate broth (TG) was evaluated after prelabeling resident M phi with the ...fluorescent cell tracking dye, PKH-1. Most of the PKH-1-labeled resident M phi disappeared from the recoverable peritoneal cell population within the first hour after injection of TG. This disappearance coincided with the inflammatory influx of neutrophils (PMNs) and was sustained for at least 5 days after administration of TG, although the PMN number had returned to resident levels by this time. PKH-1-labeled peritoneal M phi were observed again in most animals at 7 days after injection of TG. The number of labeled M phi recovered at 7 days was approximately twice the number of resident peritoneal M phi in control animals which did not receive the TG broth. These additional M phi may include progeny of either the resident M phi or other local M phi precursors, such as omental M phi, which were labeled by the PKH-1 injection.
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