Investigation of thermogenic mechanisms in adipose tissue during recovery from sepsis

Background – Sepsis is defined as a dysregulated host response to infection resulting in organ dysfunction and, in some cases, death. Temperature impacts outcomes from sepsis – patients with fever are more likely to survive. However, in the recovery phase, thermogenesis may be detrimental. Survivors frequently develop cachexia and sepsis-induced myopathy which impairs recovery and increases long term mortality. In conditions akin to sepsis, including burn injury and cancer-associated cachexia, this has been attributed to catabolism driven by hypermetabolism due to a process called ‘browning’. Browning describes the switch of energy-storing white adipose tissue to a thermogenic energy-burning brown adipose tissue-like phenotype. Identification and prevention of browning in sepsis may improve outcomes. Hypothesis – In survivors of sepsis, browning of white adipose tissue occurs and drives cachexia and myopathy. Methods – Experimental sepsis was induced in rats using intraperitoneal zymosan. Body mass, muscle mass and myofibre cross sectional area were used to assess cachexia and myopathy. Expression of thermogenic browning mechanisms were studied in epididymal and retroperitoneal adipose tissue (eWAT and rpWAT, respectively) using thermal imaging, respirometry, RNA-sequencing and Western blot. Mitochondrial function and tissue morphology was interrogated by multiphoton imaging in live rpWAT explants. Results – Rats with zymosan peritonitis developed a sepsis-like illness with a 14-day mortality of 17%. In the recovery phase survivors developed hypermetabolism, cachexia and myopathy with reduced muscle mass and myofiber thickness. Oxygen consumption of eWAT and rpWAT per milligram of tissue was elevated at days 3, 7 4 and 14 of sepsis recovery. However, when controlled for protein content, lipid droplet size or mitochondrial or cell number, the increase was abolished. RNA sequencing of rpWAT demonstrated up-regulation of inflammatory genes and down-regulation of genes related to oxidative phosphorylation and thermogenesis during recovery. Notably, SERCA2 mRNA and SERCA2 protein were increased. Multiphoton microscopy showed neither increased mitochondrial density nor lipid multiloculation consistent with browning. The NAD(P)H pool was, however, more oxidised in tissue from animals recovering from sepsis, indicating altered metabolism. Conclusion – Hypermetabolism, cachexia and myopathy in the recovery phase of experimental sepsis are not caused by classical browning. Calcium cycling mechanisms in adipose tissue may be implicated and merit further investigation

Molecular evolutionary trends and feeding ecology diversification in the Hemiptera, anchored by the milkweed bug genome.

BACKGROUND: The Hemiptera (aphids, cicadas, and true bugs) are a key insect order, with high diversity for feeding ecology and excellent experimental tractability for molecular genetics. Building upon recent sequencing of hemipteran pests such as phloem-feeding aphids and blood-feeding bed bugs, we present the genome sequence and comparative analyses centered on the milkweed bug Oncopeltus fasciatus, a seed feeder of the family Lygaeidae. RESULTS: The 926-Mb Oncopeltus genome is well represented by the current assembly and official gene set. We use our genomic and RNA-seq data not only to characterize the protein-coding gene repertoire and perform isoform-specific RNAi, but also to elucidate patterns of molecular evolution and physiology. We find ongoing, lineage-specific expansion and diversification of repressive C2H2 zinc finger proteins. The discovery of intron gain and turnover specific to the Hemiptera also prompted the evaluation of lineage and genome size as predictors of gene structure evolution. Furthermore, we identify enzymatic gains and losses that correlate with feeding biology, particularly for reductions associated with derived, fluid nutrition feeding. CONCLUSIONS: With the milkweed bug, we now have a critical mass of sequenced species for a hemimetabolous insect order and close outgroup to the Holometabola, substantially improving the diversity of insect genomics. We thereby define commonalities among the Hemiptera and delve into how hemipteran genomes reflect distinct feeding ecologies. Given Oncopeltus's strength as an experimental model, these new sequence resources bolster the foundation for molecular research and highlight technical considerations for the analysis of medium-sized invertebrate genomes

The V471A polymorphism in autophagy-related gene ATG7 modifies age at onset specifically in Italian Huntington disease patients

The cause of Huntington disease (HD) is a polyglutamine repeat expansion of more than 36 units in the huntingtin protein, which is inversely correlated with the age at onset of the disease. However, additional genetic factors are believed to modify the course and the age at onset of HD. Recently, we identified the V471A polymorphism in the autophagy-related gene ATG7, a key component of the autophagy pathway that plays an important role in HD pathogenesis, to be associated with the age at onset in a large group of European Huntington disease patients. To confirm this association in a second independent patient cohort, we analysed the ATG7 V471A polymorphism in additional 1,464 European HD patients of the “REGISTRY” cohort from the European Huntington Disease Network (EHDN). In the entire REGISTRY cohort we could not confirm a modifying effect of the ATG7 V471A polymorphism. However, analysing a modifying effect of ATG7 in these REGISTRY patients and in patients of our previous HD cohort according to their ethnic origin, we identified a significant effect of the ATG7 V471A polymorphism on the HD age at onset only in the Italian population (327 patients). In these Italian patients, the polymorphism is associated with a 6-years earlier disease onset and thus seems to have an aggravating effect. We could specify the role of ATG7 as a genetic modifier for HD particularly in the Italian population. This result affirms the modifying influence of the autophagic pathway on the course of HD, but also suggests population-specific modifying mechanisms in HD pathogenesis

Identification of genetic variants associated with Huntington's disease progression: a genome-wide association study

Background Huntington's disease is caused by a CAG repeat expansion in the huntingtin gene, HTT. Age at onset has been used as a quantitative phenotype in genetic analysis looking for Huntington's disease modifiers, but is hard to define and not always available. Therefore, we aimed to generate a novel measure of disease progression and to identify genetic markers associated with this progression measure. Methods We generated a progression score on the basis of principal component analysis of prospectively acquired longitudinal changes in motor, cognitive, and imaging measures in the 218 indivduals in the TRACK-HD cohort of Huntington's disease gene mutation carriers (data collected 2008–11). We generated a parallel progression score using data from 1773 previously genotyped participants from the European Huntington's Disease Network REGISTRY study of Huntington's disease mutation carriers (data collected 2003–13). We did a genome-wide association analyses in terms of progression for 216 TRACK-HD participants and 1773 REGISTRY participants, then a meta-analysis of these results was undertaken. Findings Longitudinal motor, cognitive, and imaging scores were correlated with each other in TRACK-HD participants, justifying use of a single, cross-domain measure of disease progression in both studies. The TRACK-HD and REGISTRY progression measures were correlated with each other (r=0·674), and with age at onset (TRACK-HD, r=0·315; REGISTRY, r=0·234). The meta-analysis of progression in TRACK-HD and REGISTRY gave a genome-wide significant signal (p=1·12 × 10−10) on chromosome 5 spanning three genes: MSH3, DHFR, and MTRNR2L2. The genes in this locus were associated with progression in TRACK-HD (MSH3 p=2·94 × 10−8 DHFR p=8·37 × 10−7 MTRNR2L2 p=2·15 × 10−9) and to a lesser extent in REGISTRY (MSH3 p=9·36 × 10−4 DHFR p=8·45 × 10−4 MTRNR2L2 p=1·20 × 10−3). The lead single nucleotide polymorphism (SNP) in TRACK-HD (rs557874766) was genome-wide significant in the meta-analysis (p=1·58 × 10−8), and encodes an aminoacid change (Pro67Ala) in MSH3. In TRACK-HD, each copy of the minor allele at this SNP was associated with a 0·4 units per year (95% CI 0·16–0·66) reduction in the rate of change of the Unified Huntington's Disease Rating Scale (UHDRS) Total Motor Score, and a reduction of 0·12 units per year (95% CI 0·06–0·18) in the rate of change of UHDRS Total Functional Capacity score. These associations remained significant after adjusting for age of onset. Interpretation The multidomain progression measure in TRACK-HD was associated with a functional variant that was genome-wide significant in our meta-analysis. The association in only 216 participants implies that the progression measure is a sensitive reflection of disease burden, that the effect size at this locus is large, or both. Knockout of Msh3 reduces somatic expansion in Huntington's disease mouse models, suggesting this mechanism as an area for future therapeutic investigation

Character and temporal evolution of apoptosis in acetaminophen-induced acute liver failure

OBJECTIVE: To evaluate the role of hepatocellular and extrahepatic apoptosis during the evolution of Acetaminophen-induced acute liver failure (AALF). DESIGN & SETTING: A prospective observational study in two tertiary liver transplant units. PATIENTS: 88 patients with AALF were recruited. Control groups included patients with non-AALF (n=13), non-hepatic multi-organ failure (MOF, n=28), chronic liver disease (CLD, n=19) and healthy controls (HC, n=11). MEASUREMENTS: Total and caspase-cleaved cytokeratin 18 (M65 and M30 measured on admission and sequentially on day 3, 7 and 10 following admission. Levels were also determined from hepatic, portal vein and systemic arterial blood in seven patients undergoing transplantation. Protein arrays of liver homogenates from AALF patients were assessed for apoptosis-associated proteins and histological assessment of liver tissue was performed. MAIN RESULTS: Admission M30 levels were significantly elevated in AALF and NALF patients compared to MOF, CLD and healthy controls. Admission M30 levels correlated with outcome with AUROC of 0.755 (0.639-0.885, p<0.001). Peak levels in ALF patients were seen on admission then fell significantly but did not normalize over ten days. A negative gradient of M30 from the portal to hepatic vein was demonstrated in AALF patients (p=0.042) at the time of liver transplant. Analysis of protein array data demonstrated lower apoptosis-associated protein and higher catalase concentrations in AALF liver compared to controls (p<0.05). Explant histological analysis revealed evidence of cellular proliferation with an absence of histological evidence of apoptosis. CONCLUSIONS: Hepatocellular apoptosis occurs in the early phases of human AALF, peaking on day 1 of hospital admission and correlates strongly with poor outcome. Hepatic regenerative/tissue repair responses prevail during the later stages of ALF where elevated levels of M30 are likely to reflect epithelial cell death in extra-hepatic organs

From agonist to antagonist: structure and dynamics of innate immune glycoprotein md-2 upon recognition of variably acylated bacterial endotoxins

The human immune response to an infection by Gram-negative bacteria involves detection of lipopolysaccharides (LPS), also known as endotoxins, which comprise the bacterial outer cell wall. Distinct from mammalian glycolipid structures. LPS have a conserved chemical pattern that is recognized by the pattern recognition receptor complex formed by myeloid differentiation protein 2 (MD-2) and toll-like receptor 4 (TLR4). A remarkable immune-mediated structure-toxicity relationship has been defined that relates to the number of acyl chains in the endotoxin. While there is a clear correlation between endotoxin acylation and elicited agonist or antagonist responses, the 3D structural basis of this relationship remains unclear. In order to explore, at atomic-resolution, the effects of a range of chemically distinct endotoxins on the structure and dynamics of their MD-2.endotoxin complexes, we examined a series of variably acylated lipid A molecules from Escherichia coli and Neisseria meningitidis in complex with human MD-2. Through the application of molecular dynamics simulations, in concert with experimental data, we have identified specific structural and dynamic features of the MD-2-endotoxin complexes that may control dimerization of TLR4 molecules. As dimerization is central to the release of downstream chemical mediators, the results provide a structural foundation for the ability of endotoxins to act as either agonists or antagonists of the TLR4 pathway. Published by Elsevier Ltd