Biology subjects

Kelly, M.

Publications and source records attributed to Kelly, M..

At least 55 records · Page 3Linked to original sources

bioRxiv2023

Shigella O-specific polysaccharide functional IgA responses mediate protection against shigella infection in an endemic high-burden setting

Shigella is the second leading cause of diarrheal disease-related death in young children in low and middle income countries. The mechanism of protection against shigella infection and disease in endemic areas is uncertain. While historically LPS-specific IgG titers have been associated with protection in endemic settings, emerging deeper immune approaches have recently elucidated a protective role for IpaB-specific antibody responses in a controlled human challenge model in North American volunteers. To deeply interrogate potential correlates of immunity in areas endemic for shigellosis, here we applied a systems approach to analyze the serological response to shigella across endemic and non-endemic populations. Additionally, we analyzed shigella-specific antibody responses over time in the context of endemic resistance or breakthrough infections in a high shigella burden location. Individuals with endemic exposure to shigella possessed broad and functional antibody responses across both glycolipid and protein antigens compared to individuals from non-endemic regions. In high shigella burden settings, elevated levels of OSP-specific FcR binding antibodies were associated with resistance to shigellosis. OSP-specific FcR binding IgA found in resistant individuals activated bactericidal neutrophil functions including phagocytosis, degranulation and reactive oxygen species production. Moreover, IgA depletion from resistant serum significantly reduced binding of OSP-specific antibodies to FcR and antibody mediated activation of neutrophils and monocytes. Overall, our findings suggest that OSP-specific functional IgA responses contribute to protective immunity against shigella infection in high-burden settings. These findings will assist in the development and evaluation of shigella vaccines.

immunology↗

bioRxiv2023

Lactate transporter MCT1 in hepatic stellate cells promotes fibrotic collagen expression in nonalcoholic steatohepatitis

Circulating lactate is a fuel source for liver metabolism but may exacerbate metabolic diseases such as nonalcoholic steatohepatitis (NASH). Indeed, haploinsufficiency of lactate transporter monocarboxylate transporter 1 (MCT1) in mice reportedly promotes resistance to hepatic steatosis and inflammation. Here, we used adeno-associated virus (AAV) vectors to deliver thyroxin binding globulin (TBG)-Cre or lecithin-retinol acyltransferase (Lrat)-Cre to MCT1fl/fl mice on a choline deficient, high fat NASH diet to deplete hepatocyte or stellate cell MCT1, respectively. Stellate cell MCT1KO (AAV-Lrat-Cre) attenuated liver type 1 collagen protein expression and caused a downward trend in trichrome staining. MCT1 depletion in cultured human LX2 stellate cells also diminished collagen 1 protein expression. Tetra-ethylenglycol-cholesterol (Chol)-conjugated siRNAs, which enter all hepatic cell types, and hepatocyte-selective tri-N-acetyl galactosamine (GN)-conjugated siRNAs were then used to evaluate MCT1 function in a genetically obese NASH mouse model. MCT1 silencing by Chol-siRNA decreased liver collagen 1 levels, while hepatocyte-selective MCT1 depletion by AAV-TBG-Cre or by GN-siRNA unexpectedly increased collagen 1 and total fibrosis without effect on triglyceride accumulation. These findings demonstrate that stellate cell lactate transporter MCT1 significantly contributes to liver fibrosis through increased collagen 1 protein expression in vitro and in vivo, while hepatocyte MCT1 appears not to be an attractive therapeutic target for NASH.

molecular biology↗

bioRxiv2023

mTOR Inhibition Suppresses Salinomycin-Induced Ferroptosis in Breast Cancer Stem Cells by Ironing Out Mitochondrial Dysfunctions

Ferroptosis constitutes a promising therapeutic strategy against cancer by efficiently targeting the highly tumorigenic and treatment-resistant cancer stem cells (CSCs). We previously showed that the lysosomal iron-targeting drug Salinomycin (Sal) was able to eliminate CSCs by triggering ferroptosis. Here, in a well-established breast CSCs model (human mammary epithelial HMLER CD24low/CD44high), we identified that pharmacological inhibition of mechanistic target of rapamycin (mTOR), suppresses Sal-induced ferroptosis. Mechanistically, mTOR inhibition modulates iron cellular flux and prevents the iron and ROS bursts induced by Sal. Besides, integration of multi-omics data identified mitochondria as a key target of Sal action. We demonstrated that mTOR inhibition prevents Sal-induced mitochondrial functional and structural alteration, and that Sal-induced metabolic plasticity is mainly dependent on the mTOR pathway. Overall, our findings provide experimental evidences on the detailed mechanisms of mTOR as a crucial effector of Sal-induced ferroptosis, and gives proof-of-concept that careful evaluation of such combination therapy (here mTOR and ferroptosis co-targeting) is required for effective treatment. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=120 SRC="FIGDIR/small/539040v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@8b8772org.highwire.dtl.DTLVardef@695e0forg.highwire.dtl.DTLVardef@19e3f83org.highwire.dtl.DTLVardef@5bc0a6_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2022

Biologic that disrupts PDE11A4 homodimerization in hippocampus CA1 reverses age-related proteinopathies in PDE11A4 and cognitive decline of social memories

Age-related proteinopathies in phosphodiesterase 11A (PDE11A), an enzyme that degrades 3,5-cAMP/cGMP and is enriched in the ventral hippocampal formation (VHIPP), drive age-related cognitive decline (ARCD) of social memories. In the VHIPP, age-related increases in PDE11A4 occur specifically within the membrane compartment and ectopically accumulate in filamentous structures termed ghost axons. Previous in vitro studies show that disrupting PDE11 homodimerization by expressing an isolated PDE11A-GAFB domain, which acts as a "negative sink" for monomers, selectively degrades membrane-associated PDE11A4 and prevents the punctate accumulation of PDE11A4. Therefore, we determined if disrupting PDE11A4 homodimerization in vivo via the expression of an isolated PDE11A4-GAFB domain would be sufficient to reverse 1) age-related accumulations of PDE11A4 in VHIPP ghost axons and 2) ARCD of social memories. Indeed, in vivo lentiviral expression of the isolated PDE11A4-GAFB domain in hippocampal CA1 reversed the age-related accumulation of PDE11A4 in ghost axons, reversed ACRD of social transmission of food preference memory (STFP), and improved remote long-term memory for social odor recognition (SOR) without affecting memory for non-social odor recognition. In vitro studies suggest that disrupting homodimerization of PDE11A4 does not directly alter the catalytic activity of the enzyme but may reverse age-related decreases in cGMP by dispersing the accumulation of the enzyme independently of other intramolecular mechanisms previously established to disperse PDE11A4 (e.g., phosphorylation of PDE11A4 at serine 162). Altogether, these data suggest that a biologic designed to disrupt PDE11A4 homodimerization may serve to ameliorate age-related deficits in hippocampal cyclic nucleotide signaling and subsequent ARCD of remote social memory.

neuroscience↗

bioRxiv2022

Genome-wide determination of barriers to horizontal gene transfer

Horizontal gene transfer (HGT) is a major contributor to bacterial genome evolution, generating phenotypic diversity, driving the expansion of protein families, and facilitating the evolution of new phenotypes, new metabolic pathways, and new species. Comparative studies of gene gain in bacteria suggest that the frequency with which individual genes successfully undergo HGT varies considerably and may be associated with the number of protein-protein interactions in which the gene participates--its connectivity. Two non-exclusive hypotheses have emerged to explain why transferability should decrease with connectivity: the Complexity Hypothesis (Jain, Rivera, & Lake, 1999) and the Balance Hypothesis (Papp, Pal, & Hurst, 2003). These hypotheses predict that the functional costs of HGT arise from a failure of divergent homologues to make normal protein-protein interactions or from gene mis-expression, respectively. Here we describe genome-wide assessments of these hypotheses in which we used 74 existing prokaryotic whole genome shotgun libraries to estimate rates of horizontal transfer of genes from taxonomically diverse prokaryotic donors into E. coli. We show that transferability declines as connectivity increases, but that this effect is driven primarily by the ribosomal genes. We also show that transferability declines as the divergence (% amino acid difference) between donor and recipient orthologs increases and that this effect of divergence increases with connectivity. We explain how these results, even the stronger effect of connectivity on the transferability of ribosomal compared to non-ribosomal genes, provide strong support for both the Balance and Complexity Hypotheses. Significance StatementComparisons between prokaryotic genomes consistently show that genes with informational functions, e.g. in genome replication, transcription, and translation, have been subject to horizontal gene transfer between species more often than genes with operational functions, e.g. in metabolism and environmental sensing. In this study, we use a genome wide analysis of transferability data obtained from 74 genomes to provide the first experimental evidence that this pattern results from differences between informational and operational genes in the number of other proteins with which they interact, i.e., their connectivity, rather than from their functional differences. The importance of our exceptionally large dataset to the detection of connectivity effects on transferability explains why past experimental studies failed to replicate the consistent finding from comparative genomic studies.

genomics↗

bioRxiv2022

De novo lipogenesis fuels adipocyte autophagosome membrane dynamics

Autophagy is a homeostatic degradative process for cell components that enables stress resilience and can determine cellular fate and function. However, lipid sources for the rapid membrane expansions of autophagosomes, the workhorses of autophagy, are poorly understood. Here, we identify de novo lipogenesis (DNL) as a critical source of fatty acids (FA) to fuel autophagosome dynamics in adipocytes. Adipocyte fatty acid synthase (Fasn) deficiency markedly impairs autophagy, evident by autophagosome accumulation, and severely compromises degradation of the autophagic substrate p62. Autophagy dependence on FA produced by Fasn is not fully alleviated by exogenous FA in cultured adipocytes even though lipid droplet size is restored. Imaging studies reveal that Fasn colocalizes with nascent autophagosomes, while loss of Fasn decreases certain membrane phosphoinositides known to be required for autophagosome assembly. Together, our studies highlight a newly appreciated function for adipocyte DNL in autophagosome membrane formation and provide evidence that localized FA synthesis contributes to autophagosome dynamics.

molecular biology↗

bioRxiv2022

Paradoxical activation of SREBP1c and de novo lipogenesis by hepatocyte-selective ACLY depletion in obese mice.

Hepatic steatosis associated with high fat diets, obesity and type 2 diabetes is thought to be the major driver of severe liver inflammation, fibrosis, and cirrhosis. Cytosolic acetyl-coenzyme A (AcCoA), a central metabolite and substrate for de novo lipogenesis (DNL), is produced from citrate by ATP-citrate lyase (ACLY) and from acetate through AcCoA synthase short chain family member 2 (ACSS2). However, the relative contributions of these two enzymes to hepatic AcCoA pools and DNL rates in response to high fat feeding is unknown. We report here that hepatocyte-selective depletion of either ACSS2 or ACLY caused similar 50% decreases in liver AcCoA levels in obese mice, showing that both pathways contribute to generation of this DNL substrate. Unexpectedly however, the hepatocyte ACLY depletion in obese mice paradoxically increased total DNL flux measured by D2O incorporation into palmitate, while in contrast ACSS2 depletion had no effect. The increase in liver DNL upon ACLY depletion was associated with increased expression of nuclear sterol regulatory element-binding protein 1c (SREBP1c) and of its target DNL enzymes. This upregulated DNL enzyme expression explains the increased rate of palmitate synthesis in ACLY depleted livers. Furthermore, this increased flux through DNL may also contribute to the observed depletion of AcCoA levels due to its increased conversion to Malonyl CoA (MalCoA) and palmitate. Together, these data indicate that in HFD fed obese mice, hepatic DNL is not limited by its immediate substrates AcCoA or MalCoA, but rather by activities of DNL enzymes.

molecular biology↗

bioRxiv2022

Yersinia pestis genomes reveal plague in Britain 4,000 years ago

Extinct lineages of Yersinia pestis, the causative agent of the plague, have been identified in several individuals from Central Europe and Asia between 5,000 and 3,500 years before present (BP). One of these, the LNBA lineage (Late Neolithic and Bronze Age), has been suggested to have spread into Central Europe with human groups expanding from the Eurasian steppes. Here, we show that LNBA plague was spread to Europes northwestern periphery by sequencing Yersinia pestis genomes from two individuals dating to ~4,000 cal BP from an unusual mass burial context in Somerset, England, UK. This represents the earliest evidence of plague in Britain documented to date. These British Yersinia pestis genomes belong to a sublineage previously observed in two Bronze Age individuals from Central Europe that had lost the putative virulence factor yapC. This sublineage is later found in Central Asia ~3,600 BP. While the severity of disease is currently unclear, the wide geographic distribution within a few centuries suggests substantial transmissibility.

evolutionary biology↗

bioRxiv2021

Multiplexed identification of RAS paralog imbalance as a driver of lung cancer growth

Oncogenic KRAS mutations occur in approximately 30% of lung adenocarcinoma. Despite several decades of effort, oncogenic KRAS-driven lung cancer remains difficult to treat, and our understanding of the positive and negative regulators of RAS signaling is incomplete. To uncover the functional impact of diverse KRAS-interacting proteins on lung cancer growth in vivo, we used multiplexed somatic CRISPR/Cas9-based genome editing in genetically engineered mouse models with tumor barcoding and high-throughput barcode sequencing. Through a series of CRISPR/Cas9 screens in autochthonous lung tumors, we identified HRAS and NRAS as key suppressors of KRASG12D-driven tumor growth in vivo and confirmed these effects in oncogenic KRAS-driven human lung cancer cell lines. Mechanistically, RAS paralogs interact with oncogenic KRAS, suppress KRAS-KRAS interactions, and reduce downstream ERK signaling. HRAS mutations identified in KRAS-driven human tumors partially abolished this effect. Comparison of the tumor-suppressive effects of HRAS and NRAS in KRAS- and BRAF-driven lung cancer models confirmed that RAS paralogs are specific suppressors of oncogenic KRAS-driven lung cancer in vivo. Our study outlines a technological avenue to uncover positive and negative regulators of oncogenic KRAS-driven cancer in a multiplexed manner in vivo and highlights the role of RAS paralog imbalance in oncogenic KRAS-driven lung cancer.

cancer biology↗

bioRxiv2021

Microbiome diversity and host immune functions may define the fate of sponge holobionts under future ocean conditions

The sponge-associated microbial community contributes to the overall health and adaptive capacity of the sponge holobiont. This community is regulated by the environment, as well as the immune system of the host. However, little is known about the effect of environmental stress on the regulation of host immune functions and how this may, in turn, affect sponge-microbe interactions. In this study, we compared the microbiomes and immune repertoire of two sponge species, the demosponge, Neopetrosia compacta and the calcareous sponge, Leucetta chagosensis, under varying levels of acidification and warming stress. Neopetrosia compacta harbors a diverse bacterial assemblage and possesses a rich repertoire of scavenger receptors while L. chagosensis has a less diverse microbiome and an expanded range of pattern recognition receptors and proteins with immunological domains. Upon exposure to warming and acidification, the microbiome and host transcriptome of N. compacta remained stable, which correlated with high survival. In contrast, the bacterial community of L. chagosensis exhibited drastic restructuring and widespread downregulation of host immune-related pathways, which accompanied tissue necrosis and mortality. Differences in microbiome diversity and immunological repertoire of diverse sponge groups highlight the central role of host-microbe interactions in predicting the fate of sponges under future ocean conditions.

microbiology↗

bioRxiv2020

CRISPR-enhanced human adipocyte 'browning' as cell therapy for metabolic disease

Obesity and type 2 diabetes (T2D) are associated with poor tissue responses to insulin1,2, disturbances in glucose and lipid fluxes3-5 and comorbidities including steatohepatitis6 and cardiovascular disease7,8. Despite extensive efforts at prevention and treatment9,10, diabetes afflicts over 400 million people worldwide11. Whole body metabolism is regulated by adipose tissue depots12-14, which include both lipid-storing white adipocytes and less abundant "brown" and "brite/beige" adipocytes that express thermogenic uncoupling protein UCP1 and secrete factors favorable to metabolic health15-18. Application of clustered regularly interspaced short palindromic repeats (CRISPR) gene editing19,20 to enhance "browning" of white adipose tissue is an attractive therapeutic approach to T2D. However, the problems of cell-selective delivery, immunogenicity of CRISPR reagents and long term stability of the modified adipocytes are formidable. To overcome these issues, we developed methods that deliver complexes of SpyCas9 protein and sgRNA ex vivo to disrupt the thermogenesis suppressor gene NRIP121,22 with near 100% efficiency in human or mouse adipocytes. NRIP1 gene disruption at discrete loci strongly ablated NRIP1 protein and upregulated expression of UCP1 and beneficial secreted factors, while residual Cas9 protein and sgRNA were rapidly degraded. Implantation of the CRISPR-enhanced human or mouse brown-like adipocytes into high fat diet fed mice decreased adiposity and liver triglycerides while enhancing glucose tolerance compared to mice implanted with unmodified adipocytes. These findings advance a therapeutic strategy to improve metabolic homeostasis through CRISPR-based genetic modification of human adipocytes without exposure of the recipient to immunogenic Cas9 or delivery vectors.

bioengineering↗

bioRxiv2020

Humans surviving cholera develop antibodies against Vibrio cholerae O-specific polysaccharide that inhibit pathogen motility

The mechanism of protection against cholera afforded by previous illness or vaccination is currently unknown. We have recently shown that antibodies targeting O-specific polysaccharide (OSP) of Vibrio cholerae correlate highly with protection against cholera. V. cholerae is highly motile and possesses a flagellum sheathed in O-specific polysaccharide (OSP), and motility of V. cholerae correlates with virulence. Using high speed video microscopy, and building upon previous animal-related work, we demonstrate that sera, polyclonal antibody fractions, and OSP-specific monoclonal antibodies recovered from humans surviving cholera block V. cholerae motility at both subagglutinating and agglutinating concentrations. This anti-motility effect is reversed by pre-adsorbing sera and polyclonal antibody fractions with purified OSP; and is associated with OSP-specific but not flagellin-specific monoclonal antibodies. F[ab] fragments of OSP-specific polyclonal antibodies do not inhibit motility, suggesting a requirement for antibody-mediated crosslinking in motility inhibition. We show that OSP-specific antibodies do not directly affect V. cholerae viability, but that OSP-specific monoclonal antibody highly protects against death in the murine cholera model. We used in vivo competitive index studies to demonstrate that OSP-specific antibodies impede colonization and survival of V. cholerae in intestinal tissues, and that this impact is motility-dependent. Our findings suggest that the impedance of motility by antibodies targeting V. cholerae OSP contributes to protection against cholera. IMPORTANCECholera is a severe dehydrating illness of humans caused by Vibrio cholerae. V. cholerae is a highly motile bacterium that has a single flagellum covered in lipopolysaccharide (LPS) displaying O-specific polysaccharide (OSP), and V. cholerae motility correlates with its ability to cause disease. The mechanisms of protection against cholera are not well understood; however, since V. cholerae is a non-invasive intestinal pathogen, it is likely that antibodies that bind the pathogen or its products in the intestinal lumen contribute to protection from infection. Here, we demonstrate that OSP-specific antibodies isolated from humans surviving cholera in Bangladesh inhibit V. cholerae motility and are associated with protection against challenge in a motility-dependent manner.

microbiology↗

bioRxiv2020

Single-cell atlas of tumor clonal evolution in liver cancer

Tumor evolution is a key feature of tumorigenesis and plays a pivotal role in driving intratumor heterogeneity, treatment failure and patients prognosis. Here we performed single-cell transcriptome profiling of 46 primary liver cancers from 37 patients enrolled for interventional studies. We surveyed the landscape of ~57,000 malignant and non-malignant cells and determined tumor cell clonality by developing a machine learning-based consensus clustering method. We found evidence of tumor cell branching evolution using hierarchical clustering, RNA velocity as well as reverse graph embedding methods. Interestingly, an increasing tumor cell clonality was tightly linked to patients prognosis, accompanied by a polarized immune cell landscape. We identified osteopontin as a key player for tumor cell evolution and microenvironmental reprogramming. Our study offers insight into the collective behavior of tumor cell communities in liver cancer as well as potential drivers for tumor evolution in response to therapy.

cancer biology↗

bioRxiv2020

Single Cell RNA Profiling Reveals Adipocyte to Macrophage Signaling Sufficient to Enhance Thermogenesis

The "browning" of inguinal white adipose tissue (iWAT) through increased abundance of thermogenic beige/brite adipocytes is induced by cold exposure and many other perturbations in association with beneficial systemic metabolic effects. Adipose browning is reported to require activation of sympathetic nerve fibers (SNF), aided by alternately activated macrophages within iWAT. Here we demonstrate the first example of a non-cell autonomous pathway for iWAT browning that is fully independent of SNF activity. Thus, the strong induction of thermogenic adipocytes prompted by deletion of adipocyte fatty acid synthase (iAdFASNKO mice) was unaffected by denervation or the deletion of SNF modulator Neuregulin-4. However, browning of iWAT in iAdFASNKO mice does require adipocyte cAMP/protein kinase A signaling, as it was blocked in adipocyte- selective Fasn/Gs double KO mice. Single-cell transcriptomic analysis of iAdFASNKO mouse adipose stromal cells revealed increased macrophages displaying gene expression signatures of the alternately activated type. Mechanistically, depletion of such phagocytic immune cells in iAdFASNKO mice fully abrogated appearance of thermogenic adipocytes in iWAT. Altogether, these findings reveal an unexpected pathway of cAMP/PKA-dependent iWAT browning that is initiated by adipocyte signals and caused by macrophage-like cells independent of sympathetic neuron involvement.

physiology↗

bioRxiv2020

Rapid event-related, BOLD, NHP: choose two out of three

Human functional magnetic resonance imaging (fMRI) typically employs the blood-oxygen-level-dependent (BOLD) contrast mechanism. In non-human primates (NHP), contrast enhancement is possible using monocrystalline iron-oxide nanoparticles (MION) contrast agent, which has a more temporally extended response function. However, using BOLD fMRI in NHP is desirable for interspecies comparison, and the faster response of the BOLD signal promises to be beneficial to rapid event-related (rER) designs. Here, we used rER BOLD fMRI in macaque monkeys while viewing real-world images, and found visual responses and category-selectivity consistent with previous studies. However, activity estimates were very noisy, suggesting that the lower contrast-to-noise ratio of BOLD, suboptimal behavioural performance, and motion artefacts, in combination, render rER BOLD fMRI challenging in NHP. Previous studies have shown that rER monkey fMRI is possible with MION, despite its prolonged response function. To understand this, we conducted simulations of the BOLD and MION response during rER designs, and found that no matter how fast the design, the greater amplitude of the MION response outweighs the contrast loss caused by greater temporal smoothing. We conclude that although any two of the three elements (rER, BOLD, NHP) have been shown to work well, the combination of all three is particularly challenging.

neuroscience↗

bioRxiv2019

Distinct roles of BRCA2 in replication fork protection in response to hydroxyurea and DNA interstrand crosslinks

DNA interstrand crosslinks (ICLs) are a form of DNA damage that requires the interplay of a number of repair proteins including those of the Fanconi anemia (FA) and the homologous recombination (HR) pathways. Pathogenic variants in the essential gene BRCA2/FANCD1, when monoallelic, predispose to breast and ovarian cancer, and when biallelic, results in a severe subtype of Fanconi anemia. BRCA2 function in the FA pathway is attributed to its role as a mediator of the RAD51 recombinase in HR repair of the programmed DNA double strand breaks (DSB). BRCA2 and RAD51 functions are also required to protect stalled replication forks from nucleolytic degradation during response to hydroxyurea (HU). While RAD51 has been shown to be necessary in the early steps of ICL repair to prevent aberrant nuclease resection, the role of BRCA2 in this process has not been described. Here, based on the analysis of BRCA2 DNA binding domain (DBD) mutants discovered in FA patients presenting with atypical FA-like phenotypes, we establish that BRCA2 is necessary for protection of DNA at an ICL. Cells carrying DBD BRCA2 mutations are sensitive to ICL inducing agents but resistant to HU treatment consistent with relatively high HR repair in these cells. BRCA2 function at an ICL protects against DNA2-WRN nuclease-helicase complex and not the MRE11 nuclease implicated in the resection of HU-stalled replication forks. Our results also indicate that unlike the processing at HU-stalled forks, function of the SNF2 translocases (SMARCAL1, ZRANB3, or HLTF), implicated in fork reversal, are not an integral component of the ICL repair, pointing to a different mechanism of fork protection at different DNA lesions.

molecular biology↗

bioRxiv2018

Resistance to Diet Induced Obesity in the Apolipoprotein E Deficient Mouse is Associated with an Attenuated Transcriptional Response in Visceral Fat

The apolipoprotein E knockout (EKO) mouse is a well-established model of atherosclerosis. Macrophages in the arterial intima of EKO mice serve a protective role, scavenging oxidatively modified LDL in order to protect cells from toxic free cholesterol. Recent studies have highlighted a similar role for macrophage foam cells in restraining the increased rates of lipolysis in adipose tissue of obese and fasting mice. Interestingly, EKO adipocytes have been shown to have increased rates of lipolysis in vitro. Therefore, the aim of this study was to examine how apoE deficiency might alter the transcriptional response of visceral adipose tissue (VAT) to high fat diet (HFD). EKO mice fed HFD for 24 weeks gained less fat mass and were more insulin sensitive than their wild type (WT) littermates. Metabolic cages showed that HFD EKO mice had increased post-prandial oxygen consumption and increased serum {beta}-hydroxybutyrate. DNA microarrays revealed that EKO VAT was comparatively insensitive to HFD in terms of alterations in gene expression, with only 0.1% of probe sets differentially expressed. In contrast, the VAT of WT mice had a 30 fold more extensive alteration in gene expression (3% of probes sets), characterized predominantly by increased expression of immune cell specific genes. In addition, analysis of a priori determined gene sets revealed broad down-regulation of PPAR{gamma} target and fatty acid catabolism genes in WT VAT, and increased expression of lipid storage and cholesterol synthesis genes. In comparison, expression of PPAR{gamma} target genes was not down-regulated in EKO VAT and expression of fatty acid oxidation genes was increased. In summary, we report three novel findings with regards to metabolism in the EKO mouse: 1) increased post-prandial oxygen consumption, 2) increased serum {beta} hydroxybutyrate concentrations and 3) a dramatically less robust transcriptional response to HFD in EKO VAT. These findings suggest that limiting adipocyte exposure to dietary fatty acids may be an attractive therapy for diet induced obesity, provided that compensatory mechanisms that prevent hyperlipidemia can be activated.

physiology↗

bioRxiv2018

Point of care influenza testing using Alere-i Influenza A & B assay: a practical assessment

We assessed the utility of the Alere i Influenza A & B point of care influenza test (Ai-POCIT) with laboratory testing using RT-PCR. 270 adult hospital patients had both Ai-POCIT and laboratory influenza tests conducted on the same sample. Overall, 30% and 32% influenza tests were positive by Ai-POCIT and RT-PCR, respectively. The sensitivity of the Ai-POCIT for influenza A, influenza B and any influenza were 93%, 100%, and 95%, respectively. Specificity was 100% for both viruses, but an 11% test failure rate indicates the need for better training of users. We believe that the use of nasopharyngeal (NP) swabs resulted in the observed high performance of the Ai-POCIT in comparison to other published studies. Ai-POCIT was regarded as very useful by front line clinical staff for clinical decision making and acute bed management.

microbiology↗