Biology subjects

Smyth, D.

Publications and source records attributed to Smyth, D..

7 recordsLinked to original sources

bioRxiv2023

Exogenous TGFβ1 and its mimic HpTGM attenuate the heart's inflammatory response to ischaemic injury and improve long term cardiac outcomes

RationaleSuccessful and timely coronary reperfusion following acute ST-elevation myocardial infarction (STEMI) is standard therapy to salvage transiently ischaemic heart muscle. However, the subsequent inflammatory response within the infarct can lead to further loss of viable myocardium. Robust interventions are required in the acute MI setting to minimise cardiac injury and reduce risk of further detrimental progression. ObjectiveTGF{beta}1 is an anti-inflammatory cytokine released endogenously in response to infection or tissue injury. The goal of this study was to investigate its protective effects when given exogenously following myocardial infarction. Methods and ResultsTGF{beta}1 is found at increased levels in the blood of STEMI patients immediately following myocardial infarction. We observe a significant correlation (p=0.003) between higher circulating TGF{beta}1 levels at 24h post MI and a reduction in infarct size over the following 3 months, suggesting that an early increase in circulating TGF{beta}1 is protective in these patients. Using a mouse model of cardiac ischaemia-reperfusion we demonstrate that additional exogenous TGF{beta}1 delivered in the acute setting has multiple beneficial outcomes. At 24 hours post-reperfusion It leads to a significantly smaller infarct size (30% reduction, p=0.025), reduced inflammatory infiltrate (28% reduction, p=0.015), lower intra-cardiac expression of inflammatory cytokines IL1{beta} and CCL2 (>50 % reduction, p=0.038 and 0.0004, respectively) and reduced scar size at 4 weeks (21% reduction, p=0.015). Furthermore exogenous delivery of an equivalent dose of HpTGM, a recently described low-fibrogenic mimic of TGF{beta}1, secreted by a helminth parasite to evade immune rejection, has an almost identical protective effect on injured mouse hearts. Furthermore using a genetic approach we show the benefit is mediated by the vascular endothelium. ConclusionsThis work reveals the potential of exogenous TGF{beta}1 and HpTGM delivered in the acute MI setting to provide protective anti-inflammatory effects and reduce infarct size, leading to a smaller scar and reduced detrimental progression.

pathology↗

bioRxiv2021

Metabolic regulation by prostaglandin E2 impairs lung group 2 innate lymphoid cell responses

Group 2 innate lymphoid cells (ILC2s) play a critical role in asthma pathogenesis. Non-steroidal anti-inflammatory drug (NSAID)-exacerbated respiratory disease (NERD) is associated with reduced signaling via EP2, a receptor for prostaglandin E2 (PGE2). However, the respective roles for the PGE2 receptors EP2 and EP4 (both share same downstream signaling) in the regulation of lung ILC2 responses has yet been deciphered. Here, we find that deficiency of EP2 rather than EP4 augments IL-33-induced lung ILC2 responses and eosinophilic inflammation in vivo. In contrast, exogenous agonism of EP4 but not EP2 markedly restricts IL-33- and Alternaria alternata-induced lung ILC2 responses and eosinophilic inflammation. Mechanistically, PGE2 directly suppresses IL-33-dependent ILC2 activation through the EP2/EP4-cAMP pathway, which downregulates STAT5 and MYC pathway gene expression and ILC2 energy metabolism. Blocking glycolysis diminishes IL-33-dependent ILC2 responses in mice lacking endogenous PG synthesis but not in PG-competent mice. Together, we have defined a mechanism for optimal suppression of lung ILC2 responses by endogenous PGE2-EP2 signaling which underpins the clinical findings of defective EP2 signaling in patients with NERD. Our findings also indicate that exogenously targeting the PGE2-EP4-cAMP and energy metabolic pathways may provide novel opportunities for treating ILC2-initiated lung inflammation in asthma and NERD. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=178 SRC="FIGDIR/small/474031v1_ufig1.gif" ALT="Figure 1"> View larger version (45K): org.highwire.dtl.DTLVardef@17dd2caorg.highwire.dtl.DTLVardef@199ce97org.highwire.dtl.DTLVardef@ca1f67org.highwire.dtl.DTLVardef@1d3a49f_HPS_FORMAT_FIGEXP M_FIG C_FIG Schematic of potential roles for activation of EP2 and EP4 by endogenous versus exogenous ligands in regulation of lung ILC2 immune responses. Endogenous PGE2 in the lung preferentially activates EP2 rather than EP4 to inhibit ILC2 responses and eosinophilic inflammation, and ablation of EP2 enhances lung ILC2 responses. Conversely, lung ILC2 responses are not altered by EP4 deficiency. However, they are markedly inhibited by EP4 agonism but not EP2 agonism. Mechanistically, PGE2-EP2/EP4 signaling activates the cAMP pathway which inhibits ILC2 energy metabolism, possibly through interruption of NF-{kappa}B (reported in Nagashima H, et al. Immunity 2019;51:682-695) and STAT5 signaling, leading to decline of ILC2 survival, proliferation and type 2 cytokine production.

immunology↗

bioRxiv2021

Immune disease risk variants regulate gene expression dynamics during CD4+ T cell activation

During activation, T cells undergo extensive changes in gene expression which shape the properties of cells to exert their effector function. Therefore, understanding the genetic regulation of gene expression during T cell activation provides essential insights into how genetic variants influence the response to infections and immune diseases. We generated a single-cell map of expression quantitative trait loci (eQTL) across a T cell activation time-course. We profiled 655,349 CD4+ naive and memory T cells, capturing transcriptional states of unstimulated cells and three time points of cell activation in 119 healthy individuals. We identified 38 cell clusters, including stable clusters such as central and effector memory T cells and transient clusters that were only present at individual time points of activation, such as interferon-responding cells. We mapped eQTLs using a T cell activation trajectory and identified 6,407 eQTL genes, of which a third (2,265 genes) were dynamically regulated during T cell activation. We integrated this information with GWAS variants for immune-mediated diseases and observed 127 colocalizations, with significant enrichment in dynamic eQTLs. Immune disease loci colocalized with genes that are involved in the regulation of T cell activation, and genes with similar functions tended to be perturbed in the same direction by disease risk alleles. Our results emphasize the importance of mapping context-specific gene expression regulation, provide insights into the mechanisms of genetic susceptibility of immune diseases, and help prioritize new therapeutic targets.

genomics↗

bioRxiv2020

Structure-based mapping of the TβRI and TβRII receptor binding sites of the parasitic TGF-β mimic, Hp-TGM

TGF-{beta} is a secreted signaling protein involved in many physiological processes: organ development, production and maintenance of the extracellular matrix, as well as regulation of the adaptive immune system. As a cytokine, TGF-{beta} stimulates the differentiation of CD4+ T-cells into regulatory T-cells (Tregs) that act to promote peripheral immune tolerance. The murine parasite Heligmosomoides polygyrus takes advantage of this pathway to induce inducing Foxp3+ Tregs in a similar manner using a TGF-{beta} mimic (TGM), comprised of five tandem complement control protein (CCP) domains, designated D1-D5. Despite having no structural homology to TGF-{beta} or to TGF-{beta} family proteins, TGM binds directly to the TGF-{beta} type I and type II receptors, T{beta}RI and T{beta}RII. To further investigate, NMR titration, and SPR and ITC binding experiments were performed, showing that TGM-D2, with the aid of D1, binds T{beta}RI and TGM-D3 binds T{beta}RII. Competition ITC experiments showed that TGM-D3 competes with TGF-{beta} for binding to T{beta}RII, consistent with TGM-D3-induced NMR chemical shift perturbations of T{beta}RII which aligned with the solvent inaccessible areas of T{beta}RII upon binding TGF-{beta}. Thus, TGM-D3 binds to the same edged {beta}-strand of T{beta}RII that is used to bind TGF-{beta}. Competition ITC experiments demonstrated that TGM-D1D2 and TGF-{beta}3:T{beta}RII compete for binding to T{beta}RI, while TGM-D2-induced NMR chemical shift perturbation of T{beta}RI showed that TGM-D2 binds to the same pre-helix extension of T{beta}RI as does the TGF-{beta}/T{beta}RII binary complex. The solution structure of TGM-D3 revealed that while it has the overall structure of a CCP domain, TGM-D3 has an insertion in the hypervariable loop uncommon to CCP domains. These findings suggest that parasitic TGM, despite its lack of structural similarity to TGF-{beta}, evolved to take advantage of the binding regions of the mammalian TGF-{beta} type I and type II receptors. The structure of this TGM domain, along with the predicted structure of other H. polygyrus secreted proteins reported in the literature, suggest that TGM is part of a larger family of evolutionarily-adapted immunomodulatory CCP-containing proteins.

biophysics↗

bioRxiv2018

A macrophage-pericyte axis directs tissue restoration via Amphiregulin-induced TGFβ activation

The Epidermal Growth Factor Receptor ligand Amphiregulin has a well-documented role in the restoration of tissue homeostasis following injury; however, the mechanism by which Amphiregulin contributes to wound repair remains unknown. Here we show that Amphiregulin functions by releasing bio-active TGFb from latent complexes via integrin-v activation. Using acute injury models in two different tissues, we found that by inducing TGFb activation on mesenchymal stromal cells (aka pericytes), Amphiregulin induced their differentiation into myo-fibroblasts, thereby selectively contributing to the restoration of vascular barrier function within injured tissue. Furthermore, we identified macrophages as a critical source of Amphiregulin, revealing a direct effector mechanism by which these cells contribute to tissue restoration following acute injury. Combined, these observations expose a so far under-appreciated mechanism of how cells of the immune system selectively control the differentiation of tissue progenitor cells during tissue repair and inflammation.

immunology↗

bioRxiv2018

Microbiome inhibition of IRAK-4 by trimethylamine mediates metabolic and immune benefits in high-fat-diet-induced insulin resistance

The global type 2 diabetes epidemic is a major health crisis and there is a critical need for innovative strategies to fight it. Although the microbiome plays important roles in the onset of insulin resistance (IR) and low-grade inflammation, the microbial compounds regulating these phenomena remain to be discovered. Here, we reveal that the microbiome inhibits a central kinase, eliciting immune and metabolic benefits. Through a series of in vivo experiments based on choline supplementation, blocking trimethylamine (TMA) production then administering TMA, we demonstrate that TMA decouples inflammation and IR from obesity in the context of high-fat diet (HFD) feeding. Through in vitro kinome screens, we reveal TMA specifically inhibits Interleukin-1 Receptor-associated Kinase 4 (IRAK4), a central kinase integrating signals from various toll-like receptors and cytokine receptors. TMA blunts TLR4 signalling in primary human hepatocytes and peripheral blood monocytic cells, and improves mouse survival after a lipopolysaccharide-induced septic shock. Consistent with this, genetic deletion and chemical inhibition of IRAK4 result in similar metabolic and immune improvements in HFD. In summary, TMA appears to be a key microbial compound inhibiting IRAK4 and mediating metabolic and immune effects with benefits upon HFD. Thereby we highlight the critical contribution of the microbial signalling metabolome in homeostatic regulation of host disease and the emerging role of the kinome in microbial-mammalian chemical crosstalk.

systems biology↗

bioRxiv2018

The clinician impact and financial cost to the NHS of litigation over pregabalin: an economic impact analysis

ObjectivesFollowing litigation over pregabalins second-use medical patent for neuropathic pain NHS England were required by the court to instruct GPs to prescribe the branded form (Lyrica) for pain. Pfizers patent was found invalid in 2015; a ruling subject to ongoing appeals. If the Supreme Court appeal in February 2018 is unsuccessful, the NHS can reclaim excess prescribing costs. We set out to describe the variation in prescribing of pregabalin as branded Lyrica, geographically and over time; to determine how clinicians responded to the NHS England instruction to GPs; and to model excess costs to the NHS attributable to the legal judgments.\n\nSettingEnglish primary care\n\nParticipantsEnglish general practices\n\nPrimary and secondary outcome measuresVariation in prescribing of branded Lyrica across the country before and after the NHS England instruction, by practice and by Clinical Commissioning Group (CCG); excess prescribing costs.\n\nResultsThe proportion of pregabalin prescribed as Lyrica increased, from 0.3% over six months before the NHS England instruction (September 2014-February 2015) to 25.7% afterwards (April - September 2015). Although 70% of pregabalin is estimated to be for neuropathic pain, only 11.6% of practices prescribed Lyrica at this level; the median proportion prescribed as Lyrica was 8.8% (IQR 1.1-41.9%). If pregabalin had come entirely off patent in September 2015, and Pfizer had not appealed, we estimate the NHS would have spent {pound}502m less on pregabalin to July 2017.\n\nConclusionNHS England instructions to GPs regarding branded prescription of pregabalin were widely ignored, and have created much debate around clinical independence in prescribing. Protecting revenue from \"skinny labels\" will pose a challenge. If Pfizers final appeal on the patent is unsuccessful the NHS can seek reimbursement of excess pregabalin prescribing costs, potentially {pound}502m.

epidemiology↗