Biology subjects

Greaves, D. R.

Publications and source records attributed to Greaves, D. R..

7 recordsLinked to original sources

bioRxiv2025

Hypercholesterolaemia promotes epigenetic memory in hematopoietic stem cells that persists after lipid lowering and causes systemic immunometabolic dysfunction mediated by macrophage metabolic reprogramming

In patients with atherosclerotic cardiovascular disease (ASCVD) lowering LDL cholesterol (LDL-C) reduces the risk of adverse cardiovascular events. However, these patients remain at continued risk of adverse cardiovascular events despite achieving optimal LDL-C lowering. We hypothesised that this residual risk is mediated by epigenetic programming of haematopoietic stem cells (HSC) which persists after cholesterol levels are lowered and results in sustained effects on innate immune cell metabolism, that is not resolved by cholesterol lowering. We found that exposure to high cholesterol (HC) in vivo induced long-term metabolic changes in macrophages differentiated ex vivo from bone marrow, leading to a pro-inflammatory profile; these changes persisted in vivo despite cholesterol lowering. HSC from HC mice had altered chromatin accessibility that persisted after cholesterol lowering and was also present in bone marrow monocytes and tissue resident macrophages. HC provoked RUNX1-dependent downregulation of stearoyl-CoA desaturase (SCD) which reduced mono-unsaturated fatty acid (MUFA) availability for OxPhos in murine and human monocytes and macrophages. Supplementation with MUFA restored OxPhos capacity and promoted a shift towards a less pro-inflammatory macrophage phenotype in HC-trained BMDM. Bone marrow chimera and lineage tracking studies revealed that prior exposure of HSC to HC conferred adverse systemic metabolic effects on normocholesterolemic mice, with increased adipose tissue mass and increased migration of macrophages derived from HC-exposed HSC into adipose tissue, resulting in increased adipose tissue inflammation and systemic glucose intolerance. These findings indicate that HC results in long-lasting immuno-epigenetic memory in HSC which is refractory to lipid lowering, and provide strong evidence that exposure to HC can have prolonged consequences that require new therapeutic approaches, beyond cholesterol lowering.

immunology↗

bioRxiv2025

Steric control of signaling bias in the immunometabolic receptor GPR84

Biased signaling in G protein-coupled receptors offers therapeutic promise, yet rational design of biased ligands remains challenging due to limited mechanistic understanding. Here, we report a molecular framework for controlling signaling bias at the immunometabolic receptor GPR84. We identified three structurally-matched ligands (OX04529, OX04954, and OX04539) with varying steric profiles that exhibit comparable Gi protein activation but dramatically different {beta}-arrestin recruitment capacities. A high- resolution cryo-EM structure of GPR84-Gi in complex with OX04529, complemented by molecular dynamics simulations and targeted mutagenesis, revealed that steric interactions between ligand substituents and Leu3366.52 and Phe1875.47 indirectly disrupt a critical polar network involving Tyr3326.48, Asn1043.36 and Asn3627.45 essential for {beta}-arrestin recruitment. Based on these insights, we developed a steric-dependent model that enabled rational design of G protein-biased agonists with predictable {beta}-arrestin recruitment profiles. This mechanistic framework provides a blueprint for designing biased agonists with customized signaling profiles at GPR84 and potentially other class A GPCRs.

pharmacology and toxicology↗

bioRxiv2023

Distinct adrenal gland macrophages regulate corticosteroid production

The adrenal glands are hormone secreting glands that sit on top of the kidneys. Adrenal glands produce glucocorticoids, mineralocorticoids, and catecholamines, and are therefore critical regulators of the stress response, the immune response, metabolism, and blood pressure. Despite being identified for more that 30 years, our understanding of adrenal macrophages remains incomplete. In numerous other tissues, macrophages carry out a plethora of physiological and homeostatic roles in addition to their classical immune functions. The aim of this study was to characterise the macrophage compartment of the adrenal gland and assess its contribution to adrenal function. Using an in vivo approach, we herein describe two morphologically and spatially distinct subsets of adrenal macrophages - dendritic-like macrophages that are present throughout the gland in young and old mice, and "foamy" lipid-laden macrophages that accumulate in the murine adrenal cortex in an age and diet-dependent manner. Furthermore, we present data showing that these foamy-like macrophages accumulate cholesterol and thereby regulate adrenal hormonal output, at steady state and in the context of obesity. We hereby provide novel insights into the physiological roles of macrophages in the adrenal gland and the mechanisms by which adrenal hormone production is regulated.

immunology↗

bioRxiv2023

The mechanical arthropod vector Stomoxys calcitrans influences the outcome of lumpy skin disease virus infection in cattle

The poxvirus lumpy skin disease virus (LSDV) is the etiological agent of lumpy skin disease (LSD), a severe disease of cattle and water buffalo that is characterised by numerous necrotic cutaneous nodules. LSD is a rapidly emerging disease, spreading into and across the Middle East, eastern Europe, and Asia in the past decade. The disease causes substantial production and economic losses in rural communities and affected regions. LSDV is mechanically transmitted by haematophagous arthropods including stable flies (Stomoxys calcitrans), however our understanding of this mechanical transmission method is sparse. A secreted saliva collection methodology using a modified artificial membrane feeding system was optimised for S. calcitrans and used to collect and characterise secreted S. calcitrans saliva. Saliva was mixed with LSDV and shown not to affect virus growth in primary bovine fibroblasts. S. calcitrans saliva or spot-feeding by S. calcitrans was then incorporated into a bovine in vivo experimental model of LSD to determine if either influenced disease pathogenesis. S. calcitrans saliva resulted in fewer animals developing disease, however this difference was not statistically significant. Spot-feeding with S. calcitrans prior to inoculation did not alter the number of animals that developed disease or the overall severity of disease however disease progression was accelerated as demonstrated by the appearance of cutaneous nodules, detection of viral DNA in the blood stream, and production of neutralising antibodies. This shows that S. calcitrans influence disease kinetics through co-incident bite trauma and/or saliva deposition. This increases our understanding of LSDV pathogenesis and highlights the overlooked importance of mechanical vectors in pathogen transmission. Author summaryInsect vectors are important conduits for the transmission of pathogens that cause diseases such as Zika, dengue, malaria, and lumpy skin disease. Biological vector-borne transmission incorporates a replication phase for the pathogen in the insect, whereas no replication occurs in the vector during mechanical transmission. When the insect bites the host it inoculates a pathogen whilst also delivering arthropod-derived factors such as saliva components and causing tissue trauma through biting and probing. Arthropod saliva and/or bite trauma have been shown to enhance the speed and severity of disease following inoculation with a range of biologically transmitted viruses. This study examined if this was true also for the mechanically transmitted pathogen lumpy skin disease virus (LSDV). LSDV is a neglected pathogen that causes severe systemic disease in cattle and is transmitted mechanically by the stable fly Stomoxys calcitrans. Using an experimental bovine model of LSD, we found that disease occurred more rapidly when virus was delivered in association with the bites of uninfected flies. This work has increased our knowledge of lumpy skin disease virus transmission, and the discovery that disease outcome can be impacted by previously overlooked mechanical insect vectors should prompt further investigation into this mechanism of transmission.

microbiology↗

bioRxiv2020

Tissue-resident macrophages regulate lymphatic vessel growth and patterning in the developing heart

Macrophages are components of the innate immune system with key roles in tissue inflammation and repair. It is now evident that macrophages also support organogenesis, but few studies have characterized their identity, ontogeny and function during heart development. Here, we show that resident macrophages in the subepicardial compartment of the developing heart coincide with the emergence of new lymphatics and interact closely with the nascent lymphatic capillaries. Consequently, global macrophage-deficiency led to extensive vessel disruption with mutant hearts exhibiting shortened and mis-patterned lymphatics. The origin of cardiac macrophages was linked to the yolk sac and fetal liver. Moreover, Csf1r+ and Cx3cr1+ myeloid sub-lineages were found to play essential functions in the remodeling of the lymphatic endothelium. Mechanistically, macrophage hyaluronan was found to be required for lymphatic sprouting by mediating direct macrophage-lymphatic endothelial cell interactions. Together, these findings reveal insight into the role of macrophages as indispensable mediators of lymphatic growth during the development of the mammalian cardiac vasculature.Summary statement Tissue-resident macrophages are indispensable mediators of lymphatic vessel formation during heart development and function to remodel the vascular plexus.View Full Text

developmental biology↗

bioRxiv2019

Inhibition of Bruton’s tyrosine kinase reduces NF-kB and NLRP3 inflammasome activity preventing insulin resistance and microvascular disease

Activation of inflammatory pathways in myeloid cells initiates insulin resistance leading to the development of type-2 diabetes and microvascular disease. Currently, there are no therapies available that target inflammation in T2D or microvascular disease. In the present study we investigate if Brutons tyrosine kinase (BTK) may represent a novel therapeutic target using the FDA approved medication ibrutinib. Ibrutinib treatment protected high fat diet (HFD)-fed mice from developing insulin resistance and improved glycaemic control by restoring signalling through IRS-1/Akt/GSK-3{beta} pathway. These improvements were independent of body weight and calorific intake. Treatment with ibrutinib to mice fed a HFD reduced NF-{kappa}B and reduced inflammatory gene expression, this was coupled with decreased activation of the NLRP3 inflammasome in the diabetic liver and kidney. Ibrutinib treatment also protected mice from the development of diabetic nephropathy by reducing monocyte/macrophage infiltration due to reduced expression of the pro-inflammatory chemokines. Ibrutinib treatment to human monocyte derived macrophages significantly reduced pro-inflammatory gene expression and a significant reduction in IL-1{beta} and TNF after LPS stimulation. In the present study we provide proof of concept evidence that BTK is a novel therapeutic target for the treatment of T2D and ibrutinib may be a candidate for drug repurposing in T2D.

systems biology↗

bioRxiv2019

Efferocytosis perpetuates substance accumulation inside macrophage populations

In both cells and animals, cannibalism can transfer harmful substances from the consumed to the consumer. Macrophages are immune cells that consume their own dead via a process called cannibalistic efferocytosis. Macrophages that contain harmful substances are found at sites of chronic inflammation, yet the role of cannibalism in this context remains unexplored. Here we take mathematical and experimental approaches to study the relationship between cannibalistic efferocytosis and substance accumulation in macrophages. Through mathematical modelling, we deduce that substances which transfer between individuals through cannibalism will concentrate inside the population via a coalescence process. This prediction was confirmed for macrophage populations inside a closed system. We used image analysis of whole slide photomicrographs to measure both latex microbead and neutral lipid accumulation inside murine bone marrow-derived macrophages (104 - 105 cells) following their stimulation into an inflammatory state ex vivo. While the total number of phagocytosed beads remained constant, cell death reduced cell numbers and efferocytosis concentrated the beads among the surviving macrophages. Since lipids are also conserved during efferocytosis, these cells accumulated lipid derived from the membranes of dead and consumed macrophages (becoming macrophage foam cells). Consequently, enhanced macrophage cell death increased the rate and extent foam cell formation. Our results demonstrate that cannibalistic efferocytosis perpetuates exogenous (e.g. beads) and endogenous (e.g. lipids) substance accumulation inside macrophage populations. As such, cannibalism has similar detrimental consequences in both cells and animals.

immunology↗