Biology subjects

McCarthy, N. E.

Publications and source records attributed to McCarthy, N. E..

7 recordsLinked to original sources

bioRxiv2025

Bacterial suppression of intestinal fungi via activation of human gut γδ T-cells

Gut symbionts condition mucosal immunity to resist infection by enteropathogens, but the specific microbes and mechanisms involved differ significantly between host species. In higher primates, bacterial metabolite HMB-PP is sensed by a specialized population of V{gamma}9V{delta}2+T-cells, which we now report can potently suppress growth of endogenous fungi in human intestinal organ cultures. In healthy intestine, HMB-PP-stimulated V{delta}2+T-cells restricted outgrowth of keystone fungus Candida albicans via a mechanism that required IL-22. In contrast, Crohns disease (CD) patients with reduced V{delta}2+T-cell numbers displayed outgrowth of C. albicans strains that readily formed toxin-producing filaments, triggered neutrophil extracellular traps, and induced macrophage IL-1{beta} release ex vivo. Genomic and proteomic analysis of the Candida isolates suggested increased tissue adhesion of CD-derived strains, which rapidly invaded the gut barrier in an intestine-on-a-chip model. These data reveal that bacterial activation of V{delta}2+T-cells suppresses fungal pathobionts in human gut via an IL-22-dependent mechanism that is dysregulated in CD.

immunology↗

bioRxiv2025

IsoDGR-Induced Endothelial Cytoskeletal Disruption Drives Age-Related Blood-Brain Barrier Breakdown

Brain aging is characterized by progressive breakdown of the blood-brain barrier (BBB), which correlates with neuroinflammation and cognitive decline. Emerging evidence implicates degenerative modifications of the vascular proteins as a key driver of BBB dysfunction. In particular, spontaneous deamidation of Asp-Gly-Arg (NGR) motifs generates isoAsp-Gly-Arg (isoDGR) sequences that structurally mimic canonical Arg-Gly-Asp (RGD) integrin-binding ligands. Here, we show that age-associated accumulation of isoDGR in the brain cortex induces endothelial cytoskeletal collapse and tight junction disorganization, leading to BBB breakdown. Using mice lacking the L-isoaspartyl repair enzyme PCMT1 (which accelerates isoDGR accumulation) and wild type aged mice, we found markedly elevated isoDGR in brain tissues accompanied by focal microhemorrhages and increased BBB permeability. Recent whole-genome sequencing suggests that a common PCMT1 variant is linked to neurodegenerative disease risk, indicating potential clinical relevance in vascular aging. Remarkably, systemic treatment with an isoDGR-neutralizing antibody largely prevented capillary breaches and leakage, and even restored barrier integrity in aged wild-type mice. To uncover the molecular mechanism, we exposed brain endothelial cells to synthetic isoDGR-peptides, which recapitulated these effects. Unbiased RNA-sequencing reinforced these findings, revealing broad transcriptomic reprogramming of cytoskeletal, cell-cell junction, inflammatory, and stress-response pathways. Functional studies demonstrated that isoDGR triggered collapse of F-actin stress fibers, disrupted junctional ZO-1 and VE-cadherin, increased monolayer permeability to macromolecules, and impaired endothelial cell migration and proliferation. IsoDGR-treated endothelial cells exhibited increased oxidative stress, upregulation of ICAM-1/VCAM-1/CCL-2, and adopted a senescent phenotype. Our results suggest that isoDGR hijacks endothelial integrin signaling to destabilize the actin cytoskeleton and tight junctions, a process that breaches the BBB and subsequently activates inflammatory and senescence programs. In summary, we identify BBB disruption via isoDGR-induced cytoskeletal dysfunction as a central pathology of vascular aging, and demonstrate that targeting isoDGR damage preserves BBB integrity and attenuates neuroinflammation.

neuroscience↗

bioRxiv2025

Anti-isoDGR Antibody Inhibits Atherosclerosis Induced by Western Diet in ApoE-/- mice

BackgroundDegenerative protein modifications (DPMs) accumulate with aging and can alter biomolecule structure and function, including via spontaneous conversion of Asn-Gly-Arg (NGR) to isoAsp-Gly-Arg (isoDGR) motifs that can bind integrins and drive chronic inflammation. Since isoDGR-modified extracellular matrix proteins are enriched in atherosclerosis and have been associated with rupture-prone plaque characteristics, we hypothesized that antibody neutralisation can inhibit key pathological features including atherosclerotic vascular plaque formation and metabolic dysfunction. MethodsWe first examined Pcmt1-/- mice which rapidly accumulate isoDGR due to lack of the corresponding repair enzyme to assess the extent of vascular protein damage. We then treated 6-8 week old atherosclerosis-prone (ApoE-/-) mice which were fed a high-fat Western diet (WD) with weekly dose of 1mg/kg isoDGR-specific monoclonal antibody (isoDGR-mAb) or isotype-matched control (while on diet) for 2 months duration. A regular chow-fed ApoE-/- group served as baseline control. Aortic atherosclerotic burden, plaque composition, systemic inflammation, lipid profiles, hepatic steatosis, and metabolic parameters (indirect calorimetry) were assessed. ResultsPcmt1-/- mice displayed extensive isoDGR deposition and degeneration of the aortic wall, linking this DPM to vascular structural damage. In the ApoE-/- mice, WD induced large aortic root plaques with abundant isoDGR and macrophage infiltration. IsoDGR-mAb treatment decreased plaque size by [~]30% with reduced lipid and collagen content (p=0.001). Furthermore, plaques in treated mice contained significantly fewer CD68+ macrophages that also exhibited limited activation. Systemically, isoDGR-mAb modified lipoprotein profiles by decreasing atherogenic VLDL/IDL/LDL cholesterol (p=0.04) while slightly increasing HDL, accompanied by a reduction in circulating inflammatory proteins. IsoDGR-mAb also protected against hepatic lipid accumulation which was reduced by [~]60% in treated animals (p<0.001), with indirect calorimetry confirming [~]30% higher oxygen consumption and energy expenditure without change in food intake or physical activity. ConclusionWe identified isoDGR as a key pathological factor involved in the progression of atherosclerosis. Remarkably, isoDGR neutralization diminished plaque inflammation and improved atherosclerotic plaque stability. Our findings support isoDGR neutralization as a promising therapeutic strategy to mitigate both atherosclerosis and aging-associated metabolic dysfunction.

pharmacology and toxicology↗

bioRxiv2023

Age-linked lung pathology is reduced by immunotherapeutic targeting of isoDGR protein damage

Advancing age is the primary risk factor for pulmonary diseases. Our investigation revealed an 8-fold increase in aging induced isoDGR-damaged proteins in lung tissue from human pulmonary fibrosis patients compared to healthy tissues, accompanied by elevated frequencies of CD68+/CD11b+ macrophages, indicating lung tissue is susceptible to time-dependent accumulation of isoDGR-proteins. To elucidate the mechanisms through which isoDGR-proteins may exacerbate aging lung disorders for potential therapeutic targeting, we assessed the functional role of this isoDGR-motif in naturally-aged mice and mice lacking the corresponding isoDGR repair enzyme (Pcmt1-/-). IsoDGR-protein accumulation in mouse lung tissue and blood vessels correlated with chronic low-grade inflammation, pulmonary edema, and hypoxemia. IsoDGR accretion induced mitochondrial and ribosomal dysfunctions, cellular senescence, and apoptosis, contributing to progressive lung damage over time. Treatment with anti-isoDGR antibodies suppressed TLR pathway activity, mitigated cytokine-driven inflammation, restored mtDNA expression, and significantly reduced lung pathology in-vivo. Similarly, exposure of lung endothelial cells to isoDGR-modified fibronectin impaired oxygen consumption, increased reactive oxygen species levels, and disrupted acidification, but these effects were efficiently reversed by target-specific antibody therapy. Collectively, our findings underscore the significant contribution of isoDGR-damaged proteins to age-linked lung pathology. IsoDGR-specific therapy emerges as a promising treatment approach for pulmonary disorders in older patients.

pathology↗

bioRxiv2020

Increased isoDGR motifs in plasma fibronectin are associated with atherosclerosis through facilitation of vascular fibrosis

Abnormal matrix deposition on vessels and recruitment of inflammatory cells into the arterial wall are critical events in atherosclerotic plaque formation. Fibronectin protein is a key matrix component that exhibits high levels of deamidation in atherosclerotic plaques and blood plasma, but it is unclear how this structural change impacts on endothelial function or modifies interactions with recruited leukocytes. This study aimed to determine how deamidation-induced isoDGR motifs in fibronectin influence extracellular matrix accumulation on endothelial cells, and to investigate possible effects on integrin outside-in signalling in matrix-bound monocytes which are key mediators of human atherosclerosis. Blood plasma fibrinogen and fibronectin displayed marked accumulation of isoDGR motifs in ischemic heart disease (IHD) as determined by ELISA analysis of patients undergoing coronary artery bypass grafting compared with age-matched healthy controls. Biochemical and functional assays confirmed that isoDGR-containing fibronectin promoted activation of integrin {beta}1 in monocytes and facilitated protein deposition and fibrillogenesis on endothelial cell layers. In addition, isoDGR interactions with integrins on the monocyte cell surface triggered an ERK:AP-1 signalling cascade that induced potent secretion of chemotactic mediators (including CCL2, CCL4, IL-8, and TNF), that promoted further leukocyte recruitment to the assembling plaque. Fibronectin deamidation forms isoDGR motifs that increase binding to {beta}1 integrins on the surface of endothelial cells and monocytes. Subsequent activation of integrin outside-in signalling pathways elicits a range of potent cytokines and chemokines that drive additional leukocyte recruitment to the developing atherosclerotic matrix and likely constitutes a key early event in progression to IHD.

biochemistry↗

bioRxiv2019

ERO1α promotes hypoxic tumour progression and is associated with poor prognosis in pancreatic cancer

Pancreatic cancer is a leading cause of mortality worldwide due to difficulty detecting early-stage disease and our poor understanding of the mediators that drive the progression of hypoxic solid tumours. We, therefore, used a heavy isotope pulse/trace proteomic approach to determine how hypoxia alters pancreatic tumour expression of proteins that confer treatment resistance, promote metastasis, and suppress host immunity. Using this method, we identified that hypoxia stress stimulates pancreatic cancer cells to rapidly translate proteins that enhance metastasis (NOTCH2, NCS1, CD151, NUSAP1), treatment resistant (ABCB6), immune suppression (NFIL3,WDR4), angiogenesis (ANGPT4, ERO1, FOS), alter cell metabolic activity (HK2, ENO2), and mediate growth-promoting cytokine responses (CLK3, ANGPTL4). Database mining confirmed that elevated gene expression of these hypoxia-induced mediators is significantly associated with poor patient survival in various stages of pancreatic cancer. Among these proteins, the oxidoreductase enzyme ERO1 was highly sensitive to induction by hypoxia stress across a range of different pancreatic cancer cell lines and was associated with particularly poor prognosis in human patients. Consistent with these data, genetic deletion of ERO1 substantially reduced growth rates and colony formation in pancreatic cancer cells when assessed in a series of functional assays in vitro. Accordingly, when transferred into a mouse xenograft model, ERO1-deficient tumour cells exhibited severe growth restriction and negligible disease progression in vivo. Together, these data indicate that ERO1 is potential prognostic biomarker and novel drug target for pancreatic cancer therapy.

cancer biology↗

bioRxiv2019

Dynamics of murine brain protein synthesis in vivo identify the hippocampus, cortex and cerebellum as highly active metabolic sites

Identification of proteins that are synthesized de novo in response to specific microenvironmental cues is critical to understanding the molecular mechanisms that underpin key physiological processes and pathologies. Here we report that a brief period of pulsed SILAC diet (Stable Isotope Labelling by Amino acids in Cell culture) enables determination of biological functions corresponding to actively translating proteins in the mouse brain. Our data demonstrate that the hippocampus, cortex and cerebellum are highly active sites of protein synthesis, rapidly expressing key mediators of nutrient sensing and lipid metabolism, as well as critical regulators of synaptic function, axon guidance, and circadian entrainment. Together, these findings confirm that protein metabolic activity varies significantly between brain regions in vivo and indicate that pSILAC-based approaches can identify specific anatomical sites and biological pathways likely to be suitable for drug targeting in neurodegenerative disorders.\n\nAbbreviationsApoA1: Apolipoprotein A1, ApoA4: Apolipoprotein A4, ApoE: Apolipoprotein E, ApoJ/Clu: Apolipoprotein J/Clusterin, App: Amyloid-{beta} precursor/A4 protein: App, HDL: high density lipoprotein, Lrp1: Low density lipoprotein receptor-related protein 1, pSILAC: pulsed SILAC, pSIVOM: pulsed-SILAC in vivo labelling in mouse, SILAC: Stable Isotope Labelling by Amino acids in Cell culture)

neuroscience↗