Biology subjects

Vella, A. T.

Publications and source records attributed to Vella, A. T..

9 recordsLinked to original sources

bioRxiv2026

Endothelial PTBP1 Deletion in Transplanted Cardiac Tissue Limits Cardiac Allograft Vasculopathy

BackgroundCardiac allograft vasculopathy (CAV) is a leading cause of late graft failure and mortality following heart transplantation, with limited therapeutic options. Endothelial cells (ECs), at the interface between the donor graft and host immune system, play a central role in CAV development. However, the molecular mechanisms driving endothelial dysfunction and vascular remodeling in chronic heart transplant rejection remain poorly understood. MethodsTo characterize endothelial alterations associated with CAV, we isolated nuclei from cardiac tissues of four human donor groups: (1) early post-transplant CAV-negative surveillance biopsies, (2) CAV-negative explanted grafts with acute cellular rejection (ACR), (3) late-stage CAV-positive explanted grafts, and (4) naive non-transplanted control hearts. We applied intranuclear cellular indexing of transcriptomes and epitopes (inCITE-seq) to profile endothelial gene expression together with nuclear protein levels of splice factor polypyrimidine tract-binding protein 1 (PTBP1), a key post-transcriptional regulator of endothelial inflammatory responses. Functional relevance of PTBP1 was assessed using endothelial-specific deletion of Ptbp1 in an F1 hybrid murine model of CAV. ResultsIn human CAV, endothelial cells exhibited increased transforming growth factor-{beta} (TGF-{beta}) signaling and reduced oxidative phosphorylation (OxPhos) transcripts. Nuclear PTBP1 protein levels were markedly elevated in CAV endothelium and were associated with TGF-{beta}-responsive transcriptional programs and correlated with clinical indices of cardiac dysfunction. In murine heart transplants, endothelial-specific deletion of Ptbp1 markedly reduced hallmarks of CAV, including neointimal hyperplasia, fibrosis, and lymphocyte activation. At the molecular level, endothelial Ptbp1 deletion prevented suppression of mitochondrial transcripts and preserved mitochondrial content and integrity under hypoxic stress, attenuating interferon signaling in endothelial cells. ConclusionThese findings identify PTBP1 as a central endothelial regulator linking pro-fibrotic stress to mitochondrial dysfunction and immune activation in chronic cardiac allograft rejection. Targeting endothelial PTBP1 may represent a strategy to limit chronic graft injury while minimizing systemic immunosuppression.

immunology↗

bioRxiv2024

Unanchored ubiquitin chains promote the non-canonical inflammasome via UBXN1

Ubiquitination is a major posttranslational covalent modification that regulates numerous cellular processes including inflammasome signaling. Cells also contain unanchored ubiquitin chains (polyUb) that bind protein targets non-covalently, but their physiological functions in immunity have been appreciated only recently. Here, we report that ubiquitin regulatory x domain-containing protein 1 (UBXN1) activates the noncanonical inflammasome via unanchored Lysin (K) 48- or 63-linked polyUb. UBXN1 deficiency impairs the activation of caspase-4/11, secretion of inflammasome-dependent cytokines and pyroptosis in response to intracellular lipopolysaccharide (LPS). UBXN1-deficient mice are protected from LPS- and cecal-ligation-and-puncture-induced sepsis, evidenced by reduced mortality and systemic inflammation, compared to UBXN1-sufficient littermates. Mechanistically, UBXN1 together with unanchored K48/63-linked polyUb bind caspase-4/11, the intracellular sensors of LPS, and promote their assembly and activation. Depleting cellular unanchored polyUb with recombinant ubiquitin-specific proteinase 5 (USP5) reduces UBXN1 binding to caspase-4/11 and inflammasome signaling, while USP5 inhibitors enhance pyroptosis in an UBXN1-dependent manner. Thus, this study identifies a critical UBXN1-dependent posttranslational mechanism involved in noncanonical inflammasome activation and UBXN1 as a potential therapeutic target for sepsis and advances a fundamental understanding of unanchored polyUb biology.

immunology↗

bioRxiv2024

Endothelial Immunosuppression in Atherosclerosis : Translational Control by Elavl1/HuR

Atherosclerotic plaques are defined by the accumulation of lipids and immune cells beneath the endothelium of the arterial intima. CD8 T cells are among the most abundant immune cell types in plaque, and conditions linked to their activation correlate with increased levels of cardiovascular disease. As lethal effectors of the immune response, CD8 T cell activation is suppressed at multiple levels. These checkpoints are critical in dampening autoimmune responses, and limiting damage in cardiovascular disease. Endothelial cells are well known for their role in recruiting CD8 T and other hematopoietic cells to low and disturbed flow (LDF) arterial regions that develop plaque, but whether they locally influence CD8 effector functions is unclear. Here, we show that endothelial cells can actively suppress CD8 T cell responses in settings of chronic plaque inflammation, but that this behavior is governed by expression of the RNA-binding protein Embryonic Lethal, Abnormal Vision-Like 1 (Elavl1). In response to immune cell recruitment in plaque, the endothelium dynamically shifts splicing of pre-mRNA and their translation to enhance expression of immune-regulatory proteins including C1q and CD27. This program is immuno-suppressive, and limited by Elavl1. We show this by Cdh5(PAC)-CreERT2-mediated deletion of Elavl1 (ECKO), and analysis of changes in translation by Translating Ribosome Affinity Purification (TRAP). In ECKO mice, the translational shift in chronic inflammation is enhanced, leading to increased ribosomal association of C1q components and other critical regulators of immune response and resulting in a [~]70% reduction in plaque CD8 T cells. CITE-seq analysis of the remaining plaque T cells shows that they exhibit lower levels of markers associated with T cell receptor (TCR) signaling, survival, and activation. To understand whether the immunosuppressive mechanism occurred through failed CD8 recruitment or local modulation of T cell responses, we used a novel in vitro co-culture system to show that ECKO endothelial cells suppress CD8 T cell expansion--even in the presence of wild-type myeloid antigen-presenting cells, antigen-specific CD8 T cells, and antigen. Despite the induction of C1q mRNA by T cell co-culture in both wild-type and ECKO endothelial cells, we find C1q protein abundantly expressed only in co-culture with ECKO cells. Together, our data define a novel immune-suppressive transition in the endothelium, reminiscent of the transition of T cells to T-regs, and demonstrate the regulation of this process by Elavl1.

cell biology↗

bioRxiv2022

Therapeutic depletion of CD8+ T-cells prevents myelin pathology in Globoid Cell Leukodystrophy

Globoid cell leukodystrophy (GLD) or Krabbes disease is a fatal genetic demyelinating disease of the central nervous system caused by loss-of-function mutations in the galactosylceramidase (galc) gene. While the metabolic basis for disease is known, the understanding of how this results in neuropathology is not well understood. Herein we report that the rapid and protracted elevation of CD8+ cytotoxic T lymphocytes occurs coincident with clinical disease in a mouse model of GLD. Administration of a function blocking antibody against CD8 effectively prevented disease onset, reduced morbidity and mortality and prevented CNS demyelination in mice. These data indicate that subsequent to the genetic cause of disease, neuropathology is driven by pathogenic CD8+ T cells, thus offering novel therapeutic potential for treatment of GLD. One-Sentence SummaryCD8 T-cells mediate demyelination and neuroinflammation in a genetic white matter disease.

neuroscience↗

bioRxiv2021

Splice Factor Polypyrimidine tract-binding protein 1 (Ptbp1) is Required for Immune Priming of the Endothelium in Atherogenic Disturbed Flow Conditions

NF{kappa}B mediated endothelial activation drives leukocyte recruitment and atherosclerosis, in part through upregulation of adhesion molecules Icam1 and Vcam. The endothelium is "primed" for cytokine activation of NF{kappa}B by exposure to low and disturbed blood flow (LDF) in vivo and by LDF or static conditions in cultured cells. While priming leads to an exaggerated expression of Icam1 and Vcam following cytokine stimulation, the molecular underpinnings are not fully understood. We showed that alternative splicing of genes regulating NF{kappa}B signaling occurs during priming, but the functional implications of this are not known. We hypothesize that the regulation of splicing by RNA-binding splice factors is critical for priming. Here, we perform a CRISPR screen in cultured aortic endothelial cells to determine whether splice factors active in the response to LDF participate in endothelial cell priming. Using Icam1 and Vcam induction by TNF stimulation as a marker of priming, we identify polypyrimidine tract binding protein (Ptbp1) as a required splice factor. Ptbp1 expression is increased and its motifs are enriched nearby alternatively spliced exons in endothelial cells exposed to LDF in vivo in a platelet dependent manner, indicating its induction by early innate immune cell recruitment. At a mechanistic level, deletion of Ptbp1 inhibited NF{kappa}B nuclear translocation and transcriptional activation. These changes coincided with altered splicing of key components of the NF{kappa}B signaling pathway that were similarly altered in the LDF response. However, these splicing and transcriptional changes could be restored by expression of human PTBP1 cDNA in Ptbp1 deleted cells. In vivo, endothelial specific deletion of Ptbp1 reduced myeloid cell infiltration at regions of LDF in atherosclerotic mice. In human coronary arteries, PTBP1 expression correlates with expression of TNF pathway genes and amount of plaque. Together, our data suggest that Ptbp1, which is activated in the endothelium by innate immune cell recruitment in regions of LDF, is required for priming of the endothelium for subsequent NF{kappa}B activation and myeloid cell recruitment in vascular inflammation. Graphical Abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY Plaque forms in low and disturbed flow regions of the vasculature, where endothelial cells are "primed" to respond to cytokines (e.g. TNF) with elevated levels of cell adhesion molecules via the NF{kappa}B signaling pathway. We show that the splice factor Ptbp1 (purple) mediates priming. Ptbp1 is induced in endothelial cells by platelet recruitment, promoting priming and subsequent myeloid cell infiltration into plaque. Mechanistically, Ptbp1 regulates splicing of genes involved in the NF{kappa}B signaling pathway and is required for efficient nuclear translocation of NF{kappa}B in endothelial cells. This provides new insight into the molecular mechanisms underlying an endothelial priming process that reinforces vascular inflammatory responses. C_FIG_DISPLAY

cell biology↗

bioRxiv2021

UBXN3B Restricts Viral Pathogenesis by Maintaining Hematopoietic Homeostasis

Hematopoiesis is finely regulated to enable timely production of the right numbers and types of mature immune cells to maintain tissue homeostasis. Dysregulated hematopoiesis may compromise antiviral immunity and/or exacerbate immunopathogenesis. Herein, we report an essential role of UBXN3B in maintenance of hematopoietic homeostasis and restriction of immunopathogenesis during respiratory viral infection. Ubxn3b deficient (Ubxn3b-/-) mice are highly vulnerable to SARS-CoV-2 and influenza A infection, characterized by more severe lung immunopathology, lower virus-specific IgG, significantly fewer B cells, but more myeloid cells than Ubxn3b+/+ littermates. This aberrant immune compartmentalization is recapitulated in uninfected Ubxn3b-/- mice. Mechanistically, UBXN3B controls precursor B-I (pre-BI) transition to pre-BII and subsequent proliferation in a cell-intrinsic manner, by maintaining BLNK protein stability and pre-BCR signaling. These results reveal an essential role of UBXN3B for the early stage of B cell development.

immunology↗

bioRxiv2020

STING deficiency-associated aberrant CXCL10 expression contributes to pathogenesis of arthritogenic alphaviruses

Arthritogenic alphaviruses such as Chikungunya virus and Onyong nyong virus cause acute and chronic crippling arthralgia associated with inflammatory immune responses. However, the physiological functions of individual immune signaling pathways in the pathogenesis of alphaviral arthritis remain poorly understood. Here we report that a deficiency in the stimulator-of-interferon-genes (STING) led to enhanced viral loads, exacerbated inflammation and selectively elevated expression of CXCL10, a chemoattractant for monocytes/macrophages/T cells, in mouse feet. Cxcl10-/- mice had the same viremia as wild-type animals, but fewer immune infiltrates and lower viral loads in footpads at the peak of arthritic disease (6-8 days post infection). Macrophages constituted the largest immune cell population in footpads following infection, which were significantly reduced in Cxcl10-/- mice. The viral RNA loads in neutrophils and macrophages were reduced in Cxcl10-/- compared to wild-type mice. In summary, our results demonstrate that STING signaling represses, while CXCL10 signaling promotes, pathogenesis of alphaviral disease.

immunology↗

bioRxiv2019

ApoC-III overexpression and LDLr-/- protect mice from DSS-colitis: identifying a new role for lipoprotein metabolism in Tregs

ObjectiveCellular metabolism is a key regulator of CD4+Foxp3+ regulatory T cell (Treg) homeostasis, but the foundational studies in this area use free fatty acid treatment as a proxy for plasma triglycerides. In vivo, plasma triglyceride is the main source of fatty acids for cells, not free fatty acids. Design/ResultsUsing apolipoprotein C-III transgenic and LDLr-/- mice, we report that the loss of lipoprotein triglycerides transport in these models results in protection from DSS-colitis and accumulation of intestinal Tregs and plasma IL-10. Total loss of apoC-III increases colitis severity. Tregs exposed to apoC-III increase lipolysis and fatty acid oxidation and apoC-III inhibits Bodipy-triglyceride uptake. Therapeutic treatment of WT mice with apoC-III-containing lipoproteins protects mice from colitis. ConclusionOur data suggest that therapies that reduce apoC-III could have negative effects in patients who are at risk of IBD, and conversely, that apoC-III could be a new therapeutic target to stimulate intestinal Tregs and IL-10 for the management of IBD. These data identify apoC-III and lipoprotein metabolism as a novel regulator of tolerance in the intestine. Summary Box* What is already known about this subject:{blacksquare} The relative capacity to use either glucose or FFA to generate acetyl CoA for mitochondrial fatty acid oxidation is a critical driver of Treg and T cell activity and proliferation. {blacksquare}ApoC-III is a known regulator of triglyceride and fatty acid metabolism in cells via LPL and LDLr endocytosis pathways {blacksquare}ApoC-III is reduced in Crohns and Colitis patients. * What are the new findings:{blacksquare} We show that Tregs express triglyceride transporters, and that LDLr expression is enriched in Tregs from the mesenteric lymph nodes. {blacksquare}We show that T cells are capable of endocytosing triglyceride from lipoproteins, and this process is inhibited by apoC-III. {blacksquare}Tregs from apoC-IIITg are metabolically unique from WT Tregs and they upregulate the genes of lipolysis, and have an increase in basal respiration. {blacksquare}The inhibition of TAG endocytosis, using 2 different models (LDLrKO and apoC-III-transgenic mice), protects mice from colitis and stimulates the accumulation of Tregs and IL-10 in the gut. {blacksquare}Intraperitoneal delivery of apoC-III on chylomicrons protects WT mice from DSS colitis. * How might it impact on clinical practice in the foreseeable future?{blacksquare} Due to the protective role apoC-III plays in these mouse models of colitis, IBD risk should be carefully considered before prescribing patient anti-apoC-III lipid-lowering therapies.

physiology↗

bioRxiv2017

Addressing current challenges in cancer immunotherapy with mathematical and computational modeling

The goal of cancer immunotherapy is to boost a patients immune response to a tumor. Yet, the design of an effective immunotherapy is complicated by various factors, including a potentially immunosuppressive tumor microenvironment, immune-modulating effects of conventional treatments, and therapy-related toxicities. These complexities can be incorporated into mathematical and computational models of cancer immunotherapy that can then be used to aid in rational therapy design. In this review, we survey modeling approaches under the umbrella of the major challenges facing immunotherapy development, which encompass tumor classification, optimal treatment scheduling, and combination therapy design. Although overlapping, each challenge has presented unique opportunities for modelers to make contributions using analytical and numerical analysis of model outcomes, as well as optimization algorithms. We discuss several examples of models that have grown in complexity as more biological information has become available, showcasing how model development is a dynamic process interlinked with the rapid advances in tumor-immune biology. We conclude the review with recommendations for modelers both with respect to methodology and biological direction that might help keep modelers at the forefront of cancer immunotherapy development.

systems biology↗