Biology subjects

Pinal-Fernandez, I.

Publications and source records attributed to Pinal-Fernandez, I..

6 recordsLinked to original sources

bioRxiv2025

Myositis-specific autoantibodies recognizing Mi2 also target the autoimmune regulator (AIRE) protein at a shared PHD-zinc finger

ObjectivesIn dermatomyositis patients with anti-Mi2 autoantibodies, autoantibodies can enter muscle cells, leading to the aberrant expression of genes normally repressed by the Mi2/nucleosome remodeling and deacetylation (NuRD) complex. However, the mechanism by which autoantibodies interfere with Mi2/NuRD function remains unclear. This study aimed to identify additional autoantibodies in anti-Mi2-positive patients as well as the specific epitopes recognized by anti-Mi2 and any novel autoantibodies. MethodsPhage ImmunoPrecipitation Sequencing (PhIP-Seq) was used to screen serum samples from anti-Mi2-positive myositis patients for autoantibodies. Enzyme-linked immunosorbent assays (ELISA) and luciferase immunoprecipitation system (LIPS) immunoassays were used to detect autoantibodies in serum samples from myositis patients and healthy controls. ResultsPhIP-Seq identified autoantibodies recognizing the autoimmune regulator (AIRE) in sera from anti-Mi2 autoantibody-positive patients. Both anti-AIRE and anti-Mi2 autoantibodies predominantly recognized a homologous region of the plant homeodomain zinc finger type I (PHD1), which is critical for AIRE and Mi2/NuRD function. ELISA and LIPS testing showed that anti-Mi2 autoantibody-positive patients were positive for anti-AIRE autoantibodies, while AIRE reactivity was largely absent in healthy comparators, anti-Mi2 autoantibody-negative-myositis, and other autoimmune diseases. Affinity-purified anti-Mi2 autoantibodies recognized both Mi2 and AIRE by ELISA, whereas anti-Mi2-depleted immunoglobulin fractions did not recognize either protein. ConclusionsAutoantibodies recognizing Mi2 also recognize AIRE at a homologous PHD1 finger. This region is required by the Mi2/NuRD complex to anchor the nucleosome and consequently repress gene expression. Our findings suggest that anti-Mi2 autoantibodies disrupt NuRD complex function by binding to the PHD1 domain. Further studies are needed to determine if anti-Mi2 autoantibodies bind other PHD1-containing proteins and their functional implications.

immunology↗

bioRxiv2024

Mitochondrial leakage and mtDNA damage trigger early immune response in Inclusion Body Myositis.

Polymyositis with mitochondrial pathology (PM-Mito) was first identified in 1997 as a subtype of idiopathic inflammatory myopathy. Recent findings demonstrated significant molecular similarities between PM-Mito and Inclusion Body Myositis (IBM), suggesting a trajectory from early to late IBM and prompting the inclusion of PM-Mito as an IBM precursor (early IBM) within the IBM spectrum. Both PM-Mito and IBM show mitochondrial abnormalities, suggesting mitochondrial disturbance is a critical element of IBM pathogenesis. The primary objective of this cross-sectional study was to characterize the mitochondrial phenotype in PM-Mito at histological, ultrastructural, and molecular levels and to study the interplay between mitochondrial dysfunction and inflammation. Skeletal muscle biopsies of 27 patients with PM-Mito and 27 with typical IBM were included for morphological and ultrastructural analysis. Mitochondrial DNA (mtDNA) copy number and deletions were assessed by qPCR and long-range PCR, respectively. In addition, full-length single-molecule sequencing of the mtDNA enabled precise mapping of deletions. Protein and RNA levels were studied using unbiased proteomic profiling, immunoblotting, and bulk RNA sequencing. Cell-free mtDNA (cfmtDNA) was measured in the serum of IBM patients. We found widespread mitochondrial abnormalities in both PM-Mito and IBM, illustrated by elevated numbers of COX-negative and SDH-positive fibers and prominent ultrastructural abnormalities with disorganized and concentric cristae within enlarged and dysmorphic mitochondria. MtDNA copy numbers were significantly reduced, and multiple large-scale mtDNA deletions were already evident in PM-Mito, compared to healthy age-matched controls, similar to the IBM group. The activation of the canonical cGAS/STING inflammatory pathway, possibly triggered by the intracellular leakage of mitochondrial DNA, was evident in PM-Mito and IBM. Elevated levels of circulating cfmtDNA also indicated leakage of mtDNA as a likely inflammatory trigger. In PM-Mito and IBM, these findings were accompanied by dysregulation of proteins and transcripts linked to the mitochondrial membranes. In summary, we identified that mitochondrial dysfunction with multiple mtDNA deletions and depletion, disturbed mitochondrial ultrastructure, and defects of the inner mitochondrial membrane are features of PM-Mito and IBM, underlining the concept of an IBM-spectrum disease (IBM-SD). The activation of inflammatory pathways related to mtDNA release indicates a significant role of mitochondria-associated inflammation in the pathogenesis of IBM-SD. Thus, mitochondrial abnormalities precede tissue remodeling and infiltration by specific T-cell subpopulations (e.g., KLRG1+) characteristic of late IBM. This study highlights the critical role of early mitochondrial abnormalities in the pathomechanism of IBM, which may lead to new approaches to therapy.

neuroscience↗

bioRxiv2022

Transcriptomic profiling reveals distinct subsets of immune checkpoint inhibitor-induced myositis

ObjectivesInflammatory myopathy or myositis is a heterogeneous family of immune-mediated diseases including dermatomyositis (DM), antisynthetase syndrome (AS), immune-mediated necrotizing myopathy (IMNM), and inclusion body myositis (IBM). Immune checkpoint inhibitors (ICI) can also cause myositis (ICI-myositis). This study was designed to define gene expression patterns in muscle biopsies from patients with ICI-myositis. MethodsBulk RNA sequencing was performed on 200 muscle biopsies (35 ICI-myositis, 44 DM, 18 AS, 54 IMNM, 16 IBM, and 33 normal muscle biopsies) and single nuclei RNA sequencing was performed on 22 muscle biopsies (7 ICI-myositis, 4 DM, 3 AS, 6 IMNM, and 2 IBM). ResultsUnsupervised clustering defined three distinct transcriptomic subsets of ICI-myositis: ICI-DM, ICI-MYO1, and ICI-MYO2. ICI-DM included patients with DM and anti-TIF1{gamma} autoantibodies who, like DM patients, overexpressed type 1 interferon-inducible genes. ICI-MYO1 patients had highly inflammatory muscle biopsies and included all patients that developed co-existing myocarditis. ICI-MYO2 was composed of patients with predominant necrotizing pathology and low levels of muscle inflammation. The type 2 interferon pathway was activated both in ICI-DM and ICI-MYO1. Unlike the other types of myositis, all three subsets of ICI-myositis patients overexpressed genes involved in the IL6 pathway. ConclusionsWe identified three distinct types of ICI-myositis based on transcriptomic analyses. The IL6 pathway was overexpressed in all groups, the type I interferon pathway activation was specific for ICI-DM, the type 2 IFN pathway was overexpressed in both ICIDM and ICI-MYO1, and only ICI-MYO1 patients developed myocarditis.

immunology↗

bioRxiv2022

Contrasting autoimmune and treatment effects reveals baseline set points of immune toxicity following checkpoint inhibitor treatment

Immune checkpoint inhibitors (ICIs) have changed the cancer treatment landscape, but severe immune-related adverse events (irAEs) can be life-threatening or fatal and may prohibit patients from receiving further ICI treatment. While the clinical features of irAEs are well documented, molecular signatures, predictive biomarkers, and mechanisms of impending irAEs are largely unknown. In addition, the markers and mechanisms of ICI-induced antitumor immunity often overlap with those for irAEs. It is thus critical to uncover signatures associated specifically with irAEs but not with antitumor immunity. To identify circulating immune cell states associated with irAEs, we applied multimodal single cell analysis (CITE-seq) to simultaneously measure the transcriptome and surface proteins from peripheral blood mononuclear cells (PBMCs) collected before and after treatment with an anti-PD-L1 antibody (avelumab) in patients with thymic cancers (thymic epithelial tumors). All patients had an antitumor response, yet a subset developed muscle autoimmunity (myositis), a potentially life-threatening irAE. Mixed-effect modeling disentangled cell type-specific transcriptional states associated with ICI treatment responses from those of irAEs to identify temporally stable pre-treatment immune set points associated with irAEs only. These pre-treatment baseline signatures of irAE developed post-avelumab irAEs reflect correlated transcriptional states of multiple innate and adaptive immune cell populations, including elevation of metabolic genes downstream of mTOR signaling in T-cell subsets. Together these findings suggest putative pre-treatment biomarkers for irAEs following ICI therapy in thymic cancer patients and raise the prospect of therapeutically dampening autoimmunity while sparing antitumor activity in cancer patients treated with ICIs. Together, pre-treatment biomarkers and interventional therapeutics could help mitigate treatment discontinuation and improve clinical outcomes.

immunology↗

bioRxiv2021

Loss of TDP-43 function and rimmed vacuoles persist after T cell depletion in a xenograft model of sporadic inclusion body myositis

Sporadic inclusion body myositis (IBM) is the most common acquired muscle disease in adults over age 50, yet it remains unclear whether the disease is primarily driven by T cell-mediated autoimmunity. IBM muscle biopsies exhibit nuclear clearance and cytoplasmic aggregation of TDP-43 in muscle cells, a pathologic finding observed initially in neurodegenerative disease, and nuclear loss of TDP-43 in neurons causes aberrant RNA splicing. Here, we show that loss of TDP-43 splicing repression, as determined by inclusion of cryptic exons, occurs in skeletal muscle of IBM patients. Out of 119 muscle biopsies tested, RT-PCR-mediated detection of cryptic exon expression is 84% sensitive and 99% specific for diagnosing IBM, indicating utility as a functional and diagnostic biomarker. To determine the role of T cells in pathogenesis, we generated a novel xenograft model by transplanting human IBM muscle into the hindlimb of immunodeficient mice. Xenografts from IBM patients display robust regeneration of human myofibers and recapitulate both inflammatory and degenerative features of the disease. Myofibers in IBM xenografts are invaded by human, oligoclonal CD8+ T cells and exhibit MHC-I upregulation, rimmed vacuoles, mitochondrial pathology, p62-positive inclusions, and nuclear clearance and cytoplasmic aggregation of TDP-43, resulting in expression of cryptic exons. Depletion of human T cells within IBM xenografts by treating mice intraperitoneally with anti-CD3 (OKT3) suppresses MHC-I upregulation, but rimmed vacuoles and loss of TDP-43 function persist. These data suggest that myofiber degeneration occurs independent of T cells, and muscle cell-intrinsic mechanisms, such as loss of TDP-43 splicing repression, drive IBM pathogenesis. One Sentence SummaryDepletion of T cells in a xenograft model of sporadic inclusion body myositis suppresses inflammation but not TDP-43 pathology or muscle degeneration.

pathology↗

bioRxiv2018

Ultra-Efficient Short Read Sequencing of Immune Receptor Repertoires

AbstractT cell receptor (TCR) repertoire sequencing is increasingly employed to characterize adaptive immune responses. However, current TCR sequencing methodologies are complex and expensive, limiting the scale of feasible studies. Here we present Framework Region 3 AmplifiKation sequencing (FR3AK-seq), a simplified multiplex PCR-based approach for the ultra-efficient analysis of TCR complementarity determining region 3 (CDR3) repertoires. By using minimal primer sets targeting a conserved region adjacent to CDR3, undistorted amplicons are analyzed via short read, single-end sequencing. We find that FR3AK-seq is sensitive and quantitative, performing comparably to two industry standards. FR3AK-seq was utilized to quickly and inexpensively characterize the T cell infiltrates of muscle biopsies obtained from 145 patients with idiopathic inflammatory myopathies and controls. A cluster of related TCRs was identified in samples from patients with sporadic inclusion body myositis, suggesting the presence of a shared antigen-driven response. The ease and minimal cost of FR3AK-seq removes critical barriers to routine, large-scale TCR CDR3 repertoire analyses.

immunology↗