Biology subjects

Pala, F.

Publications and source records attributed to Pala, F..

5 recordsLinked to original sources

bioRxiv2024

Lymphopenia drives T cell exhaustion in immunodeficient STING gain-of-function mice

STING gain-of-function (GOF) mutations are associated with the severe autoinflammatory disease designated STING Associated Vasculopathy with onset in Infancy (SAVI). Mice with the STING GOF V154M mutation develop profound T cell lymphopenia, partly due to a blockage of T cell development in the thymus. To better characterize the mechanisms of peripheral T cell dysfunctions, we conducted a transcriptomic and phenotypic analysis on sorted splenic CD4+ and CD8+ mature T cells from STING GOF V154M mice. We identify a T cell exhaustion phenotype that manifests at a terminal stage, acquired early in life but only after reaching the peripheral environment. This phenotype is independent of type I interferons and does not rely on intrinsic STING activation in either T cells or stromal cells. Mechanistically, the limited number of mature T cells that reach the periphery appear to be quickly impacted by the lymphopenic environment, experiencing heightened stimulation of the IL-7 receptor and TCR pathways, including the NFAT pathway, a key factor in T cell exhaustion. By performing transplantation experiments with STING GOF long term-hematopoietic stem cells (LT-HSCs) along with supportive wild-type bone marrow (BM) cells, we were able to prevent the T cell exhaustion of STING GOF T cells in the resulting non-lymphopenic context, demonstrating that lymphopenia is a major driver of T cell exhaustion in STING GOF mice. T cell exhaustion, although less severe, was also observed in lymphopenic mice carrying Rag1 hypomorphic mutations. In conclusion, our results, which highlight T cell exhaustion induced by lymphopenia, could have important implications for the management of patients with severe immune deficiencies. HighlightsO_LIWe describe a phenotype of T cell exhaustion in STING GOF V154M mice, which is acquired early in life and in the periphery. C_LIO_LISTING GOF-associated T cell exhaustion is independent of type I IFNs, and STING GOF/activation in T cells or in stromal cells is not sufficient for T cell exhaustion. C_LIO_LILymphopenia is a major driver of T cell exhaustion in STING GOF mice, and increased antigenic/IL-7 stimulation of T cells in the lymphopenic context of STING GOF mice could be implicated in the induction of T cell exhaustion. C_LI

immunology↗

bioRxiv2023

A spatial human thymus cell atlas mapped to a continuous tissue axis

T cells develop from circulating precursors, which enter the thymus and migrate throughout specialised sub-compartments to support maturation and selection. This process starts already in early fetal development and is highly active until the involution of the thymus in adolescence. To map the micro-anatomical underpinnings of this process in pre- vs. post-natal states, we undertook a spatially resolved analysis and established a new quantitative morphological framework for the thymus, the Cortico-Medullary Axis. Using this axis in conjunction with the curation of a multimodal single-cell, spatial transcriptomics and high-resolution multiplex imaging atlas, we show that canonical thymocyte trajectories and thymic epithelial cells are highly organised and fully established by post-conception week 12, pinpoint TEC progenitor states, find that TEC subsets and peripheral tissue genes are associated with Hassalls Corpuscles and uncover divergence in the pace and drivers of medullary entry between CD4 vs. CD8 T cell lineages. These findings are complemented with a holistic toolkit for spatial analysis and annotation, providing a basis for a detailed understanding of T lymphocyte development.

immunology↗

bioRxiv2022

Genetically Corrected RAG2-SCID Human Hematopoietic Stem CellsRestore V(D)J-Recombinase and Rescue Lymphoid Deficiency

Recombination-activating genes (RAG1 and RAG2) are critical in lymphoid cell development and function for initiating the V(D)J-recombination process to generate polyclonal lymphocytes with broad antigen-specificity. Clinical manifestations of defective RAG1/2 genes range from immune dysregulation to severe combined immunodeficiencies (SCID), causing life-threatening infections and death early in life in the absence of hematopoietic cell transplantation (HCT). Haploidentical HCT without myeloablative conditioning carries a high risk of graft failure and incomplete immune reconstitution. The RAG complex is only expressed during the G0-G1 phases of the cell cycle at the early stages of T and B cell development, underscoring that a direct gene correction would capture the precise temporal expression of the endogenous gene, is a promising therapeutic approach for RAG1/2-deficiencies. Here, we report a feasibility study using the CRISPR/Cas9-based "universal gene-correction" approach for the RAG2 locus in human hematopoietic stem/progenitor cells (HSPCs) in healthy donors and one RAG2-SCID patient. V(D)J recombinase activity was restored following gene correction of RAG2-SCID-derived HSPCs, resulting in the development of TCR {beta} and {gamma}{delta} CD3+ cells and single-positive CD4+ and CD8+ lymphocytes. TCR repertoire analysis indicated a normal distribution of the CDR3 length and preserved usage of distal TRAV genes. We confirmed in vivo rescue of B-cell development, with normal IgM surface expression and a significant decrease in CD56bright NK cells. Together, we provide specificity, toxicity, and efficacy data supporting the development of a gene-correction therapy to benefit all RAG2-deficient patients. KEY POINTSO_LIHuman hematopoietic stem cells can be corrected to restore endogenous RAG2 gene expression while preserving durable engraftment potential. C_LIO_LIGene-corrected RAG2 locus restores V(D)J recombination in RAG2-SCID patient stem cells, promoting T and B-cells receptor formation. C_LI

immunology↗

bioRxiv2019

A distinct cardiopharyngeal mesoderm genetic hierarchy establishes antero-posterior patterning of esophagus striated muscle

In most vertebrates, the upper digestive tract is composed of muscularised jaws linked to the esophagus that permit food uptake and swallowing. Masticatory and esophagus striated muscles (ESM) share a common cardiopharyngeal mesoderm (CPM) origin, however ESM are unusual among striated muscles as they are established in the absence of a primary skeletal muscle scaffold. Using mouse chimeras, we show that the transcription factors Tbx1 and Isl1 are required cell-autonomously for myogenic specification of ESM progenitors. Further, genetic loss-of-function and pharmacological studies point to Met/HGF signalling for antero-posterior migration of esophagus muscle progenitors, where HGF ligand is expressed in adjacent smooth muscle cells. These observations highlight the functional relevance of a smooth and striated muscle progenitor dialogue for ESM patterning. Our findings establish a Tbx1-Isl1-Met genetic hierarchy that uniquely regulate esophagus myogenesis and identify distinct genetic signatures that can be used as a framework to interpret pathologies arising within CPM derivatives.

developmental biology↗

bioRxiv2017

Comparison of multiple transcriptomes exposes unified and divergent features of quiescent and activated skeletal muscle stem cells

BackgroundSkeletal muscle stem cells (MuSCs) are quiescent in adult mice and can undergo multiple rounds of proliferation and self-renewal following muscle injury. Several labs have profiled transcripts of myogenic cells during developmental and adult myogenesis with the aim of identifying quiescent markers. Here, we focused on the quiescent cell state and generated new transcriptome profiles that include subfractionations of adult MuSC populations, and an artificially induced prenatal quiescent state, to identify core signatures for quiescent and proliferating MuSCs.\n\nMethodsComparison of available data offered challenges related to the inherent diversity of datasets and biological conditions. We developed a standardized workflow to homogenize the normalization, filtering, quality control steps for the analysis of gene expression profiles allowing the identification up- and down-regulated genes and the subsequent gene set enrichment analysis. To share the analytical pipeline of this work, we developed Sherpa, an interactive Shiny server that allows multiscale comparisons for extraction of desired gene sets from the analysed datasets. This tool is adaptable to cell populations in other contexts and tissues.\n\nResultsA multiscale analysis comprising eight datasets of quiescent MuSCs had 207 and 542 genes commonly up- and down-regulated, respectively. Shared up-regulated gene sets include an over-representation of the TNFa pathway via NFKb signaling, Il6-Jak-Stat3 signaling, and the apical surface processes, while shared down-regulated gene sets exhibited an over-representation of Myc and E2F targets, and genes associated to the G2M checkpoint and oxidative phosphorylation. However, virtually all datasets contained genes that are associated with activation or cell cycle entry, such as the immediate early stress response genes Fos and Jun. Empirical examination of fixed and isolated MuSCs showed that these and other genes were absent in vivo, but activated during procedural isolation of cells.\n\nConclusionsThrough the systematic comparison and individual analysis of diverse transcriptomic profiles, we identified genes that were consistently differentially expressed among the different datasets and common underlying biological processes key to the quiescent cell state. Our findings provide impetus to define and distinguish transcripts associated with true in vivo quiescence from those that are first responding genes due to disruption of the stem cell niche.

cell biology↗