Biology subjects

Roux, J.

Publications and source records attributed to Roux, J..

At least 19 recordsLinked to original sources

bioRxiv2026

Single-cell transcriptomics reveals a differential response of human bronchial epithelial cell-types to cadmium chloride

Exposure of cells or tissues to chemical compounds can be analyzed through transcriptomic signatures, which can be used to classify chemical agents. This information can also enrich Adverse Outcome Pathways (AOP). Transcriptional signatures have generally been obtained using "bulk" analysis, by which the global gene expression pattern of an entire tissue is determined. Although this approach has been useful in toxicology, some information is lost, especially when tissues containing multiple cell types are considered. With the advent of single-cell transcriptomics (scRNA-seq), it is now possible to obtain higher resolution, cell type-specific responses in complex tissues. The aim of the present study was to evaluate the added value of scRNA-seq in analysis of the acute response of human bronchial epithelial cells grown at the air/liquid interface (ALI) to a known toxic compound, CdCl2, with well described transcriptional signatures of exposure. Fully differentiated mucocilliary epithelia obtained from three independent donors were exposed to 10 {micro}M CdCl2 and scRNA-seq analysis was performed on a total of 18255 cells to obtain cell type-specific signatures. Our results show that the contribution of each cell type to the overall transcriptomic bulk response varies. For example, the classical heavy metal detoxification response was only detected in multiciliated and secreting cells, while absent in basal cells. The data demonstrate that scRNA-seq provides high-resolution transcriptional signatures with unexpected features. This added information is likely to have implications for the refinement of AOPs and could serve as a basis for a new generation of tests in predictive toxicology.

pharmacology and toxicology↗

bioRxiv2025

Chronic Infection Perturbs the Affinity Hierarchy of Antiviral B Cells

The germinal center (GC) subjects antigen-specific B cells to a Darwinian selection process. Whether and how persistent viral infection perturbs the intended affinity hierarchy remains ill-defined. Here we transferred monoclonal lymphocytic choriomeningitis virus-specific B cells into persistently infected hosts. High affinity B cells expanded vigorously, forming GCs and abundant antibody-secreting cells. When failing to gain the upper hand over the virus, the expanded B cell population contracted, ending in its quasi-complete disappearance, a process termed "attrition". In stark contrast, low-affinity B cells expanded and persisted irrespective of high viral loads. B cell attrition was associated with phenotypic and transcriptional alterations including a prominent Blimp-1 transcriptional signature in high-affinity GC B cells. Blimp-1-deficient B cells were resistant to attrition, suggesting a B cell-intrinsic process. Moreover, exogenously supplied antibody feedback prevented attrition, indicating the latter resulted from excessive stimulation. Our findings reveal that in chronic viral infection the incessant activation by overwhelming amounts of antigen perturbs B cell affinity hierarchies by preferentially dysregulating high-affinity B cells.

immunology↗

bioRxiv2024

Cultured alveolar basal cells display similar characteristics and functional properties to airway basal cells

RationaleBasal cells (BC) appear ectopically within the lung parenchyma of interstitial lung disease (ILD) patients, potentially through migration of airway BC or though trans-differentiation of alveolar epithelial type 2 (AT2) cells. The exact origin and function of these ectopic alveolar BC remains elusive. By comparing ectopic alveolar to "classical" airway BC, we aimed to get a better understanding of the origin and characteristics of alveolar BC in ILD. MethodsAlveolar and airway BC were isolated from transbronchial and airway mucosal biopsies, respectively, from the same ILD patients and expanded in culture. Samples were analyzed by single cell RNA sequencing (scRNA-seq), TaqMan RT-PCR, and immunochemistry. ResultsscRNA-seq analysis revealed several differences in gene expression that suggested a shift to a more mesenchymal-like phenotype and a decrease in keratinization genes in alveolar compared to airway BC. Specific AT2 cell marker genes were not expressed in either BC type. While the morphology, wound repair and proliferation capacities of BC from both origins were not significantly different, alveolar BC formed significantly fewer organoids, expressing more MUC5B. After instillation into bleomycin-injured mice, alveolar and airway BC showed similar engraftment, differentiation capacity and effects on fibrosis. ConclusionDespite similar overall functionality in vitro and after instillation into bleomycin-injured mice, alveolar and airway BC differed in their transcriptomes and in their capacities to form and to differentiate in organoids. Our data provide no evidence to support their potential derivation from AT2 cells. Take home messageAlveolar and airway basal cells differ in their transcriptomes and in their capacities to form and to differentiate in organoids, although with no indication of an AT2 cell origin.

cell biology↗

bioRxiv2024

Distinct circulating monocytes up-regulate CD52 and sustain innate immune function in patients with cirrhosis unless acute decompensation emerges

Background & AimsInfectious complications determine the prognosis of cirrhosis patients. Their infection susceptibility relates to the development of immuneparesis, a complex interplay of different immunosuppressive cells and soluble factors. Mechanisms underlying the dynamics of immuneparesis of innate immunity remain inconclusive. We aimed to dissect the heterogeneity of circulating monocyte states in different cirrhosis stages, and pursued the function of selected differentially expressed (DE) genes. MethodsWe systematically investigated circulating monocytes in health, compensated and not-acutely decompensated (NAD) cirrhosis using single cell RNA sequencing. Selective genes were confirmed by flow cytometry and diverse functional assays on monocytes ex vivo. ResultsWe identified seven monocyte clusters. Their abundances varied between cirrhosis stages, confirming previously reported changes i.e. reduction in CD14lowCD16++ and emergence of M-MDSC in advanced stages. DE genes between health and disease and among stages were detected, including for the first time CD52. CD52-expression on monocytes significantly enhanced throughout compensated and NAD cirrhosis. Heretofore the biological significance of CD52-expression on monocytes remained unknown. CD52highCD14+CD16highHLA-DRhigh monocytes in patients with cirrhosis revealed a functional phenotype of active phagocytes with enhanced migratory potential, increased cytokine production, but poor T cell activation. Following acute decompensation (AD), CD52 was cleaved by elevated phospholipase C (PLC), and soluble CD52 (sCD52) was detected in the circulation. Inhibition and cleavage of CD52 significantly suppressed monocyte functions ex vivo and in vitro, and the predominance of immunosuppressive CD52low circulating monocytes in patients with AD was associated with infection and low transplant-free survival. ConclusionCD52 may represent a biologically relevant target for future immunotherapy. Stabilising CD52 may enhance monocyte functions and infection control in the context of cirrhosis, guided by sCD52/PLC as biomarkers indicating immuneparesis. Lay summaryRecurrent infections are a major cause of death in patients with liver cirrhosis. A fundamental understanding of the mechanisms that suppress immune responses in patients with cirrhosis is lacking, but required for the development of strategies to restore innate immunity in cirrhosis patients and prevent infection. The current study identified a novel marker for deficient immune responses and a potential target for such a future immune-based therapy. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=134 SRC="FIGDIR/small/587894v1_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@1a42b1borg.highwire.dtl.DTLVardef@10a0df2org.highwire.dtl.DTLVardef@1199087org.highwire.dtl.DTLVardef@15fb074_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical AbstractC_FLOATNO scRNA-seq identified seven circulating monocyte states, changing in cirrhosis patients at different stages of disease. Circulating monocytes overexpress CD52 in cirrhosis, but are absent in AD/ACLF due to PLC. CD52-expressing monocytes show high capability for phagocytosis, cytokine production, adhesion and migration potential and T cell suppression. Created with BioRender.com C_FIG

immunology↗

bioRxiv2024

Targeting the expansion of myeloid-derived suppressor cells in liver cirrhosis

Background and aimsPreviously, we identified immune-suppressive circulating monocytic myeloid-derived suppressor cells (M-MDSC) in patients with cirrhosis and liver failure, which increased with disease severity and were associated with infections and mortality. Impaired immune responses and M-MDSC expansion were reversed by ex vivo polyinosinic:polycytidylic acid (poly(I:C)) treatment. Here, we aimed to investigate hepatic MDSC subsets in liver biopsies of cirrhotic patients and identify MDSC subsets in murine models to assess the safety and efficacy of poly(I:C) in vivo. Methods22 cirrhotic patients and 4 controls were clinically characterised. MDSC were identified in liver biopsies (immunofluorescence) and in the circulation (flow cytometry). M- MDSC phenotype and function following poly(I:C) stimulation were assessed ex vivo. Carbon tetrachloride-based murine models of liver fibrosis were used. Poly(I:C) was administered therapeutically. MDSC biology was investigated with flow cytometry, immunofluorescence and T-cell proliferation assay. Hepatic histopathology, transcriptomics (BulkRNAseq) and serum markers were assessed. ResultsBesides circulating M-MDSC, hepatic CD14+CD84+M-MDSC and CD15+CD84+ polymorphonuclear-MDSC expanded in cirrhotic patients and indicated disease severity, infections and poor survival. Poly(I:C) treatment reversed phenotype and function of circulating M-MDSC ex vivo. Circulating and hepatic MDSC expanded in our murine models of liver fibrosis and suppressed T-cell proliferation. Lipopolysaccharide and E.coli challenge exacerbated hepatic MDSC and fibrosis compared to CCl4 controls. Poly(I:C) therapy reduced MDSC expansion in fibrotic mice with bacterial infection and CCl4-induced fibrosis. ConclusionHepatic MDSC expanded in cirrhotic patients and were linked with disease severity and poor prognosis. Poly(I:C) reversed frequency and function of M-MDSC ex vivo. Poly(I:C) therapy reversed MDSC expansion and fibrosis in a murine model of liver fibrosis with infection. Thus, we highlighted poly(I:C) as a potential immunotherapy for the treatment of immuneparesis in cirrhosis.

immunology↗

bioRxiv2023

Multidimensional analysis of matched primary and recurrent glioblastoma identifies Fcgamma receptors upregulation on microglia as a contributor of tumor recurrence.

O_LIBackground: Glioblastoma (GBM) is a lethal brain tumor without effective treatment options. The aim of this study was to characterize longitudinal tumor immune microenvironment (iTME) changes in order to find potential actionable targets to prevent GBM-induced immune evasion mechanisms. C_LIO_LIMethods: This study included 15 patient-matched treatment-naive WHO grade 4 primary (pGBM) and recurrent (rGBM) tumors. RNA and proteins extracted from fresh frozen tumor samples from matched pGBM and rGBM were profiled via transcriptomics and proteomics, respectively. A tissue microarray containing paired formalin-fixed paraffin-embedded tumor samples was processed for spatial transcriptomics analysis. C_LIO_LIResults: Differentially expressed genes and proteins between pGBM and rGBM were involved in pathways responsible for synapse development and myelination which have been shown to play a role in GBM recurrence. By categorizing patients into short and long time-to-relapse (STTR vs LTTR), we identified genes positively or negatively associated with TTR. Expression of Fc{gamma} receptors and complement system genes such as FCGR1A (CD64), FCGR3A and C3 in rGBM samples were negatively correlated with TTR, whereas expression of DNMT1/3A, and SMARCA4, involved in DNA methylation, were positively correlated with TTR. Spatial transcriptomic analysis of the tumor cell compartment showed enrichment of oligodendrocytes in rGBM, whereas the myeloid cell compartment switched from quiescent to activated microglia, was enriched in B and T cells, specifically in rGBM with STTR. C_LIO_LIConclusions: Our results uncover a role for CD64-expressing activated microglia in GBM recurrence and suggest that interfering with these cells may represent a therapeutic option for hindering GBM relapse. C_LI Key pointsO_LITranscriptomic and proteomic differences exist between patient-paired primary and recurrent GBM tumors C_LIO_LIHigh expression of Fcy receptors genes on activated microglia at tumor recurrence is associated with shorter time to relapse. C_LI Importance of this studyIn glioblastoma (GBM), the tumor recurs in almost all cases after standard treatment such as surgery and chemo-radiotherapy. In this study, we longitudinally evaluated the immune- and neoplastic compartments using transcriptomic, proteomic, and spatial transcriptomics in patient-matched treatment-naive and recurrent tumor samples. By correlating gene expression with time-to-relapse, we identified a geneset associated with treatment resistance and faster tumor recurrence. Moreover, this study highlighted the plasticity of the myeloid compartment during disease progression and an unfavorable role of activated microglia in tumor recurrence.

cancer biology↗

bioRxiv2022

The Siglec-sialic acid-axis is a target for innate immunotherapy of glioblastoma

Glioblastoma (GBM) is the most aggressive form of primary brain tumor, for which effective therapies are urgently needed. Cancer cells are capable of evading clearance by phagocytes such as microglia and monocyte-derived cells through engaging tolerogenic programs. Here, we found that high level of Siglec-9 expression correlates with reduced survival in GBM patients. Using conditional knockouts of Siglec-E, the murine functional homologue of Siglec-9, together with single-cell RNA sequencing, we demonstrated significant pro-phagocytosis effects in microglia and monocyte-derived cells in the absence of Siglec-E. Loss of Siglec-E on monocyte-derived cells enhances antigen cross-presentation and production of pro-inflammatory cytokines, resulting in more efficient T cell priming. This bridging of innate and adaptive responses delays tumor growth and results in prolonged survival. Further, we showed synergistic activity of Siglec-E blockade in combinatorial immunotherapies and demonstrate its translational potential against GBM.

cancer biology↗

bioRxiv2022

Single-cell characterization of human GBM reveals regional differences in tumor-infiltrating leukocyte activation

Glioblastoma (GBM) harbors a highly immunosuppressive tumor microenvironment (TME) which influences glioma growth. Major efforts have been undertaken to describe the TME on a single-cell level. However, human data on regional differences within the TME remain scarce. Here, we performed high-depth single-cell RNA sequencing (scRNAseq) on paired biopsies from the tumor center, peripheral infiltration zone and blood of five primary GBM patients. Through analysis of > 45000 cells, we revealed a regionally distinct transcription profile of microglia (MG) and monocyte-derived macrophages (MdMs) and an impaired activation signature in the tumor-peripheral cytotoxic-cell compartment. Comparing tumor-infiltrating CD8+ T cells with circulating cells identified CX3CR1high and CX3CR1int CD8+ T cells with effector and memory phenotype, respectively, enriched in blood but absent in the TME. Tumor CD8+ T cells displayed a tissue-resident memory phenotype with dysfunctional features. Our analysis provides a regionally resolved mapping of transcriptional states in GBM-associated leukocytes, serving as an additional asset in the effort towards novel therapeutic strategies to combat this fatal disease.

cancer biology↗

bioRxiv2022

Robust inference of expression state in bulk and single-cell RNA-Seq using curated intergenic regions

RNA-Seq is a powerful technique to provide quantitative information on gene expression. While many applications focus on measuring expression levels, accurately distinguishing between actively and inactively transcribed genes is equally important for understanding gene function, development, and disease mechanisms. However, setting a biologically meaningful threshold for calling genes expressed is challenging due to variability in noise levels across different protocols, experiments or biological samples. We propose to define this threshold per sample relative to the background level observed in inactive genomic features, inferred by the amount of reads mapped to intergenic regions of the genome, and to call genes expressed if their level of expression is significantly higher than the estimated background noise. This approach can be applied to a single RNA-Seq library as well as to a combination of libraries from the same condition, in model and non-model organisms. We show that our method yields a more accurate prediction of expression state than existing methods, illustrated by consistent expression calls for biological replicates in the same tissue.

genomics↗

bioRxiv2021

Processing body dynamics drive non-genetic MEK inhibitors tolerance by fine-tuning KRAS and NRAS translation

Overactivation of the Mitogen-activated protein kinase (MAPK) pathway is a critical driver of many human cancers. However, therapies targeting this pathway have proven effective in only a few cancers, as cancers inevitably develop resistance. Puzzling observations suggest that MAPK targeting fails in tumors due to early compensatory RAS overexpression, albeit by unexplained mechanisms. We identified a novel mechanism of drug tolerance to MEK inhibitors (MEKi) that involves Processing Bodies (PBs), a membraneless organelle (MLO). MEKi promoted translation of the oncogenes KRAS and NRAS, which in turn triggered BRAF phosphorylation. This overexpression, which occurred in the absence of neotranscription, depended on PB dissolution as the source of the RAS mRNA. Moreover, in response to MEKi removal, the process was dynamic as PBs rapidly reformed and reduced MAPK signaling. These results highlight a dynamic spatiotemporal negative feedback loop of MAPK signaling via RAS mRNA sequestration. Furthermore, we observed a phenotype with a low number of PBs along with strong KRAS and NRAS induction capacities. Overall, we describe a new intricate mechanism involving PBs in the translational regulation of essential cellular signaling pathways like MAPKs, paving the way for future therapies altering MLO and thereby improving targeted cancer therapies.

cancer biology↗

bioRxiv2021

The Endocytic Recycling Compartment Serves as a Viral Factory for Hepatitis E Virus

Background & AimsAlthough Hepatitis E virus (HEV) is the major leading cause of enterically transmitted viral hepatitis worldwide, many gaps remain in the understanding of the HEV lifecycle. Notably, viral factories induced by HEV have not been documented yet and it is currently unknown whether HEV infection leads to cellular membrane modelling as many positive-strand RNA viruses. HEV genome encodes three proteins, the ORF1 replicase, the ORF2 capsid protein and the ORF3 protein involved in virion egress. Previously, we demonstrated that HEV produces different ORF2 isoforms including the virion-associated ORF2i form. Here, we aimed to probe infectious particles and viral factories in HEV-producing cells, using antibodies directed against the different ORF2 isoforms. MethodsWe generated monoclonal antibodies that specifically recognize the particle-associated ORF2i form, and antibodies that recognize the different ORF2 isoforms. We used them in confocal and electron microscopy approaches to probe viral factories in HEV-producing cells. We performed an extensive colocalization study of viral proteins with subcellular markers. We analyzed the impact of silencing Rab11, a central player of the endocytic recycling compartment (ERC). ResultsOne of the antibodies, named P1H1 and targeting the N-terminus of ORF2i, recognized delipidated HEV particles. Confocal and ultrastructural microscopy analyses of HEV-producing cells revealed an unprecedented HEV-induced membrane network containing tubular and vesicular structures. These subcellular structures were enriched in ORF2 and ORF3 proteins, and were dependent on the ORF3 expression and ORF2i capsid protein assembly. Colocalization and silencing analyses revealed that these structures are derived from the ERC. ConclusionsOur study reveals that HEV hijacks the ERC and forms a membrane network of vesicular and tubular structures that might be the hallmark of HEV infection. Lay summaryHepatitis E virus (HEV) is the leading cause of acute hepatitis worldwide but many steps of its lifecycle are still elusive. Thanks to the development of new antibodies that recognize the different forms of the HEV capsid protein, we were able to visualize vesicular and tubular structures that were established by the virus in the host cell. In addition, extensive efforts to identify these structures led us to conclude that HEV hijacks the endocytic recycling compartment of the cell to form this network of vesicles and tubules, which might be the hallmark of HEV infection.

microbiology↗

bioRxiv2021

T cell stimulation remodels the latently HIV-1 infected cell population by differential activation of proviral chromatin

The reservoir of latently HIV-1 infected cells is heterogeneous. To achieve an HIV-1 cure, the reservoir of activatable proviruses should be eliminated while permanently silenced proviruses may be tolerated. We have developed a method to assess the proviral nuclear microenvironment in single cells. In latently HIV-1 infected cells, a zinc finger protein tethered to the HIV-1 promoter produced a fluorescent signal as a protein of interest came in its proximity, such as the viral transactivator Tat when recruited to the nascent RNA. Tat is essential for viral replication. In these cells we assessed the proviral activation and chromatin composition. By linking Tat recruitment to proviral activity, we dissected the mechanisms of HIV-1 latency reversal and the consequences of HIV-1 production. A pulse of promoter-associated Tat was identified that contrasted to the continuous production of viral proteins. As expected, promoter H3K4me3 led to substantial expression of the provirus following T cell stimulation. However, the activation-induced cell cycle arrest and death led to a surviving cell fraction with proviruses encapsulated in repressive chromatin. Further, this cellular model was used to reveal mechanisms of action of small molecules. In a proof-of-concept study we determined the effect of an enhancer specific CBP/P300-inhibitor on HIV-1 latency reversal. Only proviruses resembling active enhancers, associated with H3K4me1 and H3K27ac, efficiently recruited Tat. Tat-independent HIV-1 latency reversal of unknown significance still occurred. We present a method for single cell assessment of the microenvironment of the latent HIV-1 proviruses, used here to reveal how T cell stimulation modulates the proviral activity and how the subsequent fate of the infected cell depends on the chromatin context.

microbiology↗

bioRxiv2021

Epithelial GPR35 protects from Citrobacter rodentium infection by preserving goblet cells and mucosal barrier integrity

Goblet cells secrete mucin to create a protective mucus layer against invasive bacterial infection and are therefore essential for maintaining intestinal health. However, the molecular pathways that regulate goblet cell function remain largely unknown. Although GPR35 is highly expressed in colonic epithelial cells, its importance in promoting the epithelial barrier is unclear. In this study, we show that epithelial Gpr35 plays a critical role in goblet cell function. In mice, cell type-specific deletion of Gpr35 in epithelial cells but not in macrophages results in goblet cell depletion and dysbiosis, rendering these animals more susceptible to Citrobacter rodentium infection. Mechanistically, scRNA-seq analysis indicates that signaling of epithelial Gpr35 is essential to maintain normal pyroptosis levels in goblet cells. Our work shows that the epithelial presence of Gpr35 is a critical element for the function of goblet cell-mediated symbiosis between host and microbiota.

immunology↗

bioRxiv2021

Gene prioritization based on random walks with restarts and absorbing states, to define gene sets regulating drug pharmacodynamics from single-cell analyses

Prioritizing genes for their role in drug sensitivity, is an important step in understanding drugs mechanisms of action and discovering new molecular targets for co-treatment. To formalize this problem, we consider two sets of genes X and P respectively composing the predictive gene signature of sensitivity to a drug and the genes involved in its mechanism of action, as well as a protein interaction network (PPIN) containing the products of X and P as nodes. We introduce Genetrank, a method to prioritize the genes in X for their likelihood to regulate the genes in P. Genetrank uses asymmetric random walks with restarts, absorbing states, and a suitable renormalization scheme. Using novel so-called saturation indices, we show that the conjunction of absorbing states and renormalization yields an exploration of the PPIN which is much more progressive than that afforded by random walks with restarts only. Using MINT as underlying network, we apply Genetrank to a predictive gene signature of cancer cells sensitivity to tumor-necrosis-factor-related apoptosis-inducing ligand (TRAIL), performed in single-cells. Our ranking provides biological insights on drug sensitivity and a gene set considerably enriched in genes regulating TRAIL pharmacodynamics when compared to the most significant differentially expressed genes obtained from a statistical analysis framework alone. We also introduce gene expression radars, a visualization tool to assess all pairwise interactions at a glance. Genetrank is made available in the Structural Bioinformatics Library (https://sbl.inria.fr/doc/Genetrank-user-manual.html). It should prove useful for mining gene sets in conjunction with a signaling pathway, whenever other approaches yield relatively large sets of genes.

systems biology↗

bioRxiv2020

The non-coding RNA miR-17~92 is a central mediator of T cell activation

T cell activation is paramount for productive adaptive immune responses. CD28 is a key clinically targeted immunoregulatory receptor because it provides the prototypical costimulatory signal required for T cell activation. Therefore, a precise understanding of the molecular consequences of CD28 costimulation has direct therapeutic relevance. Here, we uncover that the microRNA cluster miR-17~92 is part of the molecular program triggered by CD28 costimulation and hence T cell activation. Combining genetics, transcriptomics, bioinformatics and biochemical miRNA:mRNA interaction maps we demonstrate that transgenic miR-17~92 can cell-intrinsically largely overcome defects caused by CD28-deficiency. miR-17~92 promotes transcription of a proinflammatory gene signature by enhancing the calcineurin/NFAT pathway. miR-17~92 binds to and represses a network of genes including several negative regulators of T cell activation. Finally, CD28-deficient T cells exhibit derepressed miR-17~92 target genes during activation, demonstrating that this non-coding RNA is required to shape the transcriptome. Thus, we propose that miR-17~92 constitutes a central mediator for T cell activation, integrating signals by the TCR and CD28 costimulation. In this model miR-17~92 facilitates T cell activation by dampening the breaks that prevent T cell activation.

immunology↗

bioRxiv2020

In vitro culture of aberrant basal-like cells from fibrotic lung tissue

RationaleIn idiopathic pulmonary fibrosis (IPF) atypical epithelial cells are present in the alveolar compartment. Their origin and contribution to IPF pathogenesis is unknown. We recently cultured a distinct population of cells, which readily grew from fibrotic lung tissue, but only rarely from non-fibrotic tissue. Here we aimed to characterize these fibrosis-enriched cells and determine transcriptomic differences between cells derived from IPF and patients with other interstitial lung diseases (ILD). MethodsCells were cultured from peripheral lung tissue of ILD patients and analysed by bulk or single cell RNA sequencing (scRNA-seq), TaqMan-PCR, immunofluorescence (IF), immunoblotting or electron microscopy (EM). ResultsscRNA-seq demonstrated an overall homogeneity and epithelial origin of the cells. The majority of cells expressed basal cell markers (Cytokeratin (KRT) 5 and 17, TP63), of which a fraction co-expressed mesenchymal cell markers (VIM, FN1, CDH2), alveolar (SLC34A2, ABCA3, LPCAT1, EMP2, HOPX) and/or secretory epithelial cell markers (SCGB1A1, MUC4). Interestingly, most of the cells showed closest transcriptomic similarity to recently described aberrant basal-like cells. Cells derived from IPF versus other ILD patients revealed significant transcriptomic differences with an up-regulation of fibrosis-associated and a down-regulation of inflammatory pathways in IPF cells. ConclusionWe here confirm the presence of aberrant basal-like cells in fibrotic lung tissue and, importantly, are the first to describe their in vitro characteristics and a way of culturing these cells in vitro. Cultured basal-like cells co-express epithelial and mesenchymal markers, suggesting a partial epithelial to mesenchymal transition (EMT). A subset of cells co-express alveolar, ciliated or secretory epithelial cell markers, possibly indicating differentiation towards these cell linages. Furthermore, cultured basal-like cells display a disease-specific transcriptome, possibly induced by their specific microenvironment. Our findings will contribute to a better understanding of the cells origin and their potential contribution to IPF pathogenesis.

cell biology↗

bioRxiv2020

The Bgee suite: integrated curated expression atlas and comparative transcriptomics in animals

Bgee is a database to retrieve and compare gene expression patterns in multiple animal species, produced by integrating multiple data types (RNA-Seq, Affymetrix, in situ hybridization, and EST data). It is based exclusively on curated healthy wild-type expression data (e.g., no gene knock-out, no treatment, no disease), to provide a comparable reference of normal gene expression. Curation includes very large datasets such as GTEx (re-annotation of samples as "healthy" or not) as well as many small ones. Data are integrated and made comparable between species thanks to consistent data annotation and processing, and to calls of presence/absence of expression, along with expression scores. As a result, Bgee is capable of detecting the conditions of expression of any single gene, accommodating any data type and species. Bgee provides several tools for analyses, allowing, e.g., automated comparisons of gene expression patterns within and between species, retrieval of the prefered conditions of expression of any gene, or enrichment analyses of conditions with expression of sets of genes. Bgee release 14.1 includes 29 animal species, and is available at https://bgee.org/ and through its Bioconductor R package BgeeDB.

bioinformatics↗

bioRxiv2019

Air pollution mediates diabetes by disrupting gut innate mucosal immunity

Air pollution has emerged as an unexpected risk factor for diabetes1-6. The mechanisms linking air pollution and diabetes remain, however, unknown. It has been postulated that lung exposure mediates diabetes via systemic inflammation and insulin resistance7-9. By contrast, gut exposure to pollutants has received little attention, even though a large proportion of air pollution particles are swallowed after mucociliary clearance from the upper airways10. Here, we identified intestinal macrophages as key mediators linking air pollutants with impaired beta-cell function. Upon oral exposure to air pollution, intestinal macrophages up-regulated inflammatory- and interferon-response pathways, which disrupted their normal differentiation towards an anti-inflammatory/resident phenotype. The resulting pro-inflammatory milieu in the gut impaired beta-cell identity and function via local cytokine secretion from macrophages as genetic and pharmacological macrophage or IL-1{beta} ablation protected mice from developing air pollution-induced diabetes. These data establish intestinal macrophage-derived cytokines as key mediators of air pollution associated beta-cell dysfunction, thus pointing towards novel therapeutic strategies.

immunology↗