Host microbial cross-talk is essential to maintain intestinal homeostasis. However, maladaptation of this response through microbial dysbiosis or defective host defense toward invasive intestinal ...bacteria can result in chronic inflammation. We have shown that macrophages differentiated in the presence of the bacterial metabolite butyrate display enhanced antimicrobial activity. Butyrate-induced antimicrobial activity was associated with a shift in macrophage metabolism, a reduction in mTOR kinase activity, increased LC3-associated host defense and anti-microbial peptide production in the absence of an increased inflammatory cytokine response. Butyrate drove this monocyte to macrophage differentiation program through histone deacetylase 3 (HDAC3) inhibition. Administration of butyrate induced antimicrobial activity in intestinal macrophages in vivo and increased resistance to enteropathogens. Our data suggest that (1) increased intestinal butyrate might represent a strategy to bolster host defense without tissue damaging inflammation and (2) that pharmacological HDAC3 inhibition might drive selective macrophage functions toward antimicrobial host defense.
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•Butyrate induces differentiation of macrophages with potent antimicrobial function•Enhanced antimicrobial function is a consequence of glycolysis and mTOR inhibition•Single-cell RNA-sequencing identifies butyrate-induced antimicrobial peptides•Butyrate inhibits HDAC3 to drive metabolic changes and microbicidal function
Macrophages maintain gut homeostasis by eliminating invasive pathogens and regulating inflammatory responses. Schulthess et al. demonstrate that butyrate, a bacterial fermentation product, imprints potent antimicrobial activity during macrophage differentiation through HDAC3i function.
The identification of lymphocyte subsets with non-overlapping effector functions has been pivotal to the development of targeted therapies in immune-mediated inflammatory diseases (IMIDs)
. However, ...it remains unclear whether fibroblast subclasses with non-overlapping functions also exist and are responsible for the wide variety of tissue-driven processes observed in IMIDs, such as inflammation and damage
. Here we identify and describe the biology of distinct subsets of fibroblasts responsible for mediating either inflammation or tissue damage in arthritis. We show that deletion of fibroblast activation protein-α (FAPα)
fibroblasts suppressed both inflammation and bone erosions in mouse models of resolving and persistent arthritis. Single-cell transcriptional analysis identified two distinct fibroblast subsets within the FAPα
population: FAPα
THY1
immune effector fibroblasts located in the synovial sub-lining, and FAPα
THY1
destructive fibroblasts restricted to the synovial lining layer. When adoptively transferred into the joint, FAPα
THY1
fibroblasts selectively mediate bone and cartilage damage with little effect on inflammation, whereas transfer of FAPα
THY1
fibroblasts resulted in a more severe and persistent inflammatory arthritis, with minimal effect on bone and cartilage. Our findings describing anatomically discrete, functionally distinct fibroblast subsets with non-overlapping functions have important implications for cell-based therapies aimed at modulating inflammation and tissue damage.
Immune-regulatory mechanisms of drug-free remission in rheumatoid arthritis (RA) are unknown. We hypothesized that synovial tissue macrophages (STM), which persist in remission, contribute to joint ...homeostasis. We used single-cell transcriptomics to profile 32,000 STMs and identified phenotypic changes in patients with early/active RA, treatment-refractory/active RA and RA in sustained remission. Each clinical state was characterized by different frequencies of nine discrete phenotypic clusters within four distinct STM subpopulations with diverse homeostatic, regulatory and inflammatory functions. This cellular atlas, combined with deep-phenotypic, spatial and functional analyses of synovial biopsy fluorescent activated cell sorted STMs, revealed two STM subpopulations (MerTK
TREM2
and MerTK
LYVE1
) with unique remission transcriptomic signatures enriched in negative regulators of inflammation. These STMs were potent producers of inflammation-resolving lipid mediators and induced the repair response of synovial fibroblasts in vitro. A low proportion of MerTK
STMs in remission was associated with increased risk of disease flare after treatment cessation. Therapeutic modulation of MerTK
STM subpopulations could therefore be a potential treatment strategy for RA.
The severe acute respiratory syndrome coronavirus 2 (SARS‑CoV‑2) disease (COVID-19) pandemic has caused millions of deaths worldwide. Genome-wide association studies identified the 3p21.31 region as ...conferring a twofold increased risk of respiratory failure. Here, using a combined multiomics and machine learning approach, we identify the gain-of-function risk A allele of an SNP, rs17713054G>A, as a probable causative variant. We show with chromosome conformation capture and gene-expression analysis that the rs17713054-affected enhancer upregulates the interacting gene, leucine zipper transcription factor like 1 (LZTFL1). Selective spatial transcriptomic analysis of lung biopsies from patients with COVID-19 shows the presence of signals associated with epithelial-mesenchymal transition (EMT), a viral response pathway that is regulated by LZTFL1. We conclude that pulmonary epithelial cells undergoing EMT, rather than immune cells, are likely responsible for the 3p21.31-associated risk. Since the 3p21.31 effect is conferred by a gain-of-function, LZTFL1 may represent a therapeutic target.
Increased cortical size is essential to the enhanced intellectual capacity of primates during mammalian evolution. The mechanisms that control cortical size are largely unknown. Here, we show that ...mammalian BAF170, a subunit of the chromatin remodeling complex mSWI/SNF, is an intrinsic factor that controls cortical size. We find that conditional deletion of BAF170 promotes indirect neurogenesis by increasing the pool of intermediate progenitors (IPs) and results in an enlarged cortex, whereas cortex-specific BAF170 overexpression results in the opposite phenotype. Mechanistically, BAF170 competes with BAF155 subunit in the BAF complex, affecting euchromatin structure and thereby modulating the binding efficiency of the Pax6/REST-corepressor complex to Pax6 target genes that regulate the generation of IPs and late cortical progenitors. Our findings reveal a molecular mechanism mediated by the mSWI/SNF chromatin-remodeling complex that controls cortical architecture.
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•BAF170 regulates cortical size and thickness•BAF170 controls the mode of cortical neurogenesis•Interaction of BAF170, Pax6, and REST controls the timing of cortical layer generation•Competition between BAF155 and BAF170 subunits controls chromatin structure
Tuoc et al. show that BAF chromatin remodeling complex component BAF170 controls cortical size via cortical neurogenesis. BAF170 competes with BAF155 subunit in the complex. The BAF170-containing complex interacts with Pax6 and modulates Pax6 target gene expression via REST corepressor complex recruitment to control intermediate and later cortical progenitor generation.
Neural stem cell self-renewal, neurogenesis, and cell fate determination are processes that control the generation of specific classes of neurons at the correct place and time. The transcription ...factor Pax6 is essential for neural stem cell proliferation, multipotency, and neurogenesis in many regions of the central nervous system, including the cerebral cortex. We used Pax6 as an entry point to define the cellular networks controlling neural stem cell self-renewal and neurogenesis in stem cells of the developing mouse cerebral cortex. We identified the genomic binding locations of Pax6 in neocortical stem cells during normal development and ascertained the functional significance of genes that we found to be regulated by Pax6, finding that Pax6 positively and directly regulates cohorts of genes that promote neural stem cell self-renewal, basal progenitor cell genesis, and neurogenesis. Notably, we defined a core network regulating neocortical stem cell decision-making in which Pax6 interacts with three other regulators of neurogenesis, Neurog2, Ascl1, and Hes1. Analyses of the biological function of Pax6 in neural stem cells through phenotypic analyses of Pax6 gain- and loss-of-function mutant cortices demonstrated that the Pax6-regulated networks operating in neural stem cells are highly dosage sensitive. Increasing Pax6 levels drives the system towards neurogenesis and basal progenitor cell genesis by increasing expression of a cohort of basal progenitor cell determinants, including the key transcription factor Eomes/Tbr2, and thus towards neurogenesis at the expense of self-renewal. Removing Pax6 reduces cortical stem cell self-renewal by decreasing expression of key cell cycle regulators, resulting in excess early neurogenesis. We find that the relative levels of Pax6, Hes1, and Neurog2 are key determinants of a dynamic network that controls whether neural stem cells self-renew, generate cortical neurons, or generate basal progenitor cells, a mechanism that has marked parallels with the transcriptional control of embryonic stem cell self-renewal.
Promiscuous gene expression (PGE) by thymic epithelial cells (TEC) is essential for generating a diverse T cell antigen receptor repertoire tolerant to self-antigens, and thus for avoiding ...autoimmunity. Nevertheless, the extent and nature of this unusual expression program within TEC populations and single cells are unknown. Using deep transcriptome sequencing of carefully identified mouse TEC subpopulations, we discovered a program of PGE that is common between medullary (m) and cortical TEC, further elaborated in mTEC, and completed in mature mTEC expressing the autoimmune regulator gene (Aire). TEC populations are capable of expressing up to 19,293 protein-coding genes, the highest number of genes known to be expressed in any cell type. Remarkably, in mouse mTEC, Aire expression alone positively regulates 3980 tissue-restricted genes. Notably, the tissue specificities of these genes include known targets of autoimmunity in human AIRE deficiency. Led by the observation that genes induced by Aire expression are generally characterized by a repressive chromatin state in somatic tissues, we found these genes to be strongly associated with H3K27me3 marks in mTEC. Our findings are consistent with AIRE targeting and inducing the promiscuous expression of genes previously epigenetically silenced by Polycomb group proteins. Comparison of the transcriptomes of 174 single mTEC indicates that genes induced by Aire expression are transcribed stochastically at low cell frequency. Furthermore, when present, Aire expression-dependent transcript levels were 16-fold higher, on average, in individual TEC than in the mTEC population.
Dysregulated hematopoiesis occurs in several chronic inflammatory diseases, but it remains unclear how hematopoietic stem cells (HSCs) in the bone marrow (BM) sense peripheral inflammation and ...contribute to tissue damage in arthritis. Here, we show the HSC gene expression program is biased toward myelopoiesis and differentiation skewed toward granulocyte-monocyte progenitors (GMP) during joint and intestinal inflammation in experimental spondyloarthritis (SpA). GM-CSF-receptor is increased on HSCs and multipotent progenitors, favoring a striking increase in myelopoiesis at the earliest hematopoietic stages. GMP accumulate in the BM in SpA and, unexpectedly, at extramedullary sites: in the inflamed joints and spleen. Furthermore, we show that GM-CSF promotes extramedullary myelopoiesis, tissue-toxic neutrophil accumulation in target organs, and GM-CSF prophylactic or therapeutic blockade substantially decreases SpA severity. Surprisingly, besides CD4
T cells and innate lymphoid cells, mast cells are a source of GM-CSF in this model, and its pathogenic production is promoted by the alarmin IL-33.
Microglia are increasingly implicated in brain pathology, particularly neurodegenerative disease, with many genes implicated in Alzheimer's, Parkinson's, and motor neuron disease expressed in ...microglia. There is, therefore, a need for authentic, efficient in vitro models to study human microglial pathological mechanisms. Microglia originate from the yolk sac as MYB-independent macrophages, migrating into the developing brain to complete differentiation. Here, we recapitulate microglial ontogeny by highly efficient differentiation of embryonic MYB-independent iPSC-derived macrophages then co-culture them with iPSC-derived cortical neurons. Co-cultures retain neuronal maturity and functionality for many weeks. Co-culture microglia express key microglia-specific markers and neurodegenerative disease-relevant genes, develop highly dynamic ramifications, and are phagocytic. Upon activation they become more ameboid, releasing multiple microglia-relevant cytokines. Importantly, co-culture microglia downregulate pathogen-response pathways, upregulate homeostatic function pathways, and promote a more anti-inflammatory and pro-remodeling cytokine response than corresponding monocultures, demonstrating that co-cultures are preferable for modeling authentic microglial physiology.
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•iPSC microglia co-culture model with iPSC cortical neurons•Expression of microglia markers in co-culture•Co-culture promotes microglial ramification and motility•Co-culture-specific cytokine release profiles
Microglia are implicated in neurodegenerative disease, and authentic human in vitro models are needed. The iPSC microglia model described here recapitulates microglia ontogeny by differentiating MYB-independent embryonic iPSC macrophages, then co-culturing them with iPSC cortical neurons. Co-culture microglia express microglia-specific markers and neurodegenerative disease genes, upregulate homeostatic pathways, develop dynamic ramifications, are phagocytic, and produce a cytokine profile that is co-culture specific.
The mechanisms by which early spatiotemporal expression patterns of transcription factors such as Pax6 regulate cortical progenitors in a region-specific manner are poorly understood. Pax6 is ...expressed in a gradient across the developing cortex and is essential for normal corticogenesis. We found that constitutive or conditional loss of Pax6 increases cortical progenitor proliferation by amounts that vary regionally with normal Pax6 levels. We compared the gene expression profiles of equivalent Pax6-expressing progenitors isolated from Pax6+/+ and Pax6−/− cortices and identified many negatively regulated cell-cycle genes, including Cyclins and Cdks. Biochemical assays indicated that Pax6 directly represses Cdk6 expression. Cyclin/Cdk repression inhibits retinoblastoma protein (pRb) phosphorylation, thereby limiting the transcription of genes that directly promote the mechanics of the cell cycle, and we found that Pax6 inhibits pRb phosphorylation and represses genes involved in DNA replication. Our results indicate that Pax6’s modulation of cortical progenitor cell cycles is regional and direct.
•Pax6 loss increases cortical progenitor proliferation by region-specific amounts•The size of this effect correlates directly with normal Pax6 expression levels•Expression of many key cell-cycle regulators is increased in the absence of Pax6•Pax6 directly represses Cdk6 expression and controls pRb phosphorylation
The transcription factor Pax6 regulates multiple aspects of forebrain development. Mi et al. show that it exerts regional control of cortical cell proliferation. It inhibits cell-cycle gene expression, directly repressing Cdk6, and limits the activity of the pRb-E2F pathway.
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