Biology subjects

Hume, D. A.

Publications and source records attributed to Hume, D. A..

At least 19 recordsLinked to original sources

bioRxiv2026

Macrophage calcium signaling dynamics revealed using genetically-encoded sensors

Macrophages (M{Phi}s) inhabit all mammalian organs, adapting to and influencing the tissues in which they reside. These versatile cells rapidly respond to tissue-specific signals, clean up debris and serve as the first line of defense against pathogens. Their ability to act quickly relates to their abundance and specific arrangement in tissues, as well as the systems they possess for detecting, processing and integrating emergent signals. Ca2+ is a fast second messenger that has been linked to many of the core functions of M{Phi}s. However, spatiotemporal features of physiologically-relevant Ca2+ signals in M{Phi}s remain largely uncharacterized. Using mouse models expressing genetic biosensors and fluorescently tagged channels, we visualize M{Phi} Ca2+ signal dynamics and heterogeneity, uncover a remarkable degree of mechanosensitivity in these cells, and characterize the physiological consequences of genetic ablation of Piezo1 channels via analysis of knockout models across their lifetime. Our in-depth investigation of M{Phi} Ca2+ signaling dynamics has broad relevance for the field of M{Phi} biology and the tissues that these cells support.

immunology↗

bioRxiv2026

Depletion and replacement of tissue resident macrophages in mice with germ-line deletion of a conserved enhancer in the Csf1r locus.

Expression of the Csf1r gene in cells of the mononuclear phagocyte lineage is regulated by a conserved enhancer, the fms-intronic regulatory element (FIRE). In mice with a germ-line deletion of FIRE (Fireko) CSF1R expression is undetectable in bone marrow progenitors and classical monocytes. Fireko mice lack subpopulations of macrophages in the brain and periphery but develop normally. Here we show that loss of CSF1R expression in Fireko mice is partly overcome by CSF2 in vitro and inflammatory recruitment in vitro. Analysis of heterozygous mutant mice and deletion of the conserved AP1 motif in FIRE provide evidence that continuous receptor synthesis determines CSF1 responsiveness. The absence of macrophages in kidney and heart of Fireko mice was not associated with detectable loss of physiological function. In a model of renal injury macrophage recruitment and histopathology were similar in WT and Fireko mice. Tissue resident macrophages that were depleted in Fireko mice, including microglia, were replaced by donor-derived cells following intraperitoneal adoptive transfer of wild-type bone marrow at weaning. The Fireko mouse provides a novel platform to dissect the functions of tissue resident macrophages in development, homeostasis and pathology. Summary StatementThis study describes a unique model of selective tissue resident macrophage deficiency arising from dysregulated expression of the mouse Csf1r gene.

immunology↗

bioRxiv2026

CSF1R-dependent macrophages control B cell development and function in the chicken immune system.

Acquired immunity in mammals depends upon capture and presentation of antigens by specialised macrophage populations in splenic marginal zone and lymph node sinuses and follicular dendritic cells (FDC) within germinal centres. Cells referred to as FDC in chickens express CSF1R, the receptor for macrophage colony-stimulating factor (CSF1) and IL34. We utilised single cell RNA-seq on CSF1R+ cells from chicken spleen to identify monocytes and two distinct populations of macrophages. TIMD4/C1Q/MAFB+ macrophages were enriched for expression of genes involved in iron metabolism. A MARCO/VSIG4+ population expressed SPIC, a transcription factor associated with red pulp macrophages in mammals but also expressed receptors (CR2) and trophic factors (TNFSF13, CXCL13) associated with mammalian FDC. SPIC+ cells were located within follicles in spleen, caecal tonsil and bursa. We generated a CSF1R knockout in the chicken germ line. Mutant birds lack macrophages in the embryo. They were indistinguishable from wild type at hatch and behaved and fed normally but from day 5-6 post hatch they failed to thrive. Loss of CSF1R function in hatchlings led to monocytopenia and granulocytosis and the loss of macrophage subpopulations in lymphoid organs. Consistent with their expression of B cell trophic factors, the loss of follicular macrophages in the bursa was associated with involution and severe B cell deficiency in the circulation and spleen. In summary, lymphoid tissues of chickens contain specialised macrophage populations with distinct expression profiles. The details of regulation by CSF1R, specialised functions and underlying transcriptional regulation are quite different between birds and mammals.

immunology↗

bioRxiv2025

Microglia are not required for normal postnatal brain development and cognitive function in the rat.

The development, maintenance and survival of microglia, the brains resident immune cells, requires signalling via the colony stimulating factor 1 receptor (CSF1R). In mice, the Fms intronic regulatory element (FIRE), a highly conserved super enhancer, is required for CSF1R expression in microglia and germ-line deletion of FIRE leads to microglial deficiency. We have generated Csf1r{Delta}FIRE/{Delta}FIRE (Fireko) rats using CRISPR/Cas9. Fireko rats are born at expected Mendelian frequency, grow normally and survive to adulthood. Like Fireko mice, they lack microglia and subpopulations of resident macrophages in kidney and heart, but retain the other brain associated macrophages and resident populations in the liver, spleen and lungs. In contrast to mutant mice, macrophages in the peritoneum are unaffected and CSF1R expression is not completely abolished in the blood. Aside from the loss of microglia-specific transcripts, the Fireko has no effect on expression of genes associated with neurons, oligodendrocytes or astrocytes in adult rats but is associated with detectable calcification in the thalamus. Despite the absence of microglia, we detected no significant differences in ethologically relevant behaviours compared to littermates. The results suggest that microglia are not essential for normal cognitive development. Fireko rats provide an alternative to mice as a model for human microglial deficiency and the functions of microglia in development and homeostasis in the brain.

immunology↗

bioRxiv2025

A conserved distal element in the mouse Csf1r locus contributes to transcription in hematopoietic and trophoblast cells.

Expression of the Csf1r gene in mice is restricted to cells of the mononuclear phagocyte system and placental trophoblasts. A conserved element (Csf1r upstream regulatory element A, CUREA) in the mouse Csf1r locus contains transcription start sites utilised by trophoblasts and osteoclasts and an enhancer essential for expression of multicopy transgenic reporters in most tissue macrophages. Here we describe the impact of deletion of CUREA in the mouse genome, on the background of a knock-in Csf1r-FusionRed reporter. By contrast to the essential function in transgene expression, CUREA deletion had no effect on expression of FusionRed or differentiation of blood monocytes or tissue resident macrophages. The deletion reduced Csf1r mRNA in hematopoietic stem cells and committed myeloid progenitors (MPP3) leading to a subtle differentiation delay and also had a significant impact on microglial phenotype in the brain and the differentiation of osteoclasts. The expression of FusionRed in placenta confirmed expression of CSF1R in trophoblasts. 5RACE analysis demonstrated that the effect of CUREA deletion on Csf1r transcription in placenta was overcome by the use of cryptic upstream transcription start sites. We conclude that CUREA is a regulatory element controlling Csf1r transcription. The function overlaps with other enhancers identified in the locus and is therefore partly redundant. Key PointsO_LIA regulatory element (CUREA) in the mouse Csf1r locus has both promoter and enhancer activity. C_LIO_LIGerm-line deletion of CUREA impacts differentiation of marrow progenitors, microglia, osteoclasts and placental trophoblasts. C_LI

cell biology↗

bioRxiv2025

CSF1R regulates monocyte subset differentiation and intracellular metabolism.

Monocytes are key circulating effectors of vascular homeostasis, innate immunity and inflammation. Following their generation in mouse bone marrow, classical (Ly6Chigh) monocytes are mobilized into the blood circulation where they mature into non-classical (Ly6Clow) patrolling monocytes or are recruited into peripheral tissues where they differentiate into tissue resident or inflammatory macrophages. Monocytes and macrophages express CSF1R (CD115), the receptor for lineage-specific growth factors CSF1 and IL34. Here, we report that acute CSF1R blockade or genetic deletion negatively interferes with monocyte intracellular metabolism and reduces blood Ly6Clow monocytes in part by blunting differentiation of Ly6Chigh monocytes. Based upon lineage-specific deletion of GFPT1 (Glutamine-Fructose-6-Phosphate Transaminase 1), the hexosamine biosynthetic pathway (HBP) is identified as a novel regulator of CSF1R expression and monocyte subset diversity. Our findings provide new insights into the link between CSF1R signaling, metabolic regulation, and monocyte survival and differentiation.

immunology↗

bioRxiv2025

Homozygous kinase-dead Csf1r mutation in outbred mice reveals essential and redundant functions of tissue resident macrophages

The proliferation, differentiation and survival of cells of the macrophage lineage depends on signals from the macrophage colony-stimulating factor receptor (CSF1R). On a C57BL/6J background homozygous kinase-dead Csf1r mutation (Csf1rE631K/E631K - E631Km/m) causes perinatal lethality. Here we demonstrate that E631Km/m mice on a mixed genetic background (C57 x BALB/c F2) are osteopetrotic and growth retarded but viable as adults with no other gross developmental deficits. They lack osteoclasts, microglia and most peripheral tissue resident macrophages and exhibit perturbed hematopoiesis. Although CD169+ tissue resident macrophages in bone marrow are considered an essential component of the hematopoietic niche, CD169 is undetectable in E631Km/m marrow and F4/80+ macrophages are depleted. These changes are associated with expansion of mature and immature granulocytes and reduced B cells, whereas monocytes and stem and progenitor populations are unaffected as a proportion of total cells. Erythropoiesis in bone marrow is maintained in E631Km/m mice, associated with a residual population of CSF1R-independent CD169-ve/F4/80+ macrophages. Nevertheless, splenic extramedullary hematopoiesis in E631Km/m mice indicates a degree of bone marrow insufficiency. Red pulp macrophages are retained but CD169+ marginal metallophil macrophages are absent and CD209b+ (SIGNR1) macrophages are present but disorganized. Circulating white blood cell count is unchanged in E631Km/m mice, but the proportion of neutrophils is greatly increased whilst B cells and monocytes are reduced. This novel model reveals the essential roles of CSF1R-dependent macrophages in hematopoiesis and demonstrates that many developmental and homeostatic functions attributed to murine resident tissue macrophages are redundant and/or specific to inbred mouse strains.

immunology↗

bioRxiv2025

Perisurgical Colony Stimulating Factor 1 (CSF1) treatment ameliorates liver 1 ischaemia/reperfusion injury in rats

BackgroundIn the context of hepatobiliary and liver transplant surgery, ischemia-reperfusion (I/R) injury can occur due to temporary interruption of blood flow to the organ followed by a potentially damaging inflammatory response to reperfusion. As macrophages can promote liver growth and contribute to resolution of chronic liver injury and fibrosis, we tested the hypothesis that stimulation of monocytes and macrophages with Colony Stimulating Factor 1 (CSF1) could have a beneficial impact on liver repair after I/R injury. MethodsWe investigated the impact of perisurgical treatment with a long-circulating CSF1-Fc conjugate on liver injury and hepatocyte proliferation after 70% ischemia for 60 minutes at 6 h, 48 h and 7 days post reperfusion in male rats. Changes in the circulating and liver tissue monocyte and macrophage subsets in the ischaemic and oxygenated lobes were assessed using quantitative PCR and flow cytometry. ResultsCSF1-Fc treatment did not affect the extent of hepatocellular injury post-reperfusion, as indicated by serum transaminases. Liver I/R injury, especially necrotic area, was reduced in CSF1-Fc-treated rats 48 h post-surgery. This was associated with increased accumulation of macrophages in both the oxygenated and ischemic lobes, and localization to necrotic tissue in the ischemic lobe. CSF1-Fc treatment also promoted liver growth, associated with increased parenchymal and non-parenchymal cell proliferation. Flow cytometry and gene expression analysis demonstrated increased monocyte infiltration in ischemic compared to oxygenated lobes. CSF1-Fc increased the abundance of CD43+ non-classical monocytes, consistent with the role of CSF1 signaling in monocyte maturation, and increased CD163 expression on mature macrophages. ConclusionThis study suggests CSF1 stimulation drives monocytes/macrophages towards a pro-regenerative response and perisurgical CSF1 treatment might augment liver regeneration in patients undergoing liver resection. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=140 SRC="FIGDIR/small/642926v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@181eb0borg.highwire.dtl.DTLVardef@6e3370org.highwire.dtl.DTLVardef@f32022org.highwire.dtl.DTLVardef@1c2c1b3_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2025

Mutation in the rat interleukin 34 gene impacts macrophage development, homeostasis and inflammation in the brain and periphery

Interleukin-34 (IL34) and colony stimulating factor 1 (CSF1) signal through a shared receptor (CSF1R) to control macrophage survival, differentiation and function. Here we describe the impact of loss of function mutation in the rat Il34 gene. Il34-/- rats showed a partial reduction in macrophages within squamous epithelia (Langerhans-like cells) and in the testis. In the brain, microglia and brain-associated macrophages were selectively depleted in grey matter. A gradient of microglial density in Il34-/- cortex suggests that CSF1 can diffuse outwards from the corpus callosum. The reduced density of microglia was not associated with detectable neuropathology or behavioural alterations. In RNA-seq analysis of cortex, hippocampus and thalamus the only change is selective and uniform loss of the microglial signature. In the periphery, increased Il34 expression has been associated with epithelial injury. In the adenine diet model of renal interstitial fibrosis both Il34 and Csf1 were induced. The absence of IL34 led to a significant reduction in macrophage recruitment compared to controls, but pathology assessed histologically or by detection of damage-associated mRNA signature was unaffected. We suggest that IL34 and CSF1 provide redundant signals to sustain microglia and to direct macrophage recruitment and repair tissue injury in the periphery.

immunology↗

bioRxiv2025

Repopulation of the brain with microglia-like cells following intraperitoneal bone marrow cell transfer in microglia-deficient mice.

Germ-line deletion of a conserved enhancer (the Fms intrinsic regulatory element, FIRE) in the mouse Csf1r locus causes congenital absence of microglia. Homozygous FIRE deletion on a C57BL/6J background leads to perinatal lethality and hydrocephalus (HC) in surviving pups. We developed a congenic C57BL/6J line with defined regions of non-C57BL/6J genomic DNA, increased postnatal viability and reduced incidence of HC. Both perinatal lethality and HC were eliminated in F2 mice following outcross of the congenic line to CBA/J or BALBc/J backgrounds. To assess the impacts of microglial deficiency in postnatal neurodevelopment we analyzed deep total RNA-seq data from multiple brain regions of wild-type and Csf1r{Delta}FIRE/{Delta}FIRE mice. Aside from the loss of microglial-specific transcripts, we found no significant alterations in relative abundance of any cell-type or region-specific transcriptomic signature. Transcripts associated with endosome/lysosome function, which are enriched in microglia, were not affected, suggesting compensatory expression by other cell types. On the C57BL/6J x CBA/J F2 background, congenital absence of microglia did not affect motor activity, behavior or myelination up to 7 months of age but was associated with astrocytosis and calcification in the thalamus. In the congenic C57BL/6J Csf1r{Delta}FIRE/{Delta}FIRE mouse line, intraperitoneal transfer of wild-type bone marrow cells (BMT) at weaning led to complete repopulation of the brain with microglia-like cells without giving rise to monocytic intermediates. Our results suggest novel strategies for treatment of microglial deficiency.

neuroscience↗

bioRxiv2025

Interleukin 10 controls the balance between tolerance, pathogen elimination and immunopathology in birds.

Effective mucosal immunity in the intestine involves a fine balance between tolerance of the microbiome, recognition and elimination of pathogens, and inflammatory tissue injury. The anti-inflammatory cytokine IL10 regulates these processes in the intestines of mice and humans; the anti-inflammatory activity of IL10 is also conserved in birds. To determine the function of IL10 in avian mucosal immunity, we generated germ line modifications of the chicken IL10 locus to abolish or reduce IL10 expression. In vitro analysis of macrophage response to lipopolysaccharide confirmed the loss of IL10 protein expression, the lack of dosage compensation in heterozygotes, and prevention of autocrine inhibition of nitric oxide production in homozygous IL10 knockout macrophages. IL10-deficiency significantly altered the composition of the caecal microbiome, but unlike IL10-deficient mice and humans, IL10-deficient chickens did not exhibit spontaneous colitis. Following experimental challenge with Salmonella enterica serovar Typhimurium or Campylobacter jejuni in IL10-deficient chickens, enhanced clearance of the pathogens was associated with elevated transcription of pro-inflammatory genes and increased infiltration of inflammatory cells into gut mucosa. In IL10-deficient chickens challenged with the parasite Eimeria tenella, pathogen clearance was accelerated but caecal lesions were more severe and weight gain was compromised. Neither the heterozygous IL10 knockout nor a homozygous IL10 enhancer mutation had a major effect on pathogen clearance or inflammation in any of the challenge models. Our findings highlight the intrinsic compromise in mucosal immune response and have important implications for the development of strategies to combat avian and zoonotic pathogens in poultry.

immunology↗

bioRxiv2024

Transcriptomic signatures of host immune responses in aphthous ulcers, the earliest lesions of Crohn's disease, suggest that a bacterial invasive challenge, rather than global dysbiosis, is the initiating factor.

Crohns disease is a chronic, transmural inflammatory disease of the human gut. Changes in the faecal microbial composition and a reduction in species diversity (dysbiosis) are consistent features in studies of Crohns disease patients, but whether dysbiosis is a cause or consequence of inflammation remains unresolved. Genetic susceptibility also plays a role in the development of Crohns disease and has been linked to genes involved in recognition of intestinal bacteria by cells of the mononuclear phagocyte system. The earliest visible lesions in Crohns disease are aphthous ulcers, overlying Peyers patches and lymphoid follicles. To identify mechanisms underlying the earliest stages of disease initiation we compared gene expression in aphthous ulcers, Peyers patches, inflamed and endoscopically normal mucosa from patients and controls using total RNA-seq. The resulting data was subjected to network analysis to identify co-regulated gene expression signatures of cell types and processes. These results were compared to recent single cell RNA-seq analysis of intestinal macrophages in normal and diseased mucosa. The analysis of aphthous ulcers revealed signatures of epithelial stress and antimicrobial defence, plasma cell activation and immunoglobulin production, monocyte recruitment, inflammatory gene expression and induction of interferon-{gamma} and downstream target genes. These signatures were not present in the normal appearing mucosa adjacent to aphthous ulcers which were similar to healthy control mucosa. We conclude that the initial lesion in Crohns disease arises from an invasive bacterial challenge leading to intense activation of multiple host defence pathways rather than the breakdown of epithelial barrier integrity and widespread bacterial translocation.

immunology↗

bioRxiv2024

Typical development of synaptic and neuronal properties can proceed without microglia

Brain-resident macrophages, microglia, have been proposed to play an active role in synaptic refinement and maturation, influencing plasticity and circuit-level connectivity. Here we show that a variety of neurodevelopmental processes previously attributed to microglia can proceed without them. Using a genetically modified mouse which lacks microglia (Csf1r{Delta}FIRE/{Delta}FIRE) we find that intrinsic properties, synapse number and synaptic maturation are largely normal in the hippocampal CA1 region and somatosensory cortex at stages where microglia have been implicated. Seizure susceptibility and hippocampal-prefrontal cortex coherence in awake behaving animals, processes disrupted in mice deficient in microglia-enriched genes, are also normal. Similarly, eye-specific segregation of inputs into the lateral geniculate nucleus proceeds normally in the absence of microglia. Single cell/nucleus transcriptomic analyses of neurons and astrocytes did not uncover any substantial perturbation due to microglial absence. Thus, the brain possesses remarkable adaptability to execute developmental synaptic refinement, maturation and connectivity in the absence of microglia.

neuroscience↗

bioRxiv2024

Microglia determine beta-amyloid plaque burden but are non-essential for downstream pathology

Evidence points to a role for microglia in Alzheimers disease (AD) risk, although their position in the pathological cascade is incompletely understood, prompting us to generate a model of {beta}-amyloidopathy lacking microglia. We find evidence that microglia promote plaque formation and creation of an A{beta} fibril-rich zone surrounding the plaque core. However, plaque-proximal reactive astrogliosis, synapse loss, and neurite dystrophy are still observed in the absence of microglia.

neuroscience↗

bioRxiv2024

Transcriptomic analysis of the functions of CSF1R-dependent macrophages in postnatal development in the rat

Adaptation to existence outside the womb is a key event in the life of a mammal. The absence of macrophages in rats with a homozygous mutation in the Csf1r gene (Csf1rko) severely compromises pre-weaning somatic growth and maturation of organ function. Transfer of wild-type bone marrow cells (BMT) at weaning rescues tissue macrophage populations permitting normal development and long-term survival. To dissect the phenotype and function of macrophages in postnatal development, we generated transcriptomic profiles of all major organs of wild-type and Csf1rko rats at weaning and selected organs following rescue by BMT. The transcriptomic profiles revealed subtle effects of macrophage deficiency on development of all major organs. Network analysis revealed a common signature of CSF1R-dependent resident tissue macrophages that includes the components of complement C1Q (C1qa/b/c genes). Circulating C1Q was almost undetectable in Csf1rko rats and rapidly restored to normal levels following BMT. Tissue-specific macrophage signatures were also identified, notably including sinus macrophage populations in the lymph nodes. Their loss in Csf1rko rats was confirmed by immunohistochemical localisation of CD209B (SIGNR1). By 6-12 weeks, Csf1rko rats succumb to emphysema-like pathology associated with the selective loss of interstitial macrophages and granulocytosis. This pathology was reversed by BMT. Along with physiological rescue, BMT precisely regenerated the abundance and expression profiles of resident macrophages. The exception was the brain, where BM-derived microglia-like cells had a distinct expression profile compared to resident microglia. In addition, the transferred BM failed to restore blood monocyte or CSF1R-positive bone marrow progenitors. These studies provide a model for the pathology and treatment of CSF1R mutations in humans and the innate immune deficiency associated with prematurity.

immunology↗

bioRxiv2024

The effect of a dominant kinase-dead Csf1r mutation associated with adult-onset leukoencephalopathy on brain development and neuropathology.

Amino acid substitutions in the kinase domain of the human CSF1R protein are associated with autosomal dominant adult-onset leukoencephalopathy with axonal spheroids and pigmented glia (ALSP). To model the human disease, we created a disease-associated mutation (Glu631Lys; E631K) in the mouse Csf1r locus. Previous analysis demonstrated that heterozygous mutation (Csf1rE631K/+) had a dominant inhibitory effect on CSF1R signaling in vitro and in vivo but did not recapitulate the pathology of the human disease. We speculated that leukoencephalopathy in humans requires an environmental trigger and/or epistatic interaction with common neurodegenerative disease-associated alleles. Here we examine the impact of heterozygous Csf1r mutation on microglial phenotype, normal postnatal brain development, age-related changes in gene expression and on two distinct pathologies in which microgliosis is a prominent feature, prion disease and experimental autoimmune encephalitis (EAE). The heterozygous Csf1rE631K/+ mutation reduced microglial abundance and the expression of microglial-associated transcripts relative to wild-type controls at 12 weeks and 43 weeks of age but had no selective effect on homeostatic markers such as P2ry12. An epistatic interaction was demonstrated between Csf1rE631K/+ and Cxc3r1EGFP/+ genotypes leading to dysregulated microglial and neuronal gene expression in both hippocampus and striatum. Heterozygous Csf1rE631K mutation reduced the microgliosis associated with both diseases. There was no significant impact on disease severity or progression in prion disease. In EAE, induced expression of inflammation-associated transcripts in the hippocampus and striatum was suppressed in parallel with microglia-specific transcripts, but spinal cord demyelination was exacerbated. The results support a dominant-negative model of CSF1R-associated leukoencephalopathy and likely contributions of an environmental trigger and/or genetic background to neuropathology.

neuroscience↗

bioRxiv2023

Intraperitoneal transfer of wild-type bone marrow cells in the Csf1r knockout rat repopulates resident tissue macrophages without contributing to monocytopoiesis.

Homozygous null mutation of the macrophage colony-stimulating factor receptor (Csf1r) gene in rats leads to the loss of most tissue macrophage populations and has pleiotropic impacts on postnatal growth and organ maturation leading to mortality by 8-12 weeks of age. The phenotype of the Csf1r knockout (Csf1rko) can be reversed by intraperitoneal transfer of wild-type bone marrow cells (BMT) at weaning. Here we used a Csf1r-mApple transgenic reporter, which is expressed in neutrophils and B cells as well as monocytes and macrophages, to track the fate of donor-derived cells. Following BMT into Csf1r recipients, wild-type mApple+ve cells restored IBA1+ tissue macrophage populations in every tissue donor-derived cells also completely replaced recipient macrophages in organs such as spleen, lung and liver that were only partly macrophage-deficient in the Csf1rko. However, monocytes, neutrophils and B cells in bone marrow, blood and lymphoid tissues remained of recipient (mApple-ve) origin. An mApple+ve cell population expanded in the peritoneal cavity and invaded locally in the mesentery, fat pads, omentum and diaphragm. One week after BMT, distal organs contained foci of mApple+ve, IBA1-ve immature progenitors that appeared to proliferate, migrate and differentiate locally. We conclude that rat bone marrow contains progenitor cells that are able to restore and maintain all tissue macrophage populations in a Csf1rko rat directly without contributing to the bone marrow progenitor or blood monocyte populations.

immunology↗

bioRxiv2022

The association between transcriptional regulation of macrophage differentiation and activation and genetic susceptibility to inflammatory bowel disease

The abundant macrophage population of the intestinal lamina propria turns over rapidly and is replaced by blood monocytes. The differentiation and survival of resident intestinal macrophages depends upon signals from the macrophage colony-stimulating factor receptor (CSF1R). The response of human monocyte-derived macrophages (MDM) grown in macrophage colony-stimulating factor (CSF1) to bacterial lipopolysaccharide (LPS) has been proposed as a model for the differentiation and adaptation of monocytes entering the intestinal lamina propria. We hypothesized that dysregulation of this response leads to susceptibility to chronic inflammatory bowel disease (IBD). To address this hypothesis we analyzed transcriptomic variation in MDM from affected and unaffected sib pairs/trios from 22 IBD families and 6 healthy controls. There was no overall or inter-sib distinction between affected and unaffected individuals in basal gene expression or the stereotypical time course of the response to LPS. However, the basal or LPS-inducible expression of individual genes including inflammatory cytokines and many associated with IBD susceptibility in genome-wide association studies (GWAS) varied by as much as 100-fold between subjects. Extreme independent variation in the expression of pairs of HLA-associated transcripts (HLA-B/C, HLA-A/F and HLA-DRB1/DRB5) was associated with HLA genotype providing a novel explanation for the HLA association with disease susceptibility. The relationship between single nucleotide variant (SNV) genotype and gene expression at other loci was weaker and inconsistent suggesting that much of the variation arises from the integration of multiple trans-acting effects. For example, expression of IL1B at 2 hrs of LPS treatment was significantly associated with local SNV genotype and with peak expression of IL23A at 7 hrs. By contrast, there was no evidence of association between peak IL6 mRNA at 7hrs, IL6-associated SNV genotype or IL1B at 2 hrs. Our results support the view that gene-specific dysregulation in macrophage adaptation to the intestinal milieu provides a plausible explanation for genetic susceptibility to IBD. The analysis also suggests that the molecular basis of susceptibility is unique to each individual which may contribute to variation in the precise environmental trigger, the consequent pathology and response to treatment. Author summaryCells of the innate immune system called macrophages are abundant in the wall of the gut, providing a first line of defense against potential pathogens. These cells must also avoid an inappropriate or excessive response to the abundant microbial population (the microbiome) of the intestine. We have previously proposed that genetic differences between individuals in macrophage adaptation to the unique environment of the intestine underlie genetic susceptibility to inflammatory bowel disease (IBD). In this study we developed a model of the adaptation of macrophages and used that model to identify surprisingly extreme variation in the response amongst a cohort of affected and unaffected siblings in families with IBD. The response did not distinguish affected individuals from their unaffected siblings. Our results support the view that each individual within IBD-susceptible families carries a unique set of genetic variants of large effect that together predispose to uncontrolled gut inflammation in the face of an environmental trigger.

genetics↗