γδ TCRs Function as Innate-like Receptors in the Bovine γδ T Cell Response against Leptospira.

Leptospira serovar Hardjo are bacterial pathogens of cattle that also cause zoonotic disease in humans. Vaccine-mediated protection against Leptospira serovar Hardjo in cattle is associated with a workshop cluster 1 (WC1)+ γδ T cell response that can be recalled in vitro from PBMC by antigenic stimulation. This provides a model system in which to examine protective vaccine-induced γδ T cell responses in a γδ T cell high species. Only a small proportion (5-10%) of WC1+ γδ T cells from immunized cattle are Leptospira responders, implying that Ag specificity is determined by clonally distributed receptors. Both WC1 and TCR are known to be required for Leptospira-specific responses by bovine WC1+ γδ T cells. Through variegated expression patterns and V(D)J recombination, respectively, they have the capacity to confer Ag specificity. In this study, we develop and use a high-throughput TCR-sequencing approach to study the TCRγ and TCRδ repertoires of naive ex vivo PBMC, Leptospira-responding, and Leptospira nonresponding WC1+ γδ T cells to examine the potential role of γδ TCR in determining Ag specificity. Our results provide novel insights into the PBMC γδ TCR repertoires in cattle, demonstrating the TCRγ repertoire to be clonally stratified and essentially public, whereas the TCRδ repertoire shows much higher levels of clonal diversity and is essentially private. TCR repertoire analysis of Leptospira-responding WC1+ γδ T cells identifies no signature of TCR-mediated selection, suggesting that TCR functions largely as an innate-like receptor and does not act as a primary determinant of Ag specificity in the response to this pathogen.

Defining the pig microglial transcriptome reveals its core signature, regional heterogeneity, and similarity with human and rodent microglia.

Microglia play key roles in brain homeostasis as well as responses to neurodegeneration and neuroinflammatory processes caused by physical disease and psychosocial stress. The pig is a physiologically relevant model species for studying human neurological disorders, many of which are associated with microglial dysfunction. Furthermore, pigs are an important agricultural species, and there is a need to understand how microglial function affects their welfare. As a basis for improved understanding to enhance biomedical and agricultural research, we sought to characterize pig microglial identity at genome-wide scale and conduct inter-species comparisons. We isolated pig hippocampal tissue and microglia from frontal cortex, hippocampus, and cerebellum, as well as alveolar macrophages from the lungs and conducted RNA-sequencing (RNAseq). By comparing the transcriptomic profiles between microglia, macrophages, and hippocampal tissue, we derived a set of 239 highly enriched genes defining the porcine core microglial signature. We found brain regional heterogeneity based on 150 genes showing significant (adjusted p < 0.01) regional variations and that cerebellar microglia were most distinct. We compared normalized gene expression for microglia from human, mice and pigs using microglia signature gene lists derived from each species and demonstrated that a core microglial marker gene signature is conserved across species, but that species-specific expression subsets also exist. Our data provide a valuable resource defining the pig microglial transcriptome signature that validates and highlights pigs as a useful large animal species bridging between rodents and humans in which to study the role of microglia during homeostasis and disease.

Stem cell-derived porcine macrophages as a new platform for studying host-pathogen interactions.

BACKGROUND: Infectious diseases of farmed and wild animals pose a recurrent threat to food security and human health. The macrophage, a key component of the innate immune system, is the first line of defence against many infectious agents and plays a major role in shaping the adaptive immune response. However, this phagocyte is a target and host for many pathogens. Understanding the molecular basis of interactions between macrophages and pathogens is therefore crucial for the development of effective strategies to combat important infectious diseases. RESULTS: We explored how porcine pluripotent stem cells (PSCs) can provide a limitless in vitro supply of genetically and experimentally tractable macrophages. Porcine PSC-derived macrophages (PSCdMs) exhibited molecular and functional characteristics of ex vivo primary macrophages and were productively infected by pig pathogens, including porcine reproductive and respiratory syndrome virus (PRRSV) and African swine fever virus (ASFV), two of the most economically important and devastating viruses in pig farming. Moreover, porcine PSCdMs were readily amenable to genetic modification by CRISPR/Cas9 gene editing applied either in parental stem cells or directly in the macrophages by lentiviral vector transduction. CONCLUSIONS: We show that porcine PSCdMs exhibit key macrophage characteristics, including infection by a range of commercially relevant pig pathogens. In addition, genetic engineering of PSCs and PSCdMs affords new opportunities for functional analysis of macrophage biology in an important livestock species. PSCs and differentiated derivatives should therefore represent a useful and ethical experimental platform to investigate the genetic and molecular basis of host-pathogen interactions in pigs, and also have wider applications in livestock.

Pleiotropic Impacts of Macrophage and Microglial Deficiency on Development in Rats with Targeted Mutation of the Csf1r Locus.

We have produced Csf1r-deficient rats by homologous recombination in embryonic stem cells. Consistent with the role of Csf1r in macrophage differentiation, there was a loss of peripheral blood monocytes, microglia in the brain, epidermal Langerhans cells, splenic marginal zone macrophages, bone-associated macrophages and osteoclasts, and peritoneal macrophages. Macrophages of splenic red pulp, liver, lung, and gut were less affected. The pleiotropic impacts of the loss of macrophages on development of multiple organ systems in rats were distinct from those reported in mice. Csf1r-/- rats survived well into adulthood with postnatal growth retardation, distinct skeletal and bone marrow abnormalities, infertility, and loss of visceral adipose tissue. Gene expression analysis in spleen revealed selective loss of transcripts associated with the marginal zone and, in brain regions, the loss of known and candidate novel microglia-associated transcripts. Despite the complete absence of microglia, there was little overt phenotype in brain, aside from reduced myelination and increased expression of dopamine receptor-associated transcripts in striatum. The results highlight the redundant and nonredundant functions of CSF1R signaling and of macrophages in development, organogenesis, and homeostasis.

A high resolution atlas of gene expression in the domestic sheep (Ovis aries).

Sheep are a key source of meat, milk and fibre for the global livestock sector, and an important biomedical model. Global analysis of gene expression across multiple tissues has aided genome annotation and supported functional annotation of mammalian genes. We present a large-scale RNA-Seq dataset representing all the major organ systems from adult sheep and from several juvenile, neonatal and prenatal developmental time points. The Ovis aries reference genome (Oar v3.1) includes 27,504 genes (20,921 protein coding), of which 25,350 (19,921 protein coding) had detectable expression in at least one tissue in the sheep gene expression atlas dataset. Network-based cluster analysis of this dataset grouped genes according to their expression pattern. The principle of 'guilt by association' was used to infer the function of uncharacterised genes from their co-expression with genes of known function. We describe the overall transcriptional signatures present in the sheep gene expression atlas and assign those signatures, where possible, to specific cell populations or pathways. The findings are related to innate immunity by focusing on clusters with an immune signature, and to the advantages of cross-breeding by examining the patterns of genes exhibiting the greatest expression differences between purebred and crossbred animals. This high-resolution gene expression atlas for sheep is, to our knowledge, the largest transcriptomic dataset from any livestock species to date. It provides a resource to improve the annotation of the current reference genome for sheep, presenting a model transcriptome for ruminants and insight into gene, cell and tissue function at multiple developmental stages.

Macrophage colony-stimulating factor increases hepatic macrophage content, liver growth, and lipid accumulation in neonatal rats.

Signaling via the colony-stimulating factor 1 receptor (CSF1R) controls the survival, differentiation, and proliferation of macrophages. Mutations in CSF1 or CSF1R in mice and rats have pleiotropic effects on postnatal somatic growth. We tested the possible application of pig CSF1-Fc fusion protein as a therapy for low birth weight (LBW) at term, using a model based on maternal dexamethasone treatment in rats. Neonatal CSF1-Fc treatment did not alter somatic growth and did not increase the blood monocyte count. Instead, there was a substantial increase in the size of liver in both control and LBW rats, and the treatment greatly exacerbated lipid droplet accumulation seen in the dexamethasone LBW model. These effects were reversed upon cessation of treatment. Transcriptional profiling of the livers supported histochemical evidence of a large increase in macrophages with a resident Kupffer cell phenotype and revealed increased expression of many genes implicated in lipid droplet formation. There was no further increase in hepatocyte proliferation over the already high rates in neonatal liver. In conclusion, treatment of neonatal rats with CSF1-Fc caused an increase in liver size and hepatic lipid accumulation, due to Kupffer cell expansion and/or activation rather than hepatocyte proliferation. Increased liver macrophage numbers and expression of endocytic receptors could mitigate defective clearance functions in neonates. NEW & NOTEWORTHY This study is based on extensive studies in mice and pigs of the role of CSF1/CSF1R in macrophage development and postnatal growth. We extended the study to neonatal rats as a possible therapy for low birth weight. Unlike our previous studies in mice and pigs, there was no increase in hepatocyte proliferation and no increase in monocyte numbers. Instead, neonatal rats treated with CSF1 displayed reversible hepatic steatosis and Kupffer cell expansion.

A Csf1r-EGFP Transgene Provides a Novel Marker for Monocyte Subsets in Sheep.

Expression of Csf1r in adults is restricted to cells of the macrophage lineage. Transgenic reporters based upon the Csf1r locus require inclusion of the highly conserved Fms-intronic regulatory element for expression. We have created Csf1r-EGFP transgenic sheep via lentiviral transgenesis of a construct containing elements of the mouse Fms-intronic regulatory element and Csf1r promoter. Committed bone marrow macrophage precursors and blood monocytes express EGFP in these animals. Sheep monocytes were divided into three populations, similar to classical, intermediate, and nonclassical monocytes in humans, based upon CD14 and CD16 expression. All expressed EGFP, with increased levels in the nonclassical subset. Because Csf1r expression coincides with the earliest commitment to the macrophage lineage, Csf1r-EGFP bone marrow provides a tool for studying the earliest events in myelopoiesis using the sheep as a model.

Macrophage colony-stimulating factor (CSF1) controls monocyte production and maturation and the steady-state size of the liver in pigs.

Macrophage colony-stimulating factor (CSF1) is an essential growth and differentiation factor for cells of the macrophage lineage. To explore the role of CSF1 in steady-state control of monocyte production and differentiation and tissue repair, we previously developed a bioactive protein with a longer half-life in circulation by fusing pig CSF1 with the Fc region of pig IgG1a. CSF1-Fc administration to pigs expanded progenitor pools in the marrow and selectively increased monocyte numbers and their expression of the maturation marker CD163. There was a rapid increase in the size of the liver, and extensive proliferation of hepatocytes associated with increased macrophage infiltration. Despite the large influx of macrophages, there was no evidence of liver injury and no increase in circulating liver enzymes. Microarray expression profiling of livers identified increased expression of macrophage markers, i.e., cytokines such as TNF, IL1, and IL6 known to influence hepatocyte proliferation, alongside cell cycle genes. The analysis also revealed selective enrichment of genes associated with portal, as opposed to centrilobular regions, as seen in hepatic regeneration. Combined with earlier data from the mouse, this study supports the existence of a CSF1-dependent feedback loop, linking macrophages of the liver with bone marrow and blood monocytes, to mediate homeostatic control of the size of the liver. The results also provide evidence of safety and efficacy for possible clinical applications of CSF1-Fc.

Contribution of mammary epithelial cells to the immune response during early stages of a bacterial infection to Staphylococcus aureus.

To differentiate between the contribution of mammary epithelial cells (MEC) and infiltrating immune cells to gene expression profiles of mammary tissue during early stage mastitis, we investigated in goats the in vivo transcriptional response of MEC to an experimental intra mammary infection (IMI) with Staphylococcus aureus, using a non-invasive RNA sampling method from milk fat globules (MFG). Microarrays were used to record gene expression patterns during the first 24 hours post-infection (hpi). This approach was combined with laser capture microdissection of MEC from frozen slides of mammary tissue to analyze some relevant genes at 30 hpi. During the early stages post-inoculation, MEC play an important role in the recruitment and activation of inflammatory cells through the IL-8 signalling pathway and initiate a sharp induction of innate immune genes predominantly associated with the pro-inflammatory response. At 30 hpi, MEC express genes encoding different acute phase proteins, including SAA3, SERPINA1 and PTX3 and factors, such as S100A12, that contribute directly to fighting the infection. No significant change in the expression of genes encoding caseins was observed until 24 hpi, thus validating our experimental model to study early stages of infection before the occurrence of tissue damage, since the milk synthesis function is still operative. This is to our knowledge the first report showing in vivo, in goats, how MEC orchestrate the innate immune response to an IMI challenge with S. aureus. Moreover, the non-invasive sampling method of mammary representative RNA from MFG provides a valuable tool to easily follow the dynamics of gene expression in MEC to search for sensitive biomarkers in milk for early detection of mastitis and therefore, to successfully improve the treatment and thus animal welfare.

Comparative transcriptional analysis identifies genes associated with the attenuation of Theileria parva infected cells after long-term in vitro culture.

Autologous administration of attenuated Theileria parva-infected cells induces immunity to T. parva in cattle. The mechanism of attenuation, however, is largely unknown. Here, we used RNA sequencing of pathogenic and attenuated T. parva-infected T-cells to elucidate the transcriptional changes underpinning attenuation. We observed differential expression of several host genes, including TRAIL, PD-1, TGF-ß and granzymes that are known to regulate inflammation and proliferation of infected cells. Importantly, many genes linked with the attenuation of the related T. annulata-infected cells were not dysregulated in this study. Furthermore, known T. parva antigens were not dysregulated in attenuated relative to pathogenic cells, indicating that attenuation is not due to enhanced immunogenicity. Overall this study suggests that attenuation is driven by a decrease in proliferation and restoration of the inflammatory profile of T. parva-infected cells. Additionally, it provides a foundation for future mechanistic studies of the attenuation phenotype in Theileria-infected cells.

Phenotypic and spatial heterogeneity of brain myeloid cells after stroke is associated with cell ontogeny, tissue damage, and brain connectivity.

Acute stroke triggers extensive changes to myeloid immune cell populations in the brain that may be targets for limiting brain damage and enhancing repair. Immunomodulatory approaches will be most effective with precise manipulation of discrete myeloid cell phenotypes in time and space. Here, we investigate how stroke alters mononuclear myeloid cell composition and phenotypes at single-cell resolution and key spatial patterns. Our results show that multiple reactive microglial states and monocyte-derived populations contribute to an extensive myeloid cell repertoire in post-stroke brains. We identify important overlaps and distinctions among different cell types/states that involve ontogeny- and spatial-related properties. Notably, brain connectivity with infarcted tissue underpins the pattern of local and remote altered cell accumulation and reactivity. Our discoveries suggest a global but anatomically governed brain myeloid cell response to stroke that comprises diverse phenotypes arising through intrinsic cell ontogeny factors interacting with exposure to spatially organized brain damage and neuro-axonal cues.

Cystatin F (Cst7) drives sex-dependent changes in microglia in an amyloid-driven model of Alzheimer's disease.

Microglial endolysosomal (dys)function is strongly implicated in neurodegenerative disease. Transcriptomic studies show that a microglial state characterised by a set of genes involved in endolysosomal function is induced in both mouse Alzheimer's disease (AD) models and human AD brain, and that the emergence of this state is emphasised in females. Cst7 (encoding cystatin F) is among the most highly upregulated genes in these microglia. However, despite such striking and robust upregulation, the function of Cst7 in neurodegenerative disease is not understood. Here, we crossed Cst7-/- mice with the AppNL-G-F mouse to test the role of Cst7 in a model of amyloid-driven AD. Surprisingly, we found that Cst7 plays a sexually dimorphic role regulating microglia in this model. In females, Cst7-/-AppNL-G-F microglia had greater endolysosomal gene expression, lysosomal burden, and amyloid beta (Aß) burden in vivo and were more phagocytic in vitro. However, in males, Cst7-/-AppNL-G-F microglia were less inflammatory and had a reduction in lysosomal burden but had no change in Aß burden. Overall, our study reveals functional roles for one of the most commonly upregulated genes in microglia across disease models, and the sex-specific profiles of Cst7-/--altered microglial disease phenotypes. More broadly, the findings raise important implications for AD including crucial questions on sexual dimorphism in neurodegenerative disease and the interplay between endolysosomal and inflammatory pathways in AD pathology.

Use of quantitative real-time PCR to determine the local inflammatory response in the intestinal mucosa and muscularis of horses undergoing small intestinal resection.

BACKGROUND: Studies in rodents and humans have demonstrated that intestinal manipulation or surgical trauma initiates an inflammatory response in the intestine which results in leucocyte recruitment to the muscularis externa causing smooth muscle dysfunction. OBJECTIVES: To examine the intestinal inflammatory response in horses undergoing colic surgery by measuring relative differential gene expression in intestinal tissues harvested from surgical colic cases and control horses. STUDY DESIGN: Prospective case-control study. METHODS: Mucosa and muscularis externa were harvested from healthy margins of resected small intestine from horses undergoing colic surgery (n = 12) and from intestine derived from control horses euthanised for reasons unrelated to the gastrointestinal tract (n = 6). Tissue was analysed for genes encoding proteins involved in the inflammatory response: interleukin (IL) 6 and IL1ß, C-C motif chemokine ligand 2 (CCL2), tumour necrosis factor (TNF), prostaglandin-endoperoxide synthase 2 (PTGS2) and indoleamine 2,3-dioxygenase (IDO1). Relative expression of these genes was compared between the two groups. Further analysis was applied to the colic cases to determine whether the magnitude of relative gene expression was associated with the subsequent development of post-operative reflux (POR). RESULTS: Samples obtained from colic cases had increased relative expression of IL1ß, IL6, CCL2 and TNF in the mucosa and muscularis externa when compared with the control group. There was no difference in relative gene expression between proximal and distal resection margins and no association between duration of colic, age, resection length, short-term survival and the presence of pre-operative reflux and the relative expression of the genes of interest. Horses that developed POR had significantly greater relative gene expression of TNF in the mucosa compared with horses that did not develop POR. MAIN LIMITATIONS: Small sample size per group and variation within the colic cases. CONCLUSIONS: These preliminary data support an upregulation of inflammatory genes in the intestine of horses undergoing colic surgery.

CRISPR-Cas9 Editing of Human Histone Deubiquitinase Gene USP16 in Human Monocytic Leukemia Cell Line THP-1.

USP16 is a histone deubiquitinase which facilitates G2/M transition during the cell cycle, regulates DNA damage repair and contributes to inducible gene expression. We mutated the USP16 gene in a high differentiation clone of the acute monocytic leukemia cell line THP-1 using the CRISPR-Cas9 system and generated four homozygous knockout clones. All were able to proliferate and to differentiate in response to phorbol ester (PMA) treatment. One line was highly proliferative prior to PMA treatment and shut down proliferation upon differentiation, like wild type. Three clones showed sustained expression of the progenitor cell marker MYB, indicating that differentiation had not completely blocked proliferation in these clones. Network analysis of transcriptomic differences among wild type, heterozygotes and homozygotes showed clusters of genes that were up- or down-regulated after differentiation in all cell lines. Prior to PMA treatment, the homozygous clones had lower levels than wild type of genes relating to metabolism and mitochondria, including SRPRB, encoding an interaction partner of USP16. There was also apparent loss of interferon signaling. In contrast, a number of genes were up-regulated in the homozygous cells compared to wild type at baseline, including other deubiquitinases (USP12, BAP1, and MYSM1). However, three homozygotes failed to fully induce USP3 during differentiation. Other network clusters showed effects prior to or after differentiation in the homozygous clones. Thus the removal of USP16 affected the transcriptome of the cells, although all these lines were able to survive, which suggests that the functions attributed to USP16 may be redundant. Our analysis indicates that the leukemic line can adapt to the extreme selection pressure applied by the loss of USP16, and the harsh conditions of the gene editing and selection protocol, through different compensatory pathways. Similar selection pressures occur during the evolution of a cancer in vivo, and our results can be seen as a case study in leukemic cell adaptation. USP16 has been considered a target for cancer chemotherapy, but our results suggest that treatment would select for escape mutants that are resistant to USP16 inhibitors.

Transcriptomic Analysis of Rat Macrophages.

The laboratory rat is widely used as a model for human diseases. Many of these diseases involve monocytes and tissue macrophages in different states of activation. Whilst methods for in vitro differentiation of mouse macrophages from embryonic stem cells (ESC) and bone marrow (BM) are well established, these are lacking for the rat. The gene expression profiles of rat macrophages have also not been characterised to the same extent as mouse. We have established the methodology for production of rat ESC-derived macrophages and compared their gene expression profiles to macrophages obtained from the lung and peritoneal cavity and those differentiated from BM and blood monocytes. We determined the gene signature of Kupffer cells in the liver using rats deficient in macrophage colony stimulating factor receptor (CSF1R). We also examined the response of BM-derived macrophages to lipopolysaccharide (LPS). The results indicate that many, but not all, tissue-specific adaptations observed in mice are conserved in the rat. Importantly, we show that unlike mice, rat macrophages express the CSF1R ligand, colony stimulating factor 1 (CSF1).

The Transcriptional Network That Controls Growth Arrest and Macrophage Differentiation in the Human Myeloid Leukemia Cell Line THP-1.

The response of the human acute myeloid leukemia cell line THP-1 to phorbol esters has been widely studied to test candidate leukemia therapies and as a model of cell cycle arrest and monocyte-macrophage differentiation. Here we have employed Cap Analysis of Gene Expression (CAGE) to analyze a dense time course of transcriptional regulation in THP-1 cells treated with phorbol myristate acetate (PMA) over 96 h. PMA treatment greatly reduced the numbers of cells entering S phase and also blocked cells exiting G2/M. The PMA-treated cells became adherent and expression of mature macrophage-specific genes increased progressively over the duration of the time course. Within 1-2 h PMA induced known targets of tumor protein p53 (TP53), notably CDKN1A, followed by gradual down-regulation of cell-cycle associated genes. Also within the first 2 h, PMA induced immediate early genes including transcription factor genes encoding proteins implicated in macrophage differentiation (EGR2, JUN, MAFB) and down-regulated genes for transcription factors involved in immature myeloid cell proliferation (MYB, IRF8, GFI1). The dense time course revealed that the response to PMA was not linear and progressive. Rather, network-based clustering of the time course data highlighted a sequential cascade of transient up- and down-regulated expression of genes encoding feedback regulators, as well as transcription factors associated with macrophage differentiation and their inferred target genes. CAGE also identified known and candidate novel enhancers expressed in THP-1 cells and many novel inducible genes that currently lack functional annotation and/or had no previously known function in macrophages. The time course is available on the ZENBU platform allowing comparison to FANTOM4 and FANTOM5 data.

Chromosome-level assembly of the water buffalo genome surpasses human and goat genomes in sequence contiguity.

Rapid innovation in sequencing technologies and improvement in assembly algorithms have enabled the creation of highly contiguous mammalian genomes. Here we report a chromosome-level assembly of the water buffalo (Bubalus bubalis) genome using single-molecule sequencing and chromatin conformation capture data. PacBio Sequel reads, with a mean length of 11.5 kb, helped to resolve repetitive elements and generate sequence contiguity. All five B. bubalis sub-metacentric chromosomes were correctly scaffolded with centromeres spanned. Although the index animal was partly inbred, 58% of the genome was haplotype-phased by FALCON-Unzip. This new reference genome improves the contig N50 of the previous short-read based buffalo assembly more than a thousand-fold and contains only 383 gaps. It surpasses the human and goat references in sequence contiguity and facilitates the annotation of hard to assemble gene clusters such as the major histocompatibility complex (MHC).

A Gene Expression Atlas of the Domestic Water Buffalo (Bubalus bubalis).

The domestic water buffalo (Bubalus bubalis) makes a major contribution to the global agricultural economy in the form of milk, meat, hides, and draught power. The global water buffalo population is predominantly found in Asia, and per head of population more people depend upon the buffalo than on any other livestock species. Despite its agricultural importance, there are comparatively fewer genomic and transcriptomic resources available for buffalo than for other livestock species. We have generated a large-scale gene expression atlas covering multiple tissue and cell types from all major organ systems collected from three breeds of riverine water buffalo (Mediterranean, Pandharpuri and Bhadawari) and used the network analysis tool Graphia Professional to identify clusters of genes with similar expression profiles. Alongside similar data, we and others have generated for ruminants as part of the Functional Annotation of Animal Genomes Consortium; this comprehensive transcriptome supports functional annotation and comparative analysis of the water buffalo genome.

Deletion of a Csf1r enhancer selectively impacts CSF1R expression and development of tissue macrophage populations.

The proliferation, differentiation and survival of mononuclear phagocytes depend on signals from the receptor for macrophage colony-stimulating factor, CSF1R. The mammalian Csf1r locus contains a highly conserved super-enhancer, the fms-intronic regulatory element (FIRE). Here we show that genomic deletion of FIRE in mice selectively impacts CSF1R expression and tissue macrophage development in specific tissues. Deletion of FIRE ablates macrophage development from murine embryonic stem cells. Csf1rΔFIRE/ΔFIRE mice lack macrophages in the embryo, brain microglia and resident macrophages in the skin, kidney, heart and peritoneum. The homeostasis of other macrophage populations and monocytes is unaffected, but monocytes and their progenitors in bone marrow lack surface CSF1R. Finally, Csf1rΔFIRE/ΔFIRE mice are healthy and fertile without the growth, neurological or developmental abnormalities reported in Csf1r-/- rodents. Csf1rΔFIRE/ΔFIRE mice thus provide a model to explore the homeostatic, physiological and immunological functions of tissue-specific macrophage populations in adult animals.