It remains unclear whether activated inflammatory macrophages can adopt features of tissue-resident macrophages, or what mechanisms might mediate such a phenotypic conversion. Here we show that vitamin A is required for the phenotypic conversion of interleukin 4 (IL-4)-activated monocyte-derived F4/80intCD206+PD-L2+MHCII+ macrophages into macrophages with a tissue-resident F4/80hiCD206−PD-L2−MHCII−UCP1+ phenotype in the peritoneal cavity of mice and during the formation of liver granulomas in mice infected with Schistosoma mansoni. The phenotypic conversion of F4/80intCD206+ macrophages into F4/80hiCD206− macrophages was associated with almost complete remodeling of the chromatin landscape, as well as alteration of the transcriptional profiles. Vitamin A–deficient mice infected with S. mansoni had disrupted liver granuloma architecture and increased mortality, which indicates that failure to convert macrophages from the F4/80intCD206+ phenotype to F4/80hiCD206− may lead to dysregulated inflammation during helminth infection.
This is a preview of subscription content
Subscribe to Nature+
Get immediate online access to the entire Nature family of 50+ journals
Subscribe to Journal
Get full journal access for 1 year
only $4.92 per issue
All prices are NET prices.
VAT will be added later in the checkout.
Tax calculation will be finalised during checkout.
Get time limited or full article access on ReadCube.
All prices are NET prices.
Sieweke, M.H. & Allen, J.E. Beyond stem cells: self-renewal of differentiated macrophages. Science 342, 1242974 (2013).
Zigmond, E. & Jung, S. Intestinal macrophages: well educated exceptions from the rule. Trends Immunol. 34, 162–168 (2013).
Ingersoll, M.A., Platt, A.M., Potteaux, S. & Randolph, G.J. Monocyte trafficking in acute and chronic inflammation. Trends Immunol. 32, 470–477 (2011).
Shi, C. & Pamer, E.G. Monocyte recruitment during infection and inflammation. Nat. Rev. Immunol. 11, 762–774 (2011).
Gosselin, D. et al. Environment drives selection and function of enhancers controlling tissue-specific macrophage identities. Cell 159, 1327–1340 (2014).
Lavin, Y. et al. Tissue-resident macrophage enhancer landscapes are shaped by the local microenvironment. Cell 159, 1312–1326 (2014).
Scott, C.L. et al. Bone marrow–derived monocytes give rise to self-renewing and fully differentiated Kupffer cells. Nat. Commun. 7, 10321 (2016).
van de Laar, L. et al. Yolk sac macrophages, fetal liver, and adult monocytes can colonize an empty niche and develop into functional tissue-resident macrophages. Immunity 44, 755–768 (2016).
Hams, E., Aviello, G. & Fallon, P.G. The schistosoma granuloma: friend or foe? Front. Immunol. 4, 89 (2013).
Pearce, E.J. & MacDonald, A.S. The immunobiology of schistosomiasis. Nat. Rev. Immunol. 2, 499–511 (2002).
Wilson, M.S. et al. Immunopathology of schistosomiasis. Immunol. Cell Biol. 85, 148–154 (2007).
Murray, P.J. et al. Macrophage activation and polarization: nomenclature and experimental guidelines. Immunity 41, 14–20 (2014).
Wynn, T.A. & Vannella, K.M. Macrophages in tissue repair, regeneration, and fibrosis. Immunity 44, 450–462 (2016).
Herbert, D.R. et al. Alternative macrophage activation is essential for survival during schistosomiasis and downmodulates T helper 1 responses and immunopathology. Immunity 20, 623–635 (2004).
Girgis, N.M. et al. Ly6C(high) monocytes become alternatively activated macrophages in schistosome granulomas with help from CD4+ cells. PLoS Pathog. 10, e1004080 (2014).
Nascimento, M. et al. Ly6Chi monocyte recruitment is responsible for Th2 associated host-protective macrophage accumulation in liver inflammation due to schistosomiasis. PLoS Pathog. 10, e1004282 (2014).
Jenkins, S.J. et al. Local macrophage proliferation, rather than recruitment from the blood, is a signature of TH2 inflammation. Science 332, 1284–1288 (2011).
Jenkins, S.J. et al. IL-4 directly signals tissue-resident macrophages to proliferate beyond homeostatic levels controlled by CSF-1. J. Exp. Med. 210, 2477–2491 (2013).
Gundra, U.M. et al. Alternatively activated macrophages derived from monocytes and tissue macrophages are phenotypically and functionally distinct. Blood 123, e110–e122 (2014).
Pesce, J.T. et al. Arginase-1-expressing macrophages suppress Th2 cytokine-driven inflammation and fibrosis. PLoS Pathog. 5, e1000371 (2009).
Amit, I., Winter, D.R. & Jung, S. The role of the local environment and epigenetics in shaping macrophage identity and their effect on tissue homeostasis. Nat. Immunol. 17, 18–25 (2016).
Perdiguero, E.G. & Geissmann, F. The development and maintenance of resident macrophages. Nat. Immunol. 17, 2–8 (2016).
Stephensen, C.B. Vitamin A, infection, and immune function. Annu. Rev. Nutr. 21, 167–192 (2001).
Hall, J.A., Grainger, J.R., Spencer, S.P. & Belkaid, Y. The role of retinoic acid in tolerance and immunity. Immunity 35, 13–22 (2011).
Okabe, Y. & Medzhitov, R. Tissue-specific signals control reversible program of localization and functional polarization of macrophages. Cell 157, 832–844 (2014).
Barth, M.W., Hendrzak, J.A., Melnicoff, M.J. & Morahan, P.S. Review of the macrophage disappearance reaction. J. Leukoc. Biol. 57, 361–367 (1995).
Gevers, D. et al. The Human Microbiome Project: a community resource for the healthy human microbiome. PLoS Biol. 10, e1001377 (2012).
Parkhurst, C.N. et al. Microglia promote learning-dependent synapse formation through brain-derived neurotrophic factor. Cell 155, 1596–1609 (2013).
Kuwata, T. et al. Vitamin A deficiency in mice causes a systemic expansion of myeloid cells. Blood 95, 3349–3356 (2000).
Snippert, H.J. et al. Intestinal crypt homeostasis results from neutral competition between symmetrically dividing Lgr5 stem cells. Cell 143, 134–144 (2010).
A complex cell. Nat. Immunol. 17, 1 (2016).
Gautier, E.L. et al. Gata6 regulates aspartoacylase expression in resident peritoneal macrophages and controls their survival. J. Exp. Med. 211, 1525–1531 (2014).
Rosas, M. et al. The transcription factor Gata6 links tissue macrophage phenotype and proliferative renewal. Science 344, 645–648 (2014).
Wang, J. & Kubes, P. A reservoir of mature cavity macrophages that can rapidly invade visceral organs to affect tissue repair. Cell 165, 668–678 (2016).
Gibbings, S.L. et al. Transcriptome analysis highlights the conserved difference between embryonic and postnatal-derived alveolar macrophages. Blood 126, 1357–1366 (2015).
van Furth, R. & Cohn, Z.A. The origin and kinetics of mononuclear phagocytes. J. Exp. Med. 128, 415–435 (1968).
Bain, C.C. et al. Long-lived self-renewing bone marrow–derived macrophages displace embryo-derived cells to inhabit adult serous cavities. Nat. Commun. 7, ncomms11852 (2016).
Kim, K.K. et al. MHC ||+ resident peritoneal and pleural macrophages rely on IRF4 for development from circulating monocytes. J. Exp. Med. 213, 1951–1959 (2016).
Broadhurst, M.J. et al. Upregulation of retinal dehydrogenase 2 in alternatively activated macrophages during retinoid-dependent type-2 immunity to helminth infection in mice. PLoS Pathog. 8, e1002883 (2012).
Joyce, K.L., Morgan, W., Greenberg, R. & Nair, M.G. Using eggs from Schistosoma mansoni as an in vivo model of helminth-induced lung inflammation. J. Vis. Exp. e3905 (2012).
Buenrostro, J.D., Giresi, P.G., Zaba, L.C., Chang, H.Y. & Greenleaf, W.J. Transposition of native chromatin for fast and sensitive epigenomic profiling of open chromatin, DNA-binding proteins and nucleosome position. Nat. Methods 10, 1213–1218 (2013).
Kim, D. et al. TopHat2: accurate alignment of transcriptomes in the presence of insertions, deletions and gene fusions. Genome Biol. 14, R36 (2013).
Anders, S., Pyl, P.T. & Huber, W. HTSeq—a Python framework to work with high-throughput sequencing data. Bioinformatics 31, 166–169 (2015).
Langmead, B. & Salzberg, S.L. Fast gapped-read alignment with Bowtie 2. Nat. Methods 9, 357–359 (2012).
McCarthy, M.T. & O'Callaghan, C.A. PeaKDEck: a kernel density estimator-based peak calling program for DNaseI-seq data. Bioinformatics 30, 1302–1304 (2014).
Stark, R. & Brown, G. DiffBind: Differential Binding Analysis of ChIP-Seq Peak Data (Univ. of Cambridge, 2011).
Love, M.I., Huber, W. & Anders, S. Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol. 15, 550 (2014).
Kaufman, L. & Rousseeuw, P.J. Finding Groups in Data: An Introduction to Cluster Analysis (Wiley, 1990).
Falcon, S. & Gentleman, R. Using GOstats to test gene lists for GO term association. Bioinformatics 23, 257–258 (2007).
We thank K. Cadwell and members of the K. Cadwell, J. Ernst and E. Fisher laboratories for their reading of this manuscript. We thank W. Gan and D. Littman (New York University Medical Center, Skirball Institute of Biomolecular Medicine, New York, New York, USA) for generously providing the Cx3cr1CreERT2-IRES-EYFP mice, and J. Collins (UT Southwestern, Dallas, Texas, USA) for providing S. mansoni eggs. We thank the NYUMC Genome Technology Core, NYUMC Flow Cytometry Core, NYUMC Microscopy Core and NYUMC Histopathology Core facilities for their assistance; these shared resources are partially supported by the Cancer Center (support grant P30CA016087) at the Laura and Isaac Perlmutter Cancer Center. This work was supported by the NIH (T32 microbiology training grant T32 AI007180 to U.M.G. and M.A.G.; T32 cardiology grant 104220 to U.M.G.), NIAID (grants AI093811 and AI094166 to P.L.), NIDDK (grant DK103788 to P.L.), a Ruth L. Kirschstein NRSA fellowship (F32AI102502 to N.M.G.), the Vilcek Foundation (U.M.G.) and AAI (M.S.T.). Biomphalaria glabrata snails were provided by the NIAID Schistosomiasis Resource Center of the Biomedical Research Institute (Rockville, Maryland, USA) through NIH–NIAID contract HHSN272201000005I for distribution through BEI Resources.
The authors declare no competing financial interests.
Integrated supplementary information
Supplementary Figure 1 Monocyte-derived inflammatory macrophages adopt a tissue-resident phenotype after long-term residency in the peritoneal cavity.
(a) Schematic of short-term transfer of monocyte-derived inflammatory macrophages from thioglycollate treated CD45.1 donor mice into CD45.2 recipient mice, rested for 24hrs and subsequently treated with IL-4c. (b) Quantification of mean fluorescence intensity values of PD-L2 and CD206 expression on donor and recipient macrophages. (c and d) Flow cytometric analysis of adoptive cell transfer of macrophages flow sorted from (c) thioglycollate-treated mice or (d) IL-4c treated mice and transferred into CD45.2 recipient mice and then treated with IL-4c 24hrs after transfer. Representative histograms display surface expression of F4/80, CD206, PD-L2 and MHCII in Donor CD45.1+ CD45.2− (Black line) and recipient CD45.1−, CD45.2+ (Grey shaded) CD11b+, F4/80+ macrophages. (e) IL-4c induced tissue resident M2 macrophages rapidly disappear after transfer into an inflammatory environment. CD11b+, F4/80+ macrophages sorted from CD45.1 donor mice treated with IL-4c were adoptively transferred into CD45.2 recipient mice, 24hrs later the recipient mice were treated with thioglycollate + IL-4c prior to analysis.
(a) Schematic of timecourse analysis of peritoneal macrophages 1 week (n=4), 4 weeks (n=4) or 8 weeks (n=4) after thioglycollate injection. Stacked bar graph showing the relative proportion of F4/80 and/or CD206 expression. (b) Representative FACS plots are shown displaying the frequency of CD45.1+ (blue) donor cells in CD45.2 (grey) recipient mice treated with or without IL-4c after 8 weeks of residency. Flow cytometry analysis of transferred cells shows acquisition of tissue resident phenotype by CD45.1 donor cells in the presence or absence of IL-4c treatment. IL-4c treatment increases the frequency of CD45.1+ F4/80+ cells. (c) Representative FACS plots showing the frequency of EdU+ cells in recipient and donor populations in response to IL-4c given after transfer and resting for 24hrs for short term residency or long term residency for 8 weeks. (d) Schematic of adoptive transfer of Thio-elicited monocyte-derived macrophages from Stat6−/− CD45.2 donor mice transferred into WT CD45.1 recipient mice rested for 24hrs and then treated with IL-4c. Histograms display expression of F4/80, CD206, PD-L2 and MHCII in donor CD45.2+ CD45.1− (Black) and recipient CD45.2−, CD45.1+ (Grey shaded) CD11b+ cells. (e) Representative FACS plots showing the frequency of EdU+ cells in WT recipient and Stat6−/− donor CD11b+ F4/80+ macrophages in response to IL-4c given after resting for 24hrs.
Supplementary Figure 3 Transcriptional and chromatin landscape reprogramming during macrophage conversion.
(a) Pairwise Pearson’s correlation analysis and (b) PCA of transcriptional profiles in AAMmono, AAMconv and AAMres. (c) Heatmap visualizing the expression values of 6 specific genes across the different populations of macrophages. (d) Top 10 GO terms enriched in the 3966 genes upregulated in AAMmono when compared to AAMconv (top) and top 10 GO terms enriched in the 675 genes upregulated in AAMconv when compared to AAMres (bottom). (e) Pairwise Pearson’s correlation analysis and (f) PCA of accessible chromatin regions in AAMmono, AAMconv and AAMres. On PCA plots, red circles represent AAMmono, orange squares represent AAMconv and blue triangles represent AAMres in PCA plots.
Supplementary Figure 4 Baseline disruption of peritoneal tissue-resident macrophages in vitamin A–deficient mice.
(a) Total number of cells collected from the peritoneal cavity of vitamin A deficient (Vit-ADEF) or control (Vit-ACON) mice via peritoneal lavage. (b,c) Total number of F4/80hi CD206− (b) or F4/80int CD206+ (c) cells in the peritoneal cavity in Vit-ADEF or Vit-ACON mice. **P < 0.01. Unpaired Students T-test.
Supplementary Figure 5 Expression of UCP1 in S. mansoni infection associated with EdU localization in mature liver granulomas.
(a) Transcript expression of Ucp1 in whole liver from mice infected with S. mansoni at 9 weeks and 12 weeks post infection. (b) Representative immunofluorescence images of S. mansoni-infected liver granulomas at different timepoints post infection stained with DAPI (blue) and Click-it EdU (red) taken from mice pulsed with EdU 3 hours prior to sacrifice. Eggs are outlined in white. (c) Slide-scanned, immunofluorescence image of S. mansoni-infected liver granulomas taken at 8 weeks post infection and stained with DAPI (blue), anti-UCP1 (green) and Click-it EdU (red) in vitamin A control mice pulsed with EdU 3 hours prior to sacrifice. Scale bars represent 50 microns or 500 microns as indicated.
Supplementary Figure 6 STAT6 regulates phenotypic conversion of peritoneal AAMmono cells after S. mansoni egg injection.
(a) Schematic of S. mansoni egg injection in Cx3cr1creERT2-IRESYFP/+ Rosa26stopfl-tdTomato/+ vitamin A deficient (Vit-ADEF) or control (Vit-ACON) mice. Representative flow cytometry plots of fate-mapped monocyte-derived macrophages in the lung after S. mansoni egg challenge in the lung. (b) Schematic of S. mansoni egg injection in WT:WT (n=5) or Stat6−/−:WT (n=5) mixed bone marrow chimeric mice whereby mice were sensitized with eggs via i.p. injection then challenged i.v. with eggs after 2 weeks. PECs and lung macrophages were analyzed after 8 days of rest and pulsed with EdU 3hrs prior to sacrifice. Representative flow cytometry plots display macrophage phenotypes from the peritoneal cavity and lung in mixed bone marrow chimeric mice treated with or without eggs. (c) Schematic of S. mansoni infection in mixed bone marrow chimera Stat6−/−:WT mice. Cumulative body weight during the infection reveals loss of weight after 5 weeks post-infection. Kaplan-Meyer survival curve displays rapid mortality of Stat6−/−:WT chimeric mice at 7 weeks post-infection.
About this article
Cite this article
Gundra, U., Girgis, N., Gonzalez, M. et al. Vitamin A mediates conversion of monocyte-derived macrophages into tissue-resident macrophages during alternative activation. Nat Immunol 18, 642–653 (2017). https://doi.org/10.1038/ni.3734
High-salt diet downregulates TREM2 expression and blunts efferocytosis of macrophages after acute ischemic stroke
Journal of Neuroinflammation (2021)
Hepatic macrophages in liver homeostasis and diseases-diversity, plasticity and therapeutic opportunities
Cellular & Molecular Immunology (2021)
Nature Communications (2021)
Recruited macrophages that colonize the post-inflammatory peritoneal niche convert into functionally divergent resident cells
Nature Communications (2021)
Cellular and Molecular Life Sciences (2021)