CD64 distinguishes macrophages from dendritic cells in the gut and reveals the Th1‐inducing role of mesenteric lymph node macrophages during colitis

Abstract

Dendritic cells (DCs) and monocyte-derived macrophages (M s) are key components of intestinal immunity. However, the lack of surface markers differentiating M s from DCs has hampered understanding of their respective functions. Here, we demonstrate that, using CD64 expression, M s can be distinguished from DCs in the intestine of both mice and humans. On that basis, we revisit the phenotype of intestinal DCs in the absence of contaminating M s and we delineate a developmental pathway in the healthy intestine that leads from newly extravasated Ly-6C hi monocytes to intestinal M s. We determine how inflammation impacts this pathway and show that T cell-mediated colitis is associated with massive recruitment of monocytes to the intestine and the mesenteric lymph node (MLN). There, these monocytes differentiate into inflammatory M s endowed with phagocytic activity and the ability to produce inducible nitric oxide synthase. Eur. J. Immunol. 2012. 42: 3150-3166 HIGHLIGHTS 3151 IntroductionThe intestinal lamina propria (LP) contains cells that express high levels of CX 3 CR1, the receptor for the fractalkine chemokine [1,2]. Based on their monocytic origin and on their inability to migrate to the mesenteric lymph nodes (MLNs) such CX 3 CR1 hi cells have been defined as macrophages (M s) [1][2][3]. CX 3 CR1 hi M s contribute to the intestinal LP homeostasis through the production of anti-inflammatory cytokines and the clearance of commensal bacteria that breach the epithelial barrier [4]. In contrast, during intestinal inflammation, microenvironmental signals promote the differentiation of extravasated monocytes into proinflammatory M s with the ability to produce interleukin (IL)-12, IL-23, tumor necrosis (TNF)-α and inducible nitric oxide synthase (iNOS) [5][6][7]. However, little is known about the developmental trajectories that lead extravasated monocytes to either antior proinflammatory intestinal M s. This is primarily due to the fact that a surface marker permitting unequivocal identification of M s within the intestine and their distinction from dendritic cells (DCs) is lacking. The interstitial DCs (Int-DCs) present throughout the LP derive from blood precursors known as pre-DCs [2]. Under steady-state conditions, the Int-DCs found in the intestinal LP induce oral tolerance by carrying antigens originating from food or from harmless bacteria to the MLNs [8,9]. The CD103 + Int-DCs found in the steady-state LP have the selective ability to express aldehyde dehydrogenase (ALDH) and thereby produce retinoic acid (RA). As a result, upon migration to MLNs they trigger the differentiation of naive CD4 + T cells specific for food and microbiota antigens into induced Foxp3 + regulatory T (iTreg) cells [10][11][12][13]. In contrast, the Int-DCs that develop in inflamed LP upon exposure to pathogens lose their capacity to generate iTreg cells and, upon migration to the MLNs, trigger the differentiation of naive, antigen-responsive CD4 + T cells into T helper type 1 (Th1) cells that are specific for the invading patho...

Flow Cytometric Analysis of Macrophages and Dendritic Cell Subsets in the Mouse Lung

“…1 DCs in the mouse gut and muscle (31,32), and together with proto-oncogene tyrosine-protein kinase MER (MerTK) and CD14, is one of the most specific macrophage markers (18). We found that a combination of CD64 with CD24 and MHC II allowed not only for the separation of CD11b…”

Crucial role of plasmacytoid dendritic cells in the development of acute colitis through the regulation of intestinal inflammation

“…Fluorescence staining was analyzed with a FACSVerse flow cytometer (BD Biosciences) and FlowJo software (Tree star, Ashland, OR). In some experiments, flow cytometry analysis for the identification of leukocytes in the intestinal tissues was performed according to the previous reports 27,28,44 with some modifications.…”

Maturation Phenotype of Peripheral Blood Monocyte/Macrophage After Stimulation with Lipopolysaccharides in Irritable Bowel Syndrome

“…28 Thus, analyzing a model of MNCs stimulated with LPS is a logical focus for speculating the possible response of these cells to bacterial stimuli and their comparison between IBS patients and controls. Likewise, it is important to point out that in the 2 studies using a murine model, Bain et al 29 and Tamoutounour et al 16 proposed that the conditions of the intestine lead to the development of the macrophage in a cascade-like fashion. This is a linear concept in which complete macrophage development up to a tolerant state is possible in a normal lamina propria, whereas an inflammatory state mainly promotes the maturation of proinflammatory response macrophages.…”

“…15 This monocyte recruitment provides a constant intestinal replenishment of macrophage populations that gradually mature in a cascade-like manner until they reach the level of resident macrophages. 16,17 Our hypothesis was that patients with IBS may show an alteration in the maturation cascade of monocytes into macrophages in response to bacterial components in charge of triggering this disorder. Therefore our aim was to analyze the maturation phenotype of monocytes/macrophages (CD14+) from the peripheral blood of patients with IBS and controls, by in vitro exposing CD14+ cells to Escherichia coli-derived lipopolysaccharides (LPS).…”