Role of Monocytes and Intestinal Macrophages in Crohn's Disease and Ulcerative Colitis

Monocytes and macrophages are part of the body's first line of defence, eliminating pathogens by phagocytosis or by releasing a broad array of inflammatory mediators, such as cytokines, chemokines, and proteases. In humans, 3 subsets of monocytes are described in blood with seemingly different functions, the classical (CD14 + + CD16 -) monocytes, the intermediate (CD14 + + CD16 +) monocytes, and the nonclassical (CD14 + CD16 + +) monocytes. In the intestine, macrophages can be divided into resident and inflammatory macrophages that are distinguished by low and high expression of CD14, respectively. However, the roles and function of the 3 monocyte subsets in health and disease are not fully understood. In this review, we describe what is known about the origin of human intestinal macrophages and their blood monocytic counterparts and many of their numerous distinct mechanisms influencing the intestinal immune system

A Single-Cell Gene-Expression Profile Reveals Inter-Cellular Heterogeneity within Human Monocyte Subsets.

Human monocytes are a heterogeneous cell population classified into three different subsets: Classical CD14++CD16-, intermediate CD14++CD16+, and non-classical CD14+CD16++ monocytes. These subsets are distinguished by their differential expression of CD14 and CD16, and unique gene expression profile. So far, the variation in inter-cellular gene expression within the monocyte subsets is largely unknown. In this study, the cellular variation within each human monocyte subset from a single healthy donor was described by using a novel single-cell PCR gene-expression analysis tool. We investigated 86 different genes mainly encoding cell surface markers, and proteins involved in immune regulation. Within the three human monocyte subsets, our descriptive findings show multimodal expression of key immune response genes, such as CD40, NFⱪB1, RELA, TLR4, TLR8 and TLR9. Furthermore, we discovered one subgroup of cells within the classical monocytes, which showed alterations of 22 genes e.g. IRF8, CD40, CSF1R, NFⱪB1, RELA and TNF. Additionally one subgroup within the intermediate and non-classical monocytes also displayed distinct gene signatures by altered expression of 8 and 6 genes, respectively. Hence the three monocyte subsets can be further subdivided according to activation status and differentiation, independently of the traditional classification based on cell surface markers. Demonstrating the use and the ability to discover cell heterogeneity within defined populations of human monocytes is of great importance, and can be useful in unravelling inter-cellular variation in leukocyte populations, identifying subpopulations involved in disease pathogenesis and help tailor new therapies

Genes of interest investigated in the three monocyte subsets.

<p>Protein function adapted from <a href="http://GeneCards.org" target="_blank">GeneCards.org</a></p><p>Genes of interest investigated in the three monocyte subsets.</p

Relative Log2 transformed gene expression levels of the three subsets and statistical significance the subsets in between.

<p>C = Classical, I = Intermediate, NC = Non-Classical</p><p>Relative Log2 transformed gene expression levels of the three subsets and statistical significance the subsets in between.</p

Expression level of genes deviating in identified subgroups.

<p>Subgroups of cells were identified based on the PCA of single-cell PCR gene expression analysis data. One subgroup among the classical monocytes, intermediate monocytes and non-classical monocytes, and co-expression of genes within the subgroups were assessed using a student T test (<i>P <</i> 0.05). A) Bar graph demonstrating the differentially expressed genes by the subgroup within the classical monocyte subset identified on the PCA score plot. The subgroup is marked by filled red circles in the PCA score plot. B) Bar graph demonstrating the differentially expressed genes by the subgroup within the intermediate monocyte subset identified on the PCA score plot. The subgroup is marked by filled green triangles in the PCA score plot. C) Bar graph demonstrating the differentially expressed genes by the subgroup within the non-classical monocyte subset identified on the PCA score plot. The subgroup is marked by filled blue pluses in the PCA score plot.</p

Multimodal variation in expression levels across the three monocyte subsets.

<p>Violin plot demonstrating multimodal variation in gene expression levels of the 85 genes examined in the monocyte subsets. The classical monocytes, intermediate monocytes and non-classical monocytes are indicated in the figure by red, green and blue, respectively. The data depict the multimodal expression levels of the genes listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0144351#pone.0144351.t004" target="_blank">Table 4</a> calculated by using the Hartigans dip test (<i>P <</i> 0.05).</p

Single-cell gene expression analysis on human monocytes.

<p>Human monocytes were single-cell sorted according to the expression of the cell surface markers CD14 and CD16 and gene expression on single cells was assessed. A) Representative plot of flow cytometry analysis demonstrating the monocyte subset gating strategy, used to single-cell sort monocytes from the three classified monocyte subsets (<i>n =</i> 1). Classical (CD14⁺⁺CD16^-) monocytes, intermediate (CD14⁺⁺CD16⁺) monocytes and non-classical (CD14⁺⁺CD16⁺) monocytes are depicted in the upper left quadrant, upper right quadrant and lower right quadrant, respectively. B) Principal component analyses (PCA) of single-cell PCR gene expression analysis data showing genetic clustering of the three monocyte subsets. The PCA plot confirmed the classification of the three human monocyte subsets done by flow cytometry, visualized by gene families. Each dot represents a single cell. C) Heatmap of gene expression values for PCA showing hierarchical clustering of single-cell PCR gene expression data from the three human monocyte sub-populations. The analysis revealed cellular heterogeneity by distinct gene signatures. Red circles = classical monocytes (<i>n</i> = 94 cells), green triangles = intermediate monocytes (<i>n</i> = 92 cells), and blue pluses = non-classical monocytes (<i>n</i> = 80 cells).</p

Multimodal expression in the three monocyte subsets.

<p><b>Bold</b> = Multimodal expression <i>P</i> <0.05, <i>Italic</i> = Unimodal expression</p><p>Multimodal expression in the three monocyte subsets.</p