Transcriptomic Analysis of Human Polarized Macrophages: More than One Role of Alternative Activation?

Abstract

BackgroundMacrophages are a heterogeneous cell population which in response to the cytokine milieu polarize in either classically activated macrophages (M1) or alternatively activated macrophages (M2). This plasticity makes macrophages essential in regulating inflammation, immune response and tissue remodeling and a novel therapeutic target in inflammatory diseases such as atherosclerosis. The aim of the study was to describe the transcriptomic profiles of differently polarized human macrophages to generate new hypotheses on the biological function of the different macrophage subtypes.Methods and ResultsPolarization of circulating monocytes/macrophages of blood donors was induced in vitro by IFN-γ and LPS (M1), by IL-4 (M2a), and by IL-10 (M2c). Unstimulated cells (RM) served as time controls. Gene expression profile of M1, M2a, M2c and RM was assessed at 6, 12 and 24h after polarization with Whole Human Genome Agilent Microarray technique. When compared to RM, M1 significantly upregulated pathways involved in immunity and inflammation, whereas M2a did the opposite. Conversely, decreased and increased expression of mitochondrial metabolism, consistent with insulin resistant and insulin sensitive patterns, was seen in M1 and M2a, respectively. The time sequence in the expression of some pathways appeared to have some specific bearing on M1 function. Finally, canonical and non-canonical Wnt genes and gene groups, promoting inflammation and tissue remodeling, were upregulated in M2a compared to RM.ConclusionOur data in in vitro polarized human macrophages: 1. confirm and extend known inflammatory and anti-inflammatory gene expression patterns; 2. demonstrate changes in mitochondrial metabolism associated to insulin resistance and insulin sensitivity in M1 and M2a, respectively; 3. highlight the potential relevance of gene expression timing in M1 function; 4. unveil enhanced expression of Wnt pathways in M2a suggesting a potential dual (pro-inflammatory and anti-inflammatory) role of M2a in inflammatory diseases.

Glioblastoma-infiltrated innate immune cells resemble M0 macrophage phenotype

“…M1 polarization of macrophages was associated with the most dramatic changes in gene expression, whereas nonpolarized M0 and M2a- and M2c-polarized macrophages revealed subtle genetic differences and possessed more closely aligned expression patterns, consistent with prior reports of microarray analyses (36, 37). Among the genes that most clearly distinguished M1 macrophages were the transcription factors IRF1 and STAT1 ; the receptors CCR7 , CD80 , IL15RA , and MET ; a variety of cytokines, such as CXCL9 , CXCL10 , and CXCL11 ; the enzyme PTGS2 ; the Ca channel KCNN2 ; and the lncRNAs CCDC26 and RP11-419k12.1 .…”

“…We also make the observation that oxidative phosphorylation is upregulated in glioblastoma blood monocytes relative to normal donors. Oxidative phosphorylation has been previously associated with the M0 and M2 phenotypes (37, 50) in contrast to glycolysis utilization in M1 cells. Whether these changes in the metabolic phenotype can occur in the circulation has been previously debated (51) with the presumption being that this occurs only at the site of inflammation.…”

Tumor-associated macrophages in cancers

“…M1 macrophages highly express IL-12, IL-23, major histocompatibility complex (MHC) and B7 family members. The main function of M1 macrophages includes the promotion of antigen presentation and Th1 activation [9]. M1 macrophages can secrete proinflammatory cytokines and immune activation factors to promote inflammation and antitumor function [8].…”

Transcriptome analysis of IL-10-stimulated (M2c) macrophages by next-generation sequencing

“…While Williams et al (2002) found that just 19 genes were upregulated in M2c macrophages after 3 h of IL-10 stimulation, Derlindati et al (2015) reported that no genes were upregulated after 6, 12, and 24 h of polarization. Therefore, the purpose of this study was to identify a gene expression signature for M2c macrophages using whole transcriptome shotgun sequencing, or RNA-seq, a more sensitive method of gene expression analysis.…”