An in vitro test system for compounds that modulate human inflammatory macrophage polarization

Eur J Pharmacol. 2018 Aug 15:833:328-338. doi: 10.1016/j.ejphar.2018.06.017. Epub 2018 Jun 18.


Macrophages undergo activation by pathophysiological stimuli to pro-inflammatory and bactericidal, or wound-healing and anti-inflammatory phenotypes, termed M1 or M2, respectively. Dysregulation of the M1-M2 balance is often associated with inflammatory diseases. Therefore, mechanisms of macrophage polarization may reveal new drug targets. We profiled six compounds with claimed modulatory effects on macrophage polarization using peripheral blood monocyte-derived macrophages. Based on the distinct mRNA or protein expression in macrophages stimulated either with M1 [lipopolysaccharide (LPS) + interferon-γ, IFNγ] or M2 interleukin-4 (IL-4) stimuli, we selected a combination of M1 (IL1β, tumor necrosis factor-α,TNFα, CC chemokine receptor 7, CCR7 and CD80) and M2 (chemokine (C-C motif) ligand 22, CCL22, CD200R and mannose receptor C type 1, MRC1) markers to monitor drug effects on "M1 polarization" or cells "pre-polarized to M1". Azithromycin (25-50 μM), tofacitinib (2.5-5 μM), hydroxychloroquine (40 µg/ml) and pioglitazone (15-60 μM) exhibit an anti-inflammatory profile because they downregulated M1 markers and upregulated some M2 markers when given both before and after M1 polarization. Lovastatin given before M1 polarization downregulated M1 marker genes but enhanced the M1 phenotype in macrophages pre-polarized with LPS and IFNγ. Methotrexate (1.25-5 μM) did not modulate macrophage polarization. We have, thus, established a test system suitable to identify novel compounds or repurposed drugs that modulate inflammatory macrophage plasticity. Compounds with potential to reduce expression of molecules involved in inflammatory T cell activation (IL-1β, TNFα, CD80), while enhancing production of a major chemokine involved in recruitment of Tregs (CCL22) may be of interest for treating chronic inflammatory diseases.

Keywords: Cell surface receptor; Chronic inflammatory diseases; Cytokine; Drug screening; Human polarization marker; Macrophage.

Publication types

  • Validation Study

MeSH terms

  • Anti-Inflammatory Agents / pharmacology*
  • Anti-Inflammatory Agents / therapeutic use
  • Biomarkers / metabolism
  • Cell Culture Techniques
  • Cell Differentiation / drug effects*
  • Cell Differentiation / immunology
  • Cells, Cultured
  • Cytokines / genetics
  • Cytokines / immunology
  • Cytokines / metabolism
  • Down-Regulation
  • Drug Evaluation, Preclinical / methods
  • Humans
  • Inflammation / drug therapy*
  • Inflammation / immunology
  • Interferon-gamma / immunology
  • Interleukin-4 / immunology
  • Leukocytes, Mononuclear
  • Lipopolysaccharides / immunology
  • Macrophages / drug effects*
  • Macrophages / immunology
  • Macrophages / metabolism
  • RNA, Messenger / metabolism


  • Anti-Inflammatory Agents
  • Biomarkers
  • Cytokines
  • IL4 protein, human
  • Lipopolysaccharides
  • RNA, Messenger
  • Interleukin-4
  • Interferon-gamma