The prolyl hydroxylase PHD3 identifies proinflammatory macrophages and its expression is regulated by activin A

Area of Science:

• Immunology
• Cell Biology
• Molecular Biology

Background:

• Macrophage polarization is crucial for inflammation, with M1 (proinflammatory) and M2 (anti-inflammatory) subtypes identified by cytokine profiles.
• Identifying polarization-specific molecules is key for developing diagnostic and therapeutic strategies for inflammatory diseases.

Purpose of the Study:

• To investigate the expression and regulation of the EGLN3 gene, encoding prolyl hydroxylase PHD3, in human macrophages.
• To determine if PHD3 expression can serve as a marker for specific macrophage polarization states in vitro and in vivo.

Main Methods:

• In vitro generation of human monocyte-derived macrophages polarized to M1 (GM-CSF) or M2 (M-CSF) states.
• Quantitative real-time PCR and Western blotting to assess EGLN3 mRNA and protein expression.
• Immunohistochemical analysis of PHD3 expression in human tissues (lung, Crohn's disease, ulcerative colitis, melanoma).
• Investigating the role of activin A and hypoxia in regulating EGLN3 expression using blocking antibodies and kinase inhibitors.

Main Results:

• EGLN3 was specifically expressed at mRNA and protein levels in proinflammatory M1(GM-CSF) macrophages in vitro.
• PHD3 was detected in lung macrophages under basal conditions and abundantly in inflammatory tissues and tumors.
• PHD3 expression identified a subset of tumor-associated macrophages in melanoma with reduced M2 markers.
• EGLN3 expression in M1 macrophages was dependent on activin A, while hypoxia upregulated EGLN3 in M2 macrophages, modulated by activin A.

Conclusions:

• The EGLN3 gene and its encoded PHD3 protein are specific markers for proinflammatory M1 macrophages.
• PHD3 expression in macrophages is regulated by activin A under both basal and hypoxic conditions.
• PHD3 represents a novel molecular marker for identifying proinflammatory macrophages in diverse pathological settings.