In recent years, several novel macrophage phenotypes have been identified and characterized, such as M3, M4, Mox, Mhem, M(Hb), M(IL-10), Mregs, and TAMs. Among them, M4 macrophage is a relatively new and emerging concept that has attracted much attention in the field of immunology and inflammation.

M4 macrophages are a heterogeneous and dynamic population that can be divided into different subtypes based on their expression of Trem2, CD206, CD163, and other molecules. M4 macrophage subtypes have distinct phenotypes and functions in phagocytosis, inflammation, lipid metabolism, antigen presentation, and tissue repair.

Fig.1 Several major macrophage subtypes.¹

Creative Biolabs provides an overview of the current knowledge of M4 macrophages, including their subtypes, functions, roles in diseases, and therapeutic applications.

Subtypes and Functions of M4 Macrophage

M4 macrophages are a heterogeneous and dynamic population that can be further divided into different subtypes based on their expression of Trem2, CD206, CD163, and other molecules. The main subtypes of M4 macrophages are M4a, M4b, and M4c, which have distinct phenotypes and functions.

• M4a macrophage is characterized by high expression of Trem2 and CD206, but low expression of CD163. M4a macrophage is induced by apoptotic cells or oxLDL, and has a high capacity for macrophage phagocytosis and lipid uptake.It also produces anti-inflammatory cytokines and promotes tissue repair and resolution of inflammation. M4a macrophage is considered as a beneficial subtype that protects against atherosclerosis and Alzheimer's disease.
• M4b macrophage is characterized by high expression of Trem2 and CD163, but low expression of CD206. M4b macrophage is induced by heme or hemoglobin, and has a high capacity for iron metabolism and clearance. M4b macrophage also produces anti-inflammatory cytokines and inhibits the activation of pro-inflammatory M1 macrophage. M4b macrophage is considered as a beneficial subtype that protects against anemia and sepsis.
• M4c macrophage is characterized by low expression of Trem2, CD206, and CD163. M4c macrophage is induced by nucleic acids or Aβ, and has a high capacity for antigen presentation and cytokine production. M4c macrophage also produces pro-inflammatory cytokines and activates the adaptive immune response. M4c macrophage is considered as a detrimental subtype that contributes to autoimmune diseases and neurodegeneration.

M4 macrophage subtypes can interact with other immune cells and modulate their responses. These interactions can have significant effects on the outcome of inflammation and immunity.

• M4a macrophage can suppress the differentiation of Th1 and Th17 cells, but promote the differentiation of Treg cells.
• M4b macrophage can enhance the phagocytosis of neutrophils and eosinophils, but inhibit the degranulation of mast cells.
• M4c macrophage can stimulate the proliferation of B cells and plasma cells, but impair the function of NK cells.

Roles of M4 Macrophages in Diseases

M4 macrophages are involved in various diseases, such as leprosy, tuberculosis, atherosclerosis, obesity, diabetes, Alzheimer's disease, and cancer. The activation and dysregulation of M4 macrophages can have different effects on disease pathogenesis and progression, depending on the subtype, stimulus, and context.

• Leprosy
  • M4 macrophage expresses high levels of Trem2 and CD206, which facilitate the phagocytosis and clearance of M. leprae.
  • M4 macrophage also produces anti-inflammatory cytokines, such as IL-10 and TGF-β, which suppress the inflammatory response and prevent nerve damage.
• Tuberculosis
  • M4 macrophage expresses high levels of Trem2 and CD163, which impair the phagocytosis and killing of M. tuberculosis.
  • M4 macrophage also produces anti-inflammatory cytokines, such as IL-10 and TGF-β, which inhibit the activation of Th1 cells and macrophages.
• Alzheimer's disease
  • On one hand, M4 macrophage expresses high levels of Trem2 and CD206, which enhance the phagocytosis and clearance of Aβ. M4 macrophage also produces anti-inflammatory cytokines that attenuate the neuroinflammation and neurodegeneration.
  • On the other hand, M4 macrophage accumulates large amounts of Aβ and becomes amyloid-associated microglia (AAM). M4 macrophage also produces pro-inflammatory cytokines, such as TNF-α and IL-6, which exacerbate the neuroinflammation and neurodegeneration.
• Cancer
  • M4 macrophage expresses high levels of Trem2 and CD163, which impair the phagocytosis and killing of tumor cells.
  • M4 macrophage also produces anti-inflammatory cytokines, such as IL-10 and TGF-β, which inhibit the activation of cytotoxic T cells and natural killer cells.

M4 macrophage can also serve as a biomarker or a diagnostic tool for disease detection and monitoring.

• For example, the expression level of Trem2 or CD163 on M4 macrophage can reflect the severity or stage of leprosy, tuberculosis, atherosclerosis, obesity, diabetes, Alzheimer's disease, or cancer.
• The ratio of M4a to M4b or M4c macrophage can indicate the balance or imbalance of inflammation and resolution in these diseases.
• The detection of M4 macrophage in blood or tissue samples can help to diagnose or prognose these diseases.

Therapeutic Applications of M4 Macrophage

Several strategies and challenges of manipulating M4 macrophage for therapeutic purposes have been explored, such as enhancing their anti-inflammatory and pro-resolving functions, modulating their lipid metabolism, targeting macrophage receptors or signaling pathways, or delivering drugs or genes to them.

For instance, one strategy is to enhance the anti-inflammatory and pro-resolving functions of M4 macrophage, especially M4a and M4b subtypes, which can suppress the inflammatory response and promote tissue repair and resolution. This can be achieved by using natural or synthetic compounds that can induce or mimic the effects of the main stimuli for M4a and M4b macrophage polarization.

• For example, annexin A1, a protein that is exposed on the surface of apoptotic cells, can bind to formyl peptide receptor 2 (FPR2) on macrophages and induce M4a polarization.
• Similarly, hemopexin, a protein that binds to heme and hemoglobin, can activate CD163 on macrophages and induce M4b polarization.

References

1. Fang F, et al. "Tuning macrophages for atherosclerosis treatment". Regen Biomater, (2022).
2. Quaresma TC, et al. "Immunohistochemical Characterization of M1, M2, and M4 Macrophages in Leprosy Skin Lesions". Pathogens, (2023).