Epithelial multicellular clustering enabled by polarized macrophages on soft matrices

Area of Science:

• Biomedical Engineering
• Cell Biology
• Immunology

Background:

• Epithelial structure formation is vital for organogenesis, tumor growth, and wound repair.
• The influence of immune cells and microenvironmental mechanical cues on epithelial cell clustering remains poorly understood.

Purpose of the Study:

• To investigate how immune cells (macrophages) and matrix stiffness affect human mammary epithelial cell clustering.
• To elucidate the molecular mechanisms underlying macrophage-mediated regulation of epithelial morphogenesis.

Main Methods:

• Coculture of human mammary epithelial cells with M0, M1, or M2 macrophages on soft or stiff hydrogels.
• Analysis of epithelial cell migration, clustering, focal adhesions, fibronectin deposition, and nonmuscle myosin-IIA expression.
• Assessment of cytokine secretion (TNF-α, TGF-β) and the effect of ROCK inhibition and exogenous TGF-β.

Main Results:

• M1 macrophages on soft matrices significantly enhanced epithelial cell migration and multicellular clustering compared to M0 or M2 macrophages.
• Stiff matrices inhibited epithelial clustering by increasing cell migration and cell-extracellular matrix adhesion.
• Soft matrices with M1 macrophages modulated focal adhesions, fibronectin, and nonmuscle myosin-IIA, promoting clustering, which was dependent on cellular forces (ROCK pathway).
• Macrophage-secreted factors, particularly TGF-β, influenced epithelial clustering.

Conclusions:

• Both mechanical properties of the microenvironment and immune cell polarization are critical regulators of epithelial cell clustering.
• Understanding these interactions can inform strategies for modulating tissue repair, fibrosis, and tumor progression.