Culture Dimensionality Regulates Protein Expression and Bioactivity in THP-1-Derived Macrophages

Area of Science:

• Immunology
• Biomaterials Science
• Cell Biology

Background:

• In vitro macrophage studies often use 2D cultures lacking structural context, limiting understanding of how spatial dimensionality impacts immune cell signaling and behavior.
• Physiologically relevant environmental cues are crucial for macrophage phenotype and function, yet current models inadequately capture these influences.

Purpose of the Study:

• To investigate the effects of culture dimensionality (2D vs. 3D) and extracellular matrix (ECM) composition on monocyte-derived macrophage phenotype, signaling, and functional activity.
• To compare macrophage behavior in 3D Matrigel and type I collagen matrices versus standard 2D cultures.

Main Methods:

• THP-1 monocytes were differentiated into M0, M1, and M2 macrophages and cultured on 2D substrates or within 3D Matrigel or type I collagen matrices.
• Macrophage morphology, viability, receptor expression, and cytokine secretion were analyzed.
• Functional activity was assessed via co-culture experiments with MDA-MB-231 breast cancer cells.

Main Results:

• 3D environments induced significant morphological and viability changes in macrophages compared to 2D cultures.
• Collagen matrices supported macrophage growth, subtype-specific morphologies, and enhanced functional activity, while Matrigel promoted aggregation and reduced viability.
• Culture dimensionality and ECM composition modulated activation-associated receptors and cytokine profiles, including enhanced TNF-α expression in 3D, with functional impacts on tumor cells being more pronounced in 3D systems.

Conclusions:

• Culture dimensionality and ECM composition are critical regulators of macrophage phenotype and function.
• Collagen-based 3D systems offer a more physiologically relevant model for studying macrophage behavior than 2D platforms.
• Structurally biomimetic models are valuable for immunological research and therapeutic screening.