Protocol to study inflammasome activation in induced pluripotent stem cell-derived macrophages

Area of Science:

• Immunology
• Stem Cell Biology
• Molecular Biology

Background:

• Current in vitro macrophage models present limitations for studying inflammasome activity.
• Induced pluripotent stem cells (iPSCs) offer a promising source for generating human macrophages.

Purpose of the Study:

• To establish and validate a protocol for differentiating iPSCs into macrophages (iMacs).
• To characterize iMacs and demonstrate their utility in studying inflammasome activation.
• To provide a comprehensive method for assessing inflammasome responses in iMacs.

Main Methods:

• Differentiation of iPSCs into macrophages (iMacs).
• Characterization of iMacs using flow cytometry, phagocytosis assays, and whole-cell proteomics.
• Activation of various inflammasomes within iMacs.
• Assessment of inflammasome responses via pyroptosis, cytokine release, ASC speck formation, and western blotting.

Main Results:

• Successful differentiation and characterization of iPSCs into functional macrophages (iMacs).
• Demonstrated ability to activate diverse inflammasomes within iMacs.
• Quantified inflammasome responses including pyroptosis, cytokine release, and protein processing.

Conclusions:

• iMacs provide a robust and versatile in vitro model for inflammasome research.
• This protocol enables detailed investigation of inflammasome pathways using a renewable cell source.
• The characterized iMacs can advance our understanding of inflammasome-mediated diseases.