Induced pluripotent stem cell-derived macrophages as a model for human inflammasome signaling

Macrophage models are a mainstay of inflammasome research, however current human in vitro macrophage models have significant limitations. Here we generate induced pluripotent stem cell (iPSC)-derived macrophages (iMacs) to study inflammasome signaling and benchmark them with human monocyte-derived macrophages (HMDMs). We confirm that iMacs express high levels of macrophage markers and are highly phagocytic. Whole cell proteomics analysis shows that iMacs express many inflammasome sensors and related proteins, and in functional assays iMacs respond to multiple inflammasome stimuli. The NLRP3 inflammasome is strongly activated in iMacs and we find that nigericin alone activates NLRP3. The non-canonical inflammasome does not require a priming step in iMacs as caspase-4 is constitutively expressed. High levels of NAIP/NLRC4 inflammasome activation are also observed in response to needle toxin. Finally, unlike HMDMs, iMacs activate NLRP1. Therefore, we demonstrate that iMacs are a physiologically relevant and highly tractable model to study human inflammasome signaling and regulation. MotivationiPSC-derived macrophages (iMacs) are functionally, transcriptionally, and phenotypically similar to primary human macrophages. iMacs therefore offer new opportunities to study inflammasome activity in a human macrophage model, but to date they have not been widely used. In this study, we describe a protocol to differentiate and characterize iMacs. We then describe how to activate a range of different inflammasomes within these cells and assess the inflammasome response by measuring pyroptosis, cytokine release, ASC speck formation, and processing of inflammasome-related proteins. We also benchmark iMac responses with the current gold standard primary human monocyte derived macrophage model.