AREG / EGFR Signalling Primes Macrophage Pyroptosis Through an Autocrine Ccl2‐Driven Amplification Loop

Amphiregulin (AREG) is an epidermal growth factor receptor (EGFR) ligand that has been implicated in inflammatory responses, but its direct role in priming macrophage pyroptosis remains poorly understood. Here, we demonstrate that AREG primes bone marrow-derived macrophages (BMDMs) for ATP-induced pyroptosis through an autocrine Ccl2-dependent positive-feedback loop that requires sustained NF-κB activation. RNA-sequencing and functional clustering analyses revealed that AREG and LPS induce both convergent metabolic reprogramming and divergent transcriptional programs in macrophages. Notably, the chemokine signalling pathway was uniquely enriched in AREG-treated cells, with marked upregulation of Ccl2, Cxcl1, Cxcl3 and Cxcl16. Mechanistically, AREG induced Ccl2 secretion in an EGFR-dependent manner. Pharmacological inhibition (mNOX-E36) or antibody-mediated neutralisation of Ccl2 dose-dependently suppressed Caspase1/GSDMD cleavage and IL-1β/IL-18 release, and attenuated NF-κB p65 phosphorylation and IκBα degradation during AREG plus ATP stimulation. Consistently, siRNA-mediated Ccl2 knockdown phenocopied these effects, confirming that autocrine Ccl2 signalling sustains NF-κB activation to drive AREG-primed pyroptosis. Collectively, these findings identify a previously unrecognised AREG/EGFR/Ccl2/NF-κB axis that primes macrophages for pyroptosis, highlighting Ccl2 as a potential therapeutic target in sepsis and other inflammatory diseases driven by this amplification loop.