Matrix viscoelasticity amplifies pro-fibrotic crosstalk between cardiac fibroblasts and macrophages.

Cardiac fibrosis is driven by dynamic crosstalk between cardiac fibroblasts and macrophages, yet how tissue mechanics regulate these interactions remains poorly defined. Here, we introduce a viscoelastic coculture platform that enables precise interrogation of mechanical and paracrine signaling in a physiologically relevant context. Counterintuitively, we found that soft, viscous environments promote human-induced pluripotent stem cell-derived cardiac fibroblast activation and macrophage healing phenotypes, while stiff environments bias macrophages toward inflammation. Coculture in soft, viscous matrices amplifies reciprocal pro-fibrotic signaling, while sequential exposure to inflammatory and then healing macrophages, which mimic in vivo dynamics, further exacerbates fibroblast activation. Mechanistically, we identified a STAT1 and AP-1 mediated, viscoelasticity-driven positive feedback loop involving inflammatory cytokines IL6, CCL5, and CCL2 as well as healing cytokines VEGFA and CTGF. This work establishes tissue viscoelasticity as a central regulator of immune-stromal interactions and provides a broadly applicable platform for dissecting mechanobiological drivers of fibrosis.