Secreted Cytokines From Inflammatory Macrophages Modulate Sex Differences in Valvular Interstitial Cells on Hydrogel Biomaterials

Patients with aortic valve stenosis (AVS) experience fibrosis and/or calcification in valve tissue, which leads to heart failure if left untreated. Inflammation is a hallmark of AVS, and secreted cytokines from pro-inflammatory macrophages are thought to contribute to valve fibro-calcification by driving the activation of valvular interstitial cells (VICs) to myofibroblasts. However, the molecular mechanisms by which inflammatory cytokines differentially regulate myofibroblast activation as a function of biological sex are not fully defined. Here, we developed an in vitro hydrogel culture platform to culture male and female valvular interstitial cells (VICs) and characterize the sex-specific effects of inflammatory cytokines on VIC activation to myofibroblasts and osteoblast-like cells. Our data reveal that tumor necrosis factor alpha (TNF-α) modulates female-specific myofibroblast activation via MAPK/ERK signaling, nuclear chromatin availability, and osteoblast-like differentiation via RUNX2 nuclear localization. In parallel, our data also suggest that male-specific myofibroblast deactivation in response to TNF-α occurs via alternative pathways outside of MAPK/ERK signaling. Collectively, hydrogel biomaterials as cell culture platforms are critical for distinguishing sex differences in cellular phenotypes.