Abstract Body: Background: Mammalian hearts have evolved distinct chambers with specialized functions. Single-cell and spatial transcriptomics reveal chamber-specific cellular heterogeneity, but the mechanisms establishing transcriptomic and metabolic differences remain unclear. The Hippo pathway inhibits the transcriptional co-activator YAP, which promotes fibrosis and inflammation in cardiac fibroblasts (CFs). However, its role in regulating the metabolic microenvironment during homeostasis and fibroinflammation is not well understood.

Hypothesis: YAP in CFs regulates the metabolic microenvironment during homeostasis and fibroinflammation.

Methods: We analyzed YAP target and glycolysis gene expression in CFs using human snRNA-seq data. Glucose uptake was compared between atria via isotope-labeled glucose uptake in isolated mouse atria. YAP was activated in CFs by conditionally knocking out Hippo kinases Lats1/2 (Lats1/2 CKO) in mouse hearts. We performed metabolic assays, snRNA-seq, snATAC-seq, and spatial transcriptomics (ST) on Lats1/2 CKO and control hearts. Ligand-receptor analysis was conducted on snRNA-seq data to examine CF interactions, and functional roles were tested using pharmacologic inhibitors.

Results: YAP activity was higher in right atrial (RA) CFs than in other chambers. RA CFs exhibited increased glycolysis and glucose uptake compared to left atrial CFs. Metabolic assays, snRNA-seq, and ST analyses revealed that Hippo-deficient CFs activated YAP and glycolysis, driving fibrosis and inflammation. Pharmacologic studies showed that YAP promoted macrophage (Mac) expansion via CSF1 signaling. Macs secreted IGF1, activating IGF1 signaling in Hippo-deficient CFs to enhance proliferation and fibrosis.

Conclusion: RA CFs exhibit higher YAP and glycolysis activity during homeostasis. YAP activation in Hippo-deficient CFs further enhances glycolysis to induce RA fibroinflammation. YAP-driven CSF1 signaling promotes macrophage self-renewal, while macrophage-derived IGF1 enhances CF proliferation and fibrosis. Since CSF1 and IGF1 signaling increase glycolysis, our findings suggest that YAP fosters a glycolytic microenvironment by reinforcing CF-Mac interactions through CSF1/IGF1 signaling.