GEOapplication/xmlftp://ftp.ncbi.nlm.nih.gov/geo/series/GSE328nnn/GSE328550/primaryOK200TranscriptomicsMus musculusExpression profiling by high throughput sequencinghttps://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE328550GEOGSEfalseMacrophage-fibroblast crosstalk shapes wound repair signaling in vitroWound healing requires coordinated interactions between macrophages and fibroblasts, yet how contact-dependent signaling integrates with paracrine pathways to regulate their reciprocal behavior is not well defined. Here, we investigated macrophage-fibroblast communication using complementary 2D and 3D in vitro wound healing models combined with live-cell calcium imaging and single-cell RNA sequencing (scRNA-seq). We show that bone marrow-derived macrophages (BMDMs) promote fibroblast scratch closure in a contact-dependent manner independent of connexins, whereas fibroblasts reciprocally regulate macrophage cytokine secretion through distinct mechanisms. Direct cell-cell contact with fibroblasts enhanced macrophage IL10 production via connexin 43 (Cx43)-dependent signaling, whereas TNFα secretion was suppressed through paracrine interactions. We further demonstrate that fibroblast-macrophage contact induces connexin-dependent intermittent calcium signals selectively in macrophages. ScRNA-seq revealed that wounding reshapes macrophage and fibroblast populations, uncovering dynamic regulation of cell adhesion molecules (CAMs) and intercellular signaling pathways. Together, these findings reveal the integration of contact-dependent calcium and connexin signaling with transcriptional remodeling to coordinate macrophage-fibroblast behavior during healing.2026/04/24GSE328550GSM9685178GSM968517734328328550Mus musculus