Primary Cilium-dependent Humoral Bioactive Factors Acts in a Paracrine Manner to Control Fibroblast Cell Migration

Abstract

The primary cilium is classically recognized as a signal-reception hub, yet its role in mediating cell-to-cell communication via signal spread remains poorly defined. Here, we uncover a previously unrecognized paracrine function of the primary cilium in fibroblasts. Conditioned medium from ciliated wild-type NIH/3T3 cells enhanced wound healing in primary cilium-deficient fibroblasts, in contrast to conditioned medium derived from primary cilium-deficient NIH/3T3-Kif3a-KO or NIH/3T3-Dync2h1-KO cells. Fractionation of conditioned medium revealed that the wound healing activity resided predominantly in the 100K x g soluble supernatant (Sup-100K), rather than in extracellular vesicle (EV) fractions. Untargeted metabolomic analysis identified lysophosphatidylcholine (LPC) (14:0) as a key bioactive metabolite enriched in WT-Sup-100K secretome. Supplementation of LPC(14:0) restored wound healing capacity in NIH/3T3-Kif3a-KO cells to levels comparable to WT-Sup-100K treatment. Transcriptomic profiling of target cells revealed that WT-Sup-100K upregulated expression of extracellular matrix (ECM)-associated genes, including Ogn, Igf2, and Mfap4, while EVs modestly enhanced early ECM remodeling via induction of Nid2. Together, these findings demonstrate that the primary cilium coordinates a wound healing secretome in fibroblasts through the regulated release of LPC(14:0) and other soluble factors that activate ECM-remodeling pathways in recipient cells. This work expands the functional repertoire of the primary cilium and establishes its critical role in coordinating paracrine regenerative signaling.