Secreted RCN3 acts as an early epithelial-fibroblast mediator via TGFβR1-Smad signaling in post-ALI pulmonary fibrosis.

Pulmonary fibrosis (PF), driven by dysregulated epithelial-fibroblast interactions, contributes to poor outcomes after acute pneumonia. However, early profibrotic paracrine mediators released by injured epithelium remain incompletely defined. Using secretomics of LPS-treated pulmonary epithelial cells, we identified Reticulocalbin 3 (RCN3) as an epithelial paracrine mediator. In clinical bronchoalveolar lavage fluid (BALF) samples, RCN3 was significantly higher in organizing pneumonia patients undergoing active fibrotic organization than in idiopathic PF patients in a stable fibrotic state. In LPS-induced acute lung injury (ALI) mice, BALF RCN3 peaked earlier than TGFβ1/FGF2/CTGF, indicating an early role in PF. Mechanistically, epithelial RCN3 is secreted via an N140-glycosylation–dependent ER-Golgi pathway and engages TGFβR1, activating canonical Smad2/3 signaling in fibroblasts; RCN3 also upregulates TGFβ1 and TGFβR1/2 in a Smad3-dependent manner. In vivo, intratracheal exogenous RCN3 administration, in the absence of LPS, was sufficient to trigger fibrosis, whereas early-phase neutralization of RCN3 after LPS-ALI attenuated fibrotic remodeling. Collectively, these findings identify secreted, stress-responsive RCN3 as an early epithelial paracrine mediator engaging TGFβR1-Smad2/3 signaling and nominate it as a potential early-window therapeutic target for post-ALI pulmonary fibrosis.