PLAU promotes fibroblast activation and keloid fibrogenesis through PLAUR-Dependent TGF-β/Smad signaling.

BACKGROUND: Keloids are fibroproliferative disorders conditions of the dermis with dysregulated immune responses contributing to disease progression. However, the causal inflammatory mediators and their pathogenic mechanisms remain undefined. PURPOSE: This study aims to identify causal inflammatory mediators for keloids and their pathogenic mechanisms. METHODOLOGY: Two-sample Mendelian randomization (MR) analyses were performed using genetic instruments for 91 inflammatory proteins as exposures and genome-wide association study (GWAS) summary statistics for keloid susceptibility as the outcome. Sensitivity analyses were conducted to assess the robustness of MR findings. Single-cell RNA sequencing (scRNA-seq) datasets from keloid tissues were analyzed to identify the cellular distribution of prioritized candidates. Functional validation was performed using immunohistochemistry (IHC), immunofluorescence staining, qRT-PCR, western blotting, siRNA-mediated knockdown, recombinant protein stimulation, and transcriptome sequencing. Pharmacological inhibition and rescue experiments were further performed to investigate the downstream signaling pathway. RESULTS: MR analysis identified five inflammatory proteins with potential causal associations with keloid susceptibility, among which urokinase-type plasminogen activator (PLAU) was the only one consistently enriched in fibroblasts across two independent scRNA-seq datasets. PLAU expression was significantly elevated at both the mRNA and protein levels in keloid tissues and primary fibroblasts. PLAU knockdown in keloid fibroblasts reduced collagen I/III expression, proliferation, and migration, whereas recombinant PLAU stimulation enhanced these profibrotic phenotypes in normal dermal fibroblasts. Mechanistically, RNA-sequencing implicated activation of the TGF-β signaling pathway. Furthermore, inhibition of TGF-β receptor signaling attenuated PLAU-induced profibrotic responses, while exogenous TGF-β1 rescued the effects of PLAU depletion. Importantly, PLAUR knockdown attenuated PLAU-induced TGF-β/Smad activation and reduced PLAU-mediated enhancement of fibroblast activation. CONCLUSIONS: This integrative study identifies PLAU as a genetically supported and biologically relevant mediator of keloid fibrosis. PLAU promotes fibroblast activation through PLAUR-dependent regulation of the TGF-β/Smad signaling pathway, leading to enhanced collagen production, proliferation, and migration. These findings provide new insights into inflammatory regulation in keloid pathogenesis and highlight PLAU as a potential therapeutic target for keloid intervention.