Extracellular vesicles from lung adenocarcinoma cells induce activation of fibroblasts into different cancer-associated fibroblast subtypes

Lung cancer, particularly lung adenocarcinoma (LUAD), is the leading cause of cancer deaths worldwide, largely due to metastasis. This study investigated the impact of extracellular vesicles (EVs) derived from LUAD cells on lung fibroblasts. EVs were isolated from LUAD cell lines via ultracentrifugation and characterized using Nanoparticle Tracking Analysis and western blot. Lung fibroblasts were treated with PBS, TGFβ or EVs, and their activation was assessed through protein (Western Blot) and RNA analysis (RNA seq and RT-qPCR). Results confirmed TGFβ induced activation and showed that LUAD EVs could also activate fibroblasts, increasing cancer-associated fibroblast (CAF) markers. While EV-induced CAF activation displayed unique features, the EV and TGFβ treatments also shared some differentially expressed genes. Mesenchymal genes POSTN and SPOCK1 were significantly upregulated in TGFβ and EV-treated fibroblasts. The secretion as protein of POSTN from the TGFβ and EV-induced CAFs was confirmed through ELISA. These findings suggest that LUAD EVs play a role in CAF activation through both shared and distinct pathways compared to canonical TGFβ activation, potentially identifying novel gene expression involved in CAF activation. RNA sequencing on human lung fibroblast (H6013) primary cells after exposure to PBS, EVs derived from H1437, EVs derived from H2073 or treated with TGFb.