Lin Tian

Silicosis is a life-threatening occupational disease characterized by irreversible pulmonary fibrosis. While lung microbiota dysbiosis is implicated in its pathogenesis, the molecular mechanisms linking microbial alterations to fibrotic progression remain incompletely understood. Building upon our previous findings of silica-induced lung microbiota dysbiosis, this study investigated whether the lung microbiota drives silicosis fibrosis via epithelial–mesenchymal transition (EMT). This study reveals that antibiotic intervention significantly attenuates pulmonary fibrosis and EMT in silica-induced silicosis mouse model. Mechanistically, in vivo and in vitro results revealed that silica and lung microbiota-derived lipopolysaccharide (LPS) synergistically drive alveolar epithelial cells toward an EMT phenotype. This process is mediated by the sustained upregulation of the transcription factor Snail, which acts as a master regulator to amplify EMT signaling. Subsequently, these EMT-transformed cells orchestrate fibroblast activation and collagen deposition through paracrine communication. Crucially, silencing Snail or depleting the microbiota effectively reversed these pathological changes and inhibited fibroblast activation. These findings establish a critical “lung microbiota–Snail–EMT–fibroblast” axis in the pathogenesis of silicosis. This study provides compelling evidence that the lung microbiota is an active driver rather than a passive bystander in fibrosis, suggesting that targeting this signaling axis offers a promising therapeutic strategy for silicosis.

Background: In clinical practice, due to the lack of typical symptoms and specific diagnostic biomarkers, silicotic patients often having already developed pulmonary fibrosis by the time of clinical diagnosis. Studies have demonstrated that human umbilical cord mesenchymal stem cell-derived extracellular vesicles (hucMSC-EVs) could moderate silicosis fibrosis, which may be related to the microRNAs (miRNAs) in hucMSC-EVs. While the full extent of their antifibrotic effects and the underlying mechanisms remain to be elucidated. Methods: HucMSC-EVs were administered from day 28 to day 56 after silica exposure in mice, which in a therapeutic manner. In addition, the antifibrotic abilities of engineered hucMSC-EVs with varying levels of miR-148a-3p, a miRNA with antifibrotic properties, were evaluated. Heat shock protein 90 beta family member 1 (Hsp90b1) is reported to be a target of miR-148a-3p, the protein-protein interaction analysis was used to explore its regulated downstream factors in lung fibrosis. The underlying mechanisms were also investigated by using miR-148a-3p mimics and small interfering RNA (siRNA) targeting Hsp90b1 in vitro. Results: HucMSC-EVs could reduce the histopathological changes and the levels of fibrotic proteins in the mouse lung tissues when administered in a therapeutic manner. Meanwhile, miR-148a-3p-overexpressed hucMSC-EVs intervention exhibited the enhanced anti-fibrotic effect compared with the negative control intervention group. In vitro, the elevated level of miR-148a-3p in hucMSC-EVs was shown to enhance hucMSC-EVs’ inhibition of fibroblast collagen hypersecretion, whereas a depressed level of miR-148a-3p in hucMSC-EVs partially counteracted the inhibitory effect. Moreover, the mechanistic investigations revealed that miR-148a-3p could blunt β-catenin signaling via targeting Hsp90b1 in fibroblasts. Conclusion: This study demonstrated that hucMSC-EVs retain their antifibrotic properties in silicotic mice when administered in a therapeutic manner. Further, miR-148a-3p was confirmed to be an essential component within hucMSC-EVs, mediating their inhibition of silica-induced pulmonary fibrosis by reducing β-catenin signaling via targeting of Hsp90b1 in fibroblasts. Plain Language Summary: Silicosis is a life-threatening occupational pulmonary fibrotic disease caused by silica inhalation, and there is currently a lack of targeted drugs for treating silicosis. Extracellular vesicles (EVs) are emerging as pivotal players in cell-free therapeutics due to their crucial role in intercellular communication and the efficient transport of bioactive molecules. In this study, the antifibrotic effects of human umbilical cord mesenchymal stem cell-derived EVs (hucMSC-EVs) on experimental silicosis were investigated. HucMSC-EVs were administered therapeutically, an approach that aligns more closely with the clinical situation of silicosis. In addition, the antifibrotic abilities of engineered hucMSC-EVs with varying levels of miR-148a-3p, a microRNA (miRNA) with antifibrotic properties, were evaluated. The underlying mechanisms that contribute to these therapeutic effects were also investigated. The results demonstrated that hucMSC-EVs retain their antifibrotic properties in silicotic mice when administered in a therapeutic manner. Further, miR-148a-3p was confirmed to be an essential component within hucMSC-EVs, mediating their inhibition of silica-induced pulmonary fibrosis by reducing β-catenin signaling via targeting of heat shock protein 90 beta family member 1 (Hsp90b1) in fibroblasts. These insights provide strong empirical support for hucMSC-EVs-based therapy as a novel treatment approach for silicosis and other lung fibrotic diseases.

Silicosis is a fatal occupational respiratory disease caused by the prolonged inhalation of respirable silica. The core event of silicosis is the heightened activity of fibroblasts, which excessively synthesize extracellular matrix (ECM) proteins. Our previous studies have highlighted that human umbilical cord mesenchymal stem cell-derived extracellular vesicles (hucMSC-EVs) hold promise in mitigating silicosis and the significant role played by microRNAs (miRNAs) in this process. Delving deeper into this mechanism, we found that miR-148a-3p was the most abundant miRNA of the differential miRNAs in hucMSC-EVs, with the gene heat shock protein 90 beta family member 1 (Hsp90b1) as a potential target. Notably, miR-148a-3p’s expression was downregulated during the progression of silica-induced pulmonary fibrosis both in vitro and in vivo, but was restored after hucMSC-EVs treatment (p < 0.05). Introducing miR-148a-3p mimics effectively hindered the collagen synthesis and secretion of fibroblasts induced by transforming growth factor-β1 (TGF-β1) (p < 0.05). Confirming our hypothesis, Hsp90b1 was indeed targeted by miR-148a-3p, with significantly reduced collagen activity in TGF-β1-treated fibroblasts upon Hsp90b1 inhibition (p < 0.05). Collectively, our findings provide compelling evidence that links miR-148a-3p present in hucMSC-EVs with the amelioration of silicosis, suggesting its therapeutic potential by specifically targeting Hsp90b1, thereby inhibiting fibroblast collagen activities. This study sheds light on the role of miR-148a-3p in hucMSC-EVs, opening avenues for innovative therapeutic interventions targeting molecular pathways in pulmonary fibrosis.