Abstract: BACKGROUND: Mesenchymal stem cell exosomes have similar biological functions to mesenchymal stem cells and high safety, and can be used for the treatment of fibrosis-related diseases. However, the mechanisms underlying their therapeutic effects on pulmonary fibrosis remain unclear.
OBJECTIVE: To investigate the function and mechanism of bone marrow mesenchymal stem cell exosomes affecting bleomycin-induced pulmonary fibrosis in mice. 
METHODS: Bone marrow mesenchymal stem cells and their exosomes were extracted from C57BL/6J mice and characterized. Thirty C57BL/6J mice were randomly divided into three groups: control group, model group, and exosome group (10 mice per group). The model group and exosome group received intratracheal instillation of 5 mg/kg bleomycin to induce pulmonary fibrosis. On day 7 after bleomycin instillation, mice in the exosome group received 100 μL of 0.1 mg/mL bone marrow mesenchymal stem cell-derived exosomes via tail vein injection once daily for two weeks. After treatment, lung function indicators (airway narrowing index, respiratory rate, and tidal volume) were measured. Hematoxylin-eosin staining and Masson staining were used to assess lung tissue damage and fibrosis. Enzyme-linked immunosorbent assay was used to detect serum levels of hydroxyproline, interleukin-1β, transforming growth factor-β1, and tumor necrosis factor-α. Real-time quantitative PCR was used to detect the mRNA expression of α-smooth muscle actin, collagen I, and fibronectin in lung tissue. Western blotting was used to detect the expression of transforming growth factor-β1 and the phosphorylation level of Smad3 in mouse lung tissue.
RESULTS AND CONCLUSION: Compared with the control group, the model group mice showed a significantly increased airway narrowing index (P < 0.05), significantly decreased respiratory rate and tidal volume (P < 0.05), significantly upregulated α-smooth muscle actin, collagen I, and fibronectin mRNA expression (P < 0.05), significantly increased levels of hydroxyproline, interleukin-1β, transforming growth factor β1, and tumor necrosis factor α (P < 0.05), and significantly increased transforming growth factor β1 protein expression and Smad3 phosphorylation levels (P < 0.05). Compared with the model group, the exosome group mice showed significantly alleviated lung injury and fibrosis, a significantly decreased airway narrowing index (P < 0.05), significantly increased respiratory rate and tidal volume (P < 0.05), significantly downregulated α-smooth muscle actin, collagen I, and fibronectin mRNA expression (P < 0.05), significantly decreased levels of hydroxyproline, interleukin-1β, transforming growth factor β1, and tumor necrosis factor α (P < 0.05), and significantly decreased transforming growth factor β1 protein expression and Smad3 phosphorylation levels (P < 0.05). The results indicate that bone marrow mesenchymal stem cell-derived exosomes inhibit the transforming growth factor β1/Smad3 signaling pathway to ameliorate bleomycin-induced pulmonary fibrosis in mice.

Key words: ">bone marrow mesenchymal stem cell, exosome, bleomycin, mouse, pulmonary fibrosis, inflammation, transforming growth factor-β1/Smad3 signaling pathway, mechanism