二氧化硅诱导小鼠肺纤维化进程中miRNA-mRNA调控网络的构建与分析OA

[Background]Regulatory interactions between microRNAs(miRNAs)and messenger RNAs(mRNAs)are involved in the progression of pulmonary fibrosis,which can either promote or inhibit the development of this disease. [Objective]To explore the miRNA-mRNA regulatory network during the progression of silica(SiO2)-induced pulmonary fibrosis in mice using integrated mRNA-seq and miRNA-seq analysis. [Methods]A mouse model of pulmonary fibrosis was established by dynamic SiO2 dust exposure.The experimental design included a blank control group and four SiO2-exposed groups(7,14,28,and 56 d,n=10 per group).Successful model induction was confirmed by histopathological analysis(HE and Masson staining),hydroxyproline(HYP)quantification,and expression of key fibrosis-related cytokines[fibroblast growth factor(FGF),interleukin-6(IL-6),transforming growth factor-β(TGF-β),and tumor necrosis factor-α(TNF-α)].Lung tissues from mice in each group were subjected to sequencing,and Mfuzz was used for time-series gene clustering to identify dynamic progression patterns.DESeq2 was utilized to identify differentially expressed genes(DEGs)and differentially expressed miRNAs.Enrichment analysis of DEGs was performed to identify critical signaling pathways and biological processes underlying pulmonary fibrosis progression.Expres-sion of four selected miRNAs was subsequently validated by real-time quantitative polymerase chain reaction(RT-qPCR).The target mRNAs of key miRNAs were comprehensively predicted by integrating miRBase,starBase,and miRTarBase to construct the regulatory networks and investigate potential functions. [Results]SiO2 exposure led to time-dependent aggravation of pulmonary fibrosis in mice,evidenced by increased fibrous deposition,ele-vated HYP levels(P<0.01),and up-regulation of four kinds of pro-fibrotic cytokines(P<0.01)compared with the NT group.Mfuzz clustering revealed the stage-specific characteristics.Compared to controls,231,662,448,and 1 020 DEGs were identified after SiO2 exposure at 7,14,28,and 56 d,respectively,primarily enriched in immune responses and chemokine signaling.During critical fibrotic phases—7 d(acute inflammation and initiation)and 28 d(chronic inflammation and establishment)—18 differentially expressed miRNAs were identified;notably mmu-miR-135b-5p was significantly dysregulated at both time points.The expression trends of the four key miRNAs(mmu-miR-135b-5p,mmu-miR-708-5p,mmu-miR-21a-3p,and mmu-miR-205-5p)were consistent with the sequencing results.Further-more,bioinformatics databases were used to predict the target mRNAs of key miRNAs.The constructed network highlighted critical miR-NA-mRNA pairs—including mmu-miR-135b-5p and Meis1,mmu-miR-708-5p and Mmp25,mmu-miR-21a-3p and Cacna1d,mmu-miR-205-5p and Ereg which were closely associated with inflammatory response,extracellular matrix deposition,and fibroblast activation. [Conclusion]The progression of pulmonary fibrosis is accompanied by dynamic changes in miRNA-mRNA regulatory networks.The iden-tified miRNA-target axes(e.g.,miR-135b-5p and Meis1,mmu-miR-708-5p and Mmp25,mmu-miR-21a-3p and Cacna1d,and mmu-miR-205-5p and Ereg—)may play important roles in fibrogenesis and provide potential therapeutic targets for pulmonary fibrosis.