〔Abstract〕 Objective To screen mitochondria-targeting differential genes in alveolar macrophages of silicosis patients and explore the role of mitochondrial homeostasis in alveolar macrophages of silicosis patients. Methods High-throughput sequencing dataset GSE174725 was downloaded, and differentially expressed genes were screened with R software andP<0.05, |LogFC|>1, and then intermixed with mitochondrial gene bank MitoCarta3.0 to obtain mitochondria-targeted differentially expressed genes. Then enrichment analysis was carried out to obtain the biological processes and pathways involved in differential genes, and the protein-protein interaction network was constructed. In addition, alveolar macrophages from silicosis patients and healthy controls were collected, the expression levels of differential genes were detected by RT-qPCR, and the expressions of mitochondria-related factors mitochondrial fusion protein 1(MFN1), optic atrophy 1(OPA1) and dynamin-related protein 1(DRP1) in alveolar macrophages of silicosis patients were investigated by Western blot. Results A total of 204 differentially expressedgenes were screened in silicosis patients, among which 62 differentially expressed genes were up-regulated, 142 differentially expressed genes were down-regulated, and 22 differentially expressed genes were mitochondria-targeted. The concentration analysis of differentially expressed genes targeted by mitochondria showed that the cell components mainly enriched included mitochondrial membrane, endoplasmic membrane side components, etc. The biological processes mainly enriched included mitochondrial electron transfer from NADH to ubiquinone, inflammatory response, immune response, etc. The main molecular functions enriched included the rotation mechanism of proton transport ATP synthase activity, NADH dehydrogenase activity, chemokine activity, etc. KEGG enrichment analysis mainly focused on the involvement in chemical carcinogenesis-ROS, IL-17 signaling pathway, toll-like receptor signaling pathway, chemokine signaling pathway, TNF signaling pathway, etc. In addition, RT-qPCR results showed that the expressions of mitochondrial cytochrome coxidase 1, mitochondrial cytochrome coxidase 2, mitochondrial cytochrome coxidase 3, mitochondrially encoded NADH dehydrogenase 1, mitochondrially encoded NADH dehydrogenase 3, mitochondrially encoded NADH dehydrogenase 5, superoxide dismutase and mitochondrially encoded ATP synthase 6 gene were down-regulated in silicosis patients(P<0.05). Western blot and RT-qPCR results showed that, in silicosis patients, the expression of MFN1 and OPA1 decreased(P<0.05), while the expression of DRP1 increased(P<0.05). Conclusion Bioinformatics analysis and validation, eight mitochondrial targeted differential genes(MT-CO1, MT-CO2, MT-CO3, MT-ND1, MT-ND3, MT-ND5, SOD and MT-ATP6) were finally obtained, which were enriched in mitochondrial respiratory chain and oxidative stress pathways and might play an important role in the process of silicosis.