In this study, we utilized bioinformatics to investigate the intricate interplay between mitochondria-related genes and the immune microenvironment in sepsis, a life-threatening condition with complex pathophysiology. Through analysis of mRNA expression profiles from NCBI GEO, we identified 49 differentially expressed genes (DEGs) associated with mitochondria (MitoDEGs) by intersecting them with mitochondrial lineage genes. Gene ontology (GO) enrichment and Kyoto Gene and Genome Encyclopedia (KEGG) pathway analyses were conducted to elucidate the functional roles of MitoDEGs in sepsis. A protein‒protein interaction (PPI) network highlighted 10 central MitoDEGs as hub genes crucial in sepsis. Furthermore, utilizing NetworkAnalyst, we predicted transcription factors and constructed a competitive endogenous RNA (ceRNA) regulatory network to unveil potential interactions of MitoDEGs with miRNAs and lncRNAs. Notably, our study revealed alterations in immune cell infiltration within the sepsis microenvironment, with Spearman analysis demonstrating significant correlations between hub MitoDEGs and specific immune cell subsets. These comprehensive findings shed light on the crosstalk between mitochondrial function and immune responses in sepsis, offering novel insights for the identification of therapeutic targets in the management of this critical condition.