Identification and Analysis of Remimazolam’s Potential Therapeutic Mechanisms in Murine Sepsis-Associated Encephalopathy via Transcriptomic Analysis

Background: Sepsis-associated encephalopathy (SAE) is a common complication of sepsis, marked by neuroinflammation and cognitive impairment. Remimazolam (Rem), a novel ultra-short-acting benzodiazepine, exhibits potential anti-inflammatory and neuroprotective effects; however, its molecular mechanisms in SAE remain poorly understood. Methods: Male mice were randomly assigned to three groups: SHAM (Con, n=6), Sepsis (CLP, n=6), and remimazolam-treated Sepsis (Rem, n=6). Hippocampal tissues were collected for RNA sequencing. Differential expression analysis was performed to identify genes modulated by Rem treatment. To further screen candidate genes, weighted gene co-expression network analysis (WGCNA) and protein–protein interaction (PPI) networks were utilized, and functional enrichment was assessed through gene set enrichment analysis (GSEA). Immune microenvironment alterations were compared across groups, and potential therapeutic compounds targeting hub genes were predicted using public drug–gene interaction databases. Hub gene expression was validated at both mRNA (qRT-PCR) and protein levels. Results: RNA-seq identified 3397 and 914 differentially expressed genes (DEGs) in the Sepsis (CLP vs Con) and remimazolam-treated (Rem vs CLP) groups, respectively. A total of 647 key DEGs with opposing expression trends were screened. Through WGCNA and PPI analysis, five hub genes—ICAM1, Tlr2, Cd274, FOS, and SOCS3—were identified. Pathways related to inflammatory response, IL6/JAK/STAT3 signaling, and TNFα/NF-κB signaling were significantly enriched with these genes, suggesting their potential role in the pathogenesis of SAE. Among the twenty-one candidate drugs predicted to target these hub genes, AVELUMAB-CD274 and LIFITEGRAST-ICAM1 exhibited high binding affinity. The expressions of ICAM1, Tlr2, Cd274, and SOCS3 were further validated at both mRNA (qRT-PCR) and protein levels, corroborating the transcriptomic findings. Conclusion: This study reveals that ICAM1, Tlr2, Cd274, and SOCS3 are key molecular correlates of remimazolam treatment in a murine SAE model. Our transcriptomic analysis offers insights into the molecular basis of remimazolam’s protective effects, highlighting these genes and pathways for future research. Keywords: sepsis-associated encephalopathy, remimazolam, immune microenvironmental, RNA sequencing