Chenghui Cao ^1,2 Wenwu Liu ¹ Xin Guo ¹ Shuwei Weng ¹ Yang Chen ¹ Yonghong Luo ¹ Shuai Wang ¹ Botao Zhu ^1,2 Yuxuan Liu ^1,2 Daoquan Peng ^1*

• ¹ Department of Cardiology, The Second Xiangya Hospital, Central South University, Changsha, Hunan Province, China
• ² Xiangya School of Medicine, Central South University, Changsha, Hunan Province, China

Background: Non-alcoholic steatohepatitis (NASH) Metabolic dysfunctionassociated steatohepatitis (MASH) is a highly prevalent liver disease globally, with a significant risk of progressing to cirrhosis and even liver cancer. Efferocytosis, a process implicated in a broad spectrum of chronic inflammatory disorders, has been reported to be associated with the pathogenesis of MASH; however, its precise role remains obscure. Thus, we aimed to identify and validate efferocytosis linked signatures for detection of MASH.We retrieved gene expression patterns of MASH from the GEO database and then focused on assessing the differential expression of efferocytosisrelated genes (EFRGs) between MASH and control groups. This analysis was followed by a series of in-depth investigations, including protein-protein interaction (PPI), correlation analysis, and functional enrichment analysis, to uncover the molecular interactions and pathways at play. To screen for biomarkers for diagnosis, we applied machine learning algorithm to identify hub genes and constructed a clinical predictive model. Additionally, we conducted immune infiltration and single-cell transcriptome analyses in both MASH and control samples, providing insights into the immune cell landscape and cellular heterogeneity in these conditions.Results: This research pinpointed 39 genes exhibiting a robust correlation with efferocytosis in MASH. Among these, five potential diagnostic biomarkers-TREM2, TIMD4, STAB1, C1QC, and DYNLT1-were screened using two distinct machine learning models. Subsequent external validation and animal experimentation validated the upregulation of TREM2 and downregulation of TIMD4 in MASH samples. Notably, both TREM2 and TIMD4 demonstrated area under the curve (AUC) values exceeding 0.9, underscoring their significant potential in facilitating the diagnosis of MASH.Our study comprehensively elucidated the relationship between MASH and efferocytosis, constructing a favorable diagnostic model. Furthermore, we identified potential therapeutic targets for MASH treatment and offered novel insights into unraveling the underlying mechanisms of this disease.