AbstractObesity, characterized by accumulation of adipose, is usually accompanied by hyperlipidemia and abnormal glucose metabolism, which destroys the function and structure of islet β cells. However, the exact mechanism of islet deterioration caused by obesity has not yet been fully elucidated. Here, we fed C57BL/6 mice with a high-fat diet (HFD) for 2 (2M group) and 6 months (6M group) to construct obesity mouse models. Then, RNA-based sequencing was used to identify the molecular mechanisms in HFD-induced islet dysfunction. Compared with the control diet, a total of 262 and 428 differentially expressed genes (DEGs) were identified from islets of the 2M and 6M groups, respectively. GO and KEGG enrichment analysis revealed that the DEGs up-regulated in both the 2M and 6M groups are mainly enriched in response to endoplasmic reticulum stress and the pancreatic secretion pathway. DEGs down-regulated in both the 2M and 6M groups are mainly enriched in the neuronal cell body and protein digestion and absorption pathway. Notably, along with the HFD feeding, mRNA expression of islet cell markers was significantly down-regulated, such asIns1, Pdx1, MafA(β cell),Gcg, Arx(α cell),Sst(δcell), andPpy(PP cell). In contrast, mRNA expression of acinar cell markers was remarkably up-regulated, such asAmy1, Prss2, andPnlip. Besides, a large number of collagen genes were down-regulated, such asCol1a1, Col6a6, andCol9a2. Overall, our study provides a full-scale DEG map regarding HFD-induced islet dysfunction, which was helpful to understand the underlying molecular mechanism of islet deterioration further.