CD44 and CLDN3 as immune-metabolic regulators in acute pancreatitis: A multi-modal transcriptomics study and experimental validation

Xinwei Wang¹Cheng Hu¹Tian Liu²Rui Yang¹Yuxin Shen¹Shihang Zhang¹Lihui Deng^1,3*Qing Xia¹

• ¹West China Centre of Excellence for Pancreatitis, Institute of Integrated Traditional Chinese and Western Medicine, West China Hospital, Sichuan University, Chengdu, China
• ²Laboratory of Integrated Traditional Chinese and Western Medicine, West China Hospital, Sichuan University, Chengdu, China
• ³Sichuan University, Chengdu, China

Acute pancreatitis (AP) is an inflammatory disorder of exocrine pancreas regulated by a complex interaction between injured pancreatic acinar cells and immune cells. Recent studies indicated the crucial role of glycolysis in regulating immune cell function and inflammation. Here, we identified 43 glycolysis-related differentially expressed genes (DEGs) from transcriptomic datasets (GSE65146 and GSE109227). Further, we used three machine learning algorithms to identify Claudin-3 (CLDN3) and CD44 (CD44) were key glycolysis-related DEGs. We validate their significant upregulation in an independent dataset. Then, single-sample gene set enrichment analysis revealed CLDN3 and CD44 were significantly correlated with immune-related structural remodeling and immune infiltration patterns. Single-cell RNA-seq analysis from GSE279876 confirmed that CLDN3 was downregulated in acinar cells, while CD44 was enriched in ductal and immune cells. To validate these findings, we established AP animal model by 10 hourly intraperitoneal injections of caerulein (100 μg/kg) combined with one injection of lipopolysaccharide (10mg/kg). We validated CD44 was upregulated, which mainly expressed in inflammatory cells in AP mice. The expression of CLDN3 mRNA was downregulated, which is consistent with scRNA-seq, however, its protein expression was upregulated. We found CLDN3 mainly expressed in the cytoplasm of pancreatic acinar cells of AP mice. These results indicated there might exist post-transcriptional mechanisms of CLDN3. Our findings, for the first time, indicated that CD44 and CLDN3 were crucial biomarkers associated with immune-metabolic dysregulation between pancreatic acinar cells and immune cells. The results of this study showed the potential of these two biomarkers as therapeutic targets for AP.