Spermidine Prevents Polarity Loss of Absorptive Enterocytes in Jejunum of Lipopolysaccharide-Challenged Mice via 4D-DIA Proteomics Analysis

Identifying effective compounds to restore the polarity of absorptive enterocytes (AEs) holds promise for mitigating the severity and duration of small intestinal disorders. Spermidine (SPD) is a natural polyamine; whether it can repair inflammation-induced loss of AE polarity remains unclear. In this study, we employed lipopolysaccharide (LPS)-challenged mice models combined with 4D data-independent acquisition (DIA) proteomics to investigate the mechanisms by which SPD alleviates polarity loss in AEs. Our results demonstrated that SPD supplementation enhanced the antioxidant capacity and improved the villus/crypt ratio in the jejunum of LPS-treated mice. Proteomic analysis revealed that LPS induced acute phase and inflammatory responses, significantly downregulating the expression of cytoskeletal proteins (Pdlim3, Pdlim7) essential for epithelial morphology as well as proteins involved in apical-basal polarity (Pard6b, Pard3, Prkcz, LLGL2), apical membrane integrity (Vil1, Pdims, Akp3, Tjps, Pards), and apical SLC transporters. Conversely, SPD attenuated mucosal- and tissue-specific immune responses and reversed the downregulation of these protein groups. Furthermore, using a Caco-2 cell model, we confirmed the anti-inflammatory effect of SPD and elucidated its role in suppressing AE polarity loss via the regulation of HDAC4 signaling. These findings indicate that SPD effectively alleviates the inflammation-induced loss of AE polarity in the jejunum of LPS-challenged mice.