Objective. Vitronectin (VTN) has been reported to trigger cell pyroptosis to aggravate inflammation in our previous study. However, the function of VTN in inflammatory bowel disease (IBD) remains to be addressed. Methods. Real-time PCR and western blotting were performed to analyze VTN-regulated intestinal epithelial cell (IEC) differentiation through ferroptosis, and immunofluorescence (IF), luciferase, and chromatin immunoprecipitation were used to identify whether VTN-modulated ferroptosis is dependent on phosphodiesterase 4 (PDE4)/protein kinase A (PKA)/cyclic adenosine monophosphate-response element-binding protein (CREB) cascade pathway. In vivo experiment in mice and a pilot study in patients with IBD were used to confirm inhibition of PDE4-alleviated IECs ferroptosis, leading to cell differentiation during mucosal healing. Results. Herein, we found that caudal-related homeobox transcription factor 2-mediated IECs differentiation was impaired in response to VTN, which was attributed to enhanced ferroptosis characterized by decreased glutathione peroxidase 4 (GPX4) and solute carrier family 7 member 11 expression. Inhibition of ferroptosis in IECs rescued the inhibitory effect of VTN on cell differentiation. Further analysis showed that VTN triggered phosphorylation of PDE4, leading to inhibit PKA/CREB activation and CREB nuclear translocation, which further reduced GPX4 transactivation. Endogenous PKA interacted with CREB, and this interaction was destroyed in response to VTN stimulation. What is more, overexpression of CREB in CaCO₂ cells overcame the promotion of VTN on ferroptosis. Most importantly, inhibition of PDE4 by roflumilast or dipyridamole could alleviate dextran sulfate sodium-induced colitis in mice and in a pilot clinical study confirmed by IF. Conclusions. These findings demonstrated that highly expressed VTN disrupted IECs differentiation through PDE4-mediated ferroptosis in IBD, suggesting targeting PDE4 could be a promising therapeutic strategy for patients with IBD.

中文翻译：

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玻连蛋白通过激活 PDE4 介导的铁死亡来破坏炎症性肠病中肠上皮细胞的分化

客观的。在我们之前的研究中，有报道称玻连蛋白（VTN）会引发细胞焦亡，从而加剧炎症。然而，VTN 在炎症性肠病 (IBD) 中的作用仍有待解决。方法。进行实时 PCR 和蛋白质印迹分析 VTN 调节的肠上皮细胞 (IEC) 通过铁死亡的分化，并使用免疫荧光 (IF)、荧光素酶和染色质免疫沉淀来鉴定 VTN 调节的铁死亡是否依赖于磷酸二酯酶 4。 PDE4)/蛋白激酶A (PKA)/环磷酸腺苷反应元件结合蛋白(CREB)级联途径。小鼠体内实验和 IBD 患者的初步研究被用来确认 PDE4 减轻 IEC 铁死亡的抑制作用，从而导致粘膜愈合过程中的细胞分化。结果。在此，我们发现尾部相关的同源框转录因子 2 介导的 IEC 分化因 VTN 的反应而受损，这归因于以谷胱甘肽过氧化物酶 4 (GPX4) 和溶质载体家族 7 成员 11 表达减少为特征的铁死亡增强。IEC 中铁死亡的抑制挽救了 VTN 对细胞分化的抑制作用。进一步分析表明，VTN触发PDE4磷酸化，导致抑制PKA/CREB激活和CREB核转位，从而进一步减少GPX4反式激活。内源性 PKA 与 CREB ​​相互作用，这种相互作用在 VTN 刺激后被破坏。_{更重要的是，CaCO 2}细胞中CREB的过表达克服了VTN对铁死亡的促进作用。最重要的是，罗氟司特或双嘧达莫抑制 PDE4 可以减轻小鼠中葡聚糖硫酸钠诱导的结肠炎，这一点在 IF 的一项初步临床研究中得到证实。结论。这些发现表明，IBD 中高表达的 VTN 通过 PDE4 介导的铁死亡破坏了 IECs 分化，这表明针对 PDE4 可能是 IBD 患者的一种有前景的治疗策略。