Migration of neutrophils across endothelial barriers to capture and eliminate bacteria is served as the first line of innate immunity. Bacterial virulence factors damage endothelium to produce inflammatory cytokines interacts with neutrophils. However, the mechanisms that behind endothelial-neutrophil interaction impact on the bactericidal activity remain unclear. Therefore, we aimed to find the target proteins on endothelial cells that triggered the bactericidal activity of transendothelial neutrophils. Herein, we built the infected models on rats and endothelial-neutrophil co-cultural system (Transwell) and discovered that endothelial-derived IL-1α promoted the survival of rats under Escherichia coli infection and enhanced the bactericidal activity of transendothelial neutrophils in vivo and in vitro. Results further showed that IL-1α was inhibited by lipopolysaccharide (LPS) in the endothelial-neutrophil interaction. We found that LPS mainly damaged cell membrane and induced cell necrosis to interrupt neutrophil migration from endothelial barrier. Thus, we used the isobaric tags for relative and absolute quantification (iTRAQ) method to identify different proteins of endothelial cells. Results showed that IL-1α targeted cellular plasma membrane, endoplasmic reticulum and mitochondrial envelope and triggered eleven common proteins to persistently regulate. During the early phase, IL-1α triggered the upregulation of cell adhesion molecules (CAMs) to promote neutrophil adhesion, while oxidative phosphorylation was involved in long time regulation to induce transmigration of neutrophils against bacteria. Our results highlight the critical mechanism of endothelial-derived IL-1α on promoting bactericidal activity of transendothelial neutrophils and the findings of IL-1α triggered proteins provide the potentially important targets on the regulation of innate immunity.

中性粒细胞穿过内皮屏障迁移以捕获和消除细菌是先天免疫的第一道防线。细菌毒力因子损伤内皮产生炎症细胞因子与中性粒细胞相互作用。然而，内皮-中性粒细胞相互作用影响杀菌活性的机制仍不清楚。因此，我们的目的是寻找内皮细胞上触发跨内皮中性粒细胞杀菌活性的靶蛋白。在此，我们建立了大鼠和内皮-中性粒细胞共培养系统（Transwell）的感染模型，发现内皮源性IL-1α促进了大鼠的存活。大肠杆菌感染并增强跨内皮中性粒细胞的杀菌活性体内和体外。结果进一步表明，IL-1α在内皮-中性粒细胞相互作用中受到脂多糖（LPS）的抑制。我们发现LPS主要破坏细胞膜并诱导细胞坏死，从而中断中性粒细胞从内皮屏障的迁移。因此，我们使用同量异位标签进行相对和绝对定量（iTRAQ）方法来鉴定内皮细胞的不同蛋白质。结果表明，IL-1α靶向细胞质膜、内质网和线粒体包膜，并触发11种常见蛋白持续调节。在早期阶段，IL-1α触发细胞粘附分子（CAM）上调，促进中性粒细胞粘附，而氧化磷酸化参与长时间调节，诱导中性粒细胞针对细菌的迁移。我们的结果强调了内皮源性 IL-1α 促进跨内皮中性粒细胞杀菌活性的关键机制，并且 IL-1α 触发蛋白的发现为先天免疫调节提供了潜在的重要靶标。