Abstract：
During recovery from septic shock, circulating mitochondrial N-formyl peptides (mtFPs) predispose to secondary infection by occupying formyl peptide receptor 1 (FPR1) on the neutrophil (polymorphonuclear leukocyte, PMN) membrane, suppressing cytosolic calcium ([Ca2+]i)-dependent responses to secondarily encountered bacteria. However, no study has yet investigated therapeutic clearance of circulating mtFPs in clinical settings. Thus, we studied how to remove mtFPs from septic-shock plasma and whether such removal could preserve cell-surface FPR1 and restore sepsis-induced PMN dysfunction by normalizing [Ca2+]i flux. In in vitro model systems, mtFP removal rescued PMN FPR1-mediated [Ca2+]i flux and chemotaxis that had been suppressed by prior mtFP exposure. However, PMN functional recovery occurred in a stepwise fashion over 30 - 90 minutes. Intracellular Ca2+-calmodulin appears to contribute to this delay. In ex vivo model systems using blood samples obtained from patients with septic shock, anti-mtFP antibodies alone failed to eliminate mtFPs from septic-shock plasma or inhibit mtFP activity. We therefore created a beads-based anti-mtFP antibody cocktail (bb-AMfpA) by combining protein A/sepharose with antibodies specific for the most potent human mtFP chemoattractants. The bb-AMfpA treatment successfully removed those active mtFPs from septic-shock plasma. Furthermore, the bb-AMfpA treatment significantly restored chemotactic and bactericidal dysfunction of PMNs obtained from patients with septic shock who developed secondary infections. By clearing circulating mtFPs, the immobilized anti-mtFP antibody therapy prevented mtFP interactions with surface FPR1, thereby restoring [Ca2+]i-dependent PMN antimicrobial function in clinical septic-shock environments. This approach may help prevent the development of secondary, nosocomial infections in patients recovering from septic shock.
摘要：
在脓毒性休克恢复期间,循环线粒体N-甲酰肽（mtFP）通过占据中性粒细胞（多形核白细胞，PMN)膜，抑制胞质钙（[Ca2]i）依赖性对次生细菌的反应。然而,尚无研究调查临床环境中循环mtFP的治疗清除情况.因此，我们研究了如何从脓毒性休克血浆中去除mtFP,以及这种去除是否可以通过使[Ca2+]i通量正常化来保留细胞表面FPR1和恢复脓毒症诱导的PMN功能障碍.在体外模型系统中，mtFP去除挽救了先前mtFP暴露抑制的PMNFPR1介导的[Ca2]i通量和趋化性。然而,PMN功能恢复在30-90分钟内以逐步方式发生。细胞内Ca2-钙调蛋白似乎有助于这种延迟。在体外模型系统中，使用从感染性休克患者获得的血液样本，单独的抗mtFP抗体不能消除感染性休克血浆中的mtFP或抑制mtFP活性.因此，我们通过将蛋白A/琼脂糖与对最有效的人mtFP化学引诱物具有特异性的抗体组合，创建了基于珠子的抗mtFP抗体混合物（bb-AMfpA）。bb-AMfpA治疗成功地从脓毒性休克血浆中去除那些活性mtFP。此外，bb-AMfpA治疗显著恢复了从发生继发感染的脓毒性休克患者获得的PMN的趋化和杀菌功能障碍.通过清除循环的mtFP，固定化抗mtFP抗体治疗阻止了mtFP与表面FPR1的相互作用,从而恢复了临床感染性休克环境中[Ca2+]i依赖性PMN的抗菌功能.这种方法可能有助于防止继发性疾病的发展，感染性休克患者的医院感染。
