Substance P-regulated Leukotrien B4 Production Promotes Acute Pancreatitis-associated Lung Injury

Background and PurposeLeukotriene B4?LTB4?is a potent chemoattractant for immune cells,including neutrophils and macrophages,which play a major role in inflammatory responses.Substance P?SP?is a neuropeptide mainly involved in nociception and neurogenic inflammation.Research increasingly suggests that SP contributes to inflammation in acute pancreatitis?AP?and associated acute lung injury?ALI?,but the molecular mechanism remains unclear.Substance P?SP?has been reported to promote the production of LTB4 in vivo and in vitro.Here,we investigated the role of LTB4 in AP,AP-associated ALI and the related mechanisms of LTB4 production in AP.MethodIn vivo,murine AP model was induced by caerulein and lipopolysaccharide?LPS?or L-arginine?L-Arg?.The specific BLT1 antagonist LY293111 or the selective NK1R antagonist CP96345 was injected intraperitoneally 0.5 h before the first injection of caerulein or L-Arg.Change of LTB4/BLT1 system was analysed.The severity of AP was assessed by amylase and lipase levels and histopathological scoring.The local inflammation of AP was assessed by the infiltration of Ly6G⁺neutrophils and F4/80⁺macrophages and the mRNA levels of inflammatory cytokines?IL-6 and IL-1??in the pancreatic tissue.ALI was assessed by histopathological scoring,Ly6G⁺neutrophils infiltration and the mRNA levels of inflammatory cytokines?IL-6,IL-1?and TNF-??in lung.The proportion of reverse migrated neutrophils,which have a distinctive ICAM-1^highCXCR1^low phenotype,was detected by flow cytometry.In vitro,pancreatic acinar cells?PACs?were stimulated with SP and the supernatants were collected for LTB4 measurement.The PKC?/MAPK signaling pathway was evaluated.ResultsThe levels of LTB4 and its specific receptor BLT1 were markedly upregulated in both AP models.Blockade of BLT1 by LY293111 attenuated the severity of AP,decreased neutrophil reverse transendothelial cell migration?rTEM?into the circulation and alleviated the severity of ALI.Treatment of PACs with SP increased LTB4production.Furthermore,SP treatment increased phosphorylation of protein kinase C?PKC??and mitogen activated protein kinases?MAPKs?,including extracellular signal-regulated kinase?ERK?,p-38 MAPK and c-Jun NH2-terminal kinase?JNK?.Finally,blockade of NK1R by CP96345 significantly attenuated the severity of AP and decreased LTB4 production.ConclusionSubstance P regulates LTB4 production in AP via PKC?/MAPK pathway;LTB4increases local inflammation of AP by attracting neutrophils and macrophages and promotes AP-associated lung injury through neutrophil reverse migration.