| ObjectivePretreatment with low-dose lipopolysaccharide(LPS) induces a state of endotoxin tolerance,which was characterized by reduced inflammatory response upon subsequent high-dose LPS challenge.Polymicrobial sepsis induced by cecal ligation and puncture(CLP) reproduces many of the pathophysiologic features of septic shock and multiple organ dysfunction syndrome(MODS).The aim of the study is to investigate whether endotoxin tolerance has protective effects on CLP-induced multiple organ injury and whether these effects are associated with inhibition of NF-κB activity.Methods1,Polymicrobial sepsis was induced in male Sprague-Dawley rats via CLP. Endotoxin tolerance was reproduced by four consecutive daily intraperitoneal injections of 0.6 mg/kg LPS.2,Rats were anesthetized by intraperitoneal injection of sodium pentobarbital (40-50mg/kg),and then were randomized into three groups:the first group,rats received laparotomy only(sham group);a second group,rats received four consecutive daily intraperitoneal injections of saline and CLP on the 5th day(CLP group);a third group,rats received four consecutive daily intraperitoneal injections of 0.6 mg/kg LPS and CLP on the 5th day(ET+CLP group).Rats were killed at 0h(before CLP)and1,3,6,and 18 hrs after CLP.Blood and tissue samples(lungs, hearts,livers,and kidneys) were collected for histological examination,biochemical changes and evaluation of NF-κB.3,In the separate study,rats were randomly divided into two groups(CLP group and ET+CLP group,n=10/group),and survival rate was monitored for 7 days after CLP.Results1,CLP-induced polymicrobial sepsis was associated with high mortality rate, and endotoxin tolerance group had significantly higher survival rate in rats after CLP(24hrs survival rate 80%in ET+CLP group vs.40%in CLP group,7 days survival rate 40%in ET+CLP group vs.0%in CLP group,log-rank test p=0.021). 2,CLP-induced polymicrobial sepsis successfully induced multiple organ injury, especially at 18h after CLP,manifested by substantially increased plasma concentrations of TBIL,DBIL,ALT,AST(indicators of liver injury) and BUN,Cr (indicators of kidney injury).Endotxin tolerance group had significantly better liver and kidney function as compared with CLP group(plasma level of TBIL,DBIL,ALT, AST,BUN and Cr at 18h after CLP are 0.85±0.32mg/dL,0.57±0.23mg/dL,163.1±61.1u/L,310.8±120.5u/L,38.1±12.5 mg/dL,0.51±0.10mg/dL in CLP group vs. 0.29±0.08mg/dL,0.21±0.05mg/dL,95.3±20.1u/L,168.3±30.4u/L,22.5±5.9mg/dL,0.36±0.08mg/dL in ET+CLP group respectively,p＜0.05).3,Histopathological injuries were observed significantly in tissues of lung, liver and kidney of CLP group,characterized by inflammatory cells infiltration, profound cellular degeneration and severe vascular congestion.The histopathological changes of these tissues in the ET group were much milder than that of the CLP group.4,There was a significant increase in plasma concentrations of TNF-α,IL-6 and IL-10 in CLP group compared with sham group.Peak level of TNF-αoccurred at 6h after CLP(305.3±67.6 pg/mL),and peak level of IL-6 and IL-10 were reached at 18h after CLP(201.5±54.6 pg/mL and 95.7±38.5pg/mL respectively).Administration of endotoxin tolerance significantly reduced the cytokine production(plasma level of TNT-αwas 37.3±11.5pg/mL at 6h after CLP,plasma level of IL-6 and IL-10 were 84.2±38.5pg/mL and 35.2±11.8pg/mL at 18h after CLP,p＜0.05).5,CLP resulted in the early activation of NF-κB in lung tissue with the peak activity at 3 hrs after CLP.In contrast,endotoxin tolerance group had significantly lower activity of NF-κB at 3h and 6h after CLP(p＜0.05).ConclusionsThese data revealed that endotoxin tolerance could exert protective effects on sepsis-induced multiple organ injury.These beneficial effects appear partly due to the down-regulation of NF-κB activation and thereby attenuating the production of inflammatory cytokines.This suggests that endotoxin tolerance may be considered a potential protective approach in sepsis.
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