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The expression and role of inflammation-relatedcytokines/chemokines in acute lung injury induced by renal ischemia-reperfusion injury in neonatal ratsObjective:The purpose of this study was to investigate the expression and role of inflammation-related cytokines/chemokines in ALI induced by renal ischemia-reperfusion(I/R)injury in neonatal rats.Methods:Sprague-Dawley(SD)rats aged 10 days were randomly divided into three groups:Control group,Sham group and I/R group.The I/R group was further divided into ischemia 45min and reperfusion 1h,3h,6h,12h,24h and 48h.The Sham group only exposed kidney for 45min without ligation,just for 24h.The Control group did not undergo any treatment.Rats were sacrificed as scheduled.Kidney and lung tissues and blood were collected.Serum creatinine(sCr)concentration was detected by an automatic biochemical analyzer.Histopathological changes of kidney and lung tissues were detected by hematoxylin and eosin(HE)staining,and scores were evaluated.The protein expression levels of 27 inflammatory cytokines/chemokines in the kidney and lung tissues and serum were detected by multifunctional liquid chip technology,including interferon-induced protein 10(CXCL10),epidermal growth factor(EGF),eosinophils Cell activation chemokine(Eotaxin),chemokine(C-X-3-C Motif)ligand 1(Fractalkine),granulocyte colony-stimulating factor(G-CSF),granulocyte-macrophage colony-stimulating factor(GM-CSF),chemokine(C-X-C Motif)Ligand 1(GRO/KC/CINC-1),Interleukins(IL-1?,IL-1?,IL-2,IL-4,IL-5,IL-6,IL-10,IL-12p70,IL-13,IL-17A,IL-18),y-interferon(IFN-y),Leptin,Lipopolysaccharide-induced CXC chemokine(LIX),monocyte chemotactic protein-1(MCP-1),macrophage inflammatory protein-1?(MIP-1?),macrophage inflammatory protein-2(MIP-2),regulated upon activation normal T-cell express sequence(RANTES),tumor necrosis factor-?(TNF-?)and vascular endothelial growth factor(VEGF).The cytokine/chemokine that changed in the early stage of renal I/R and had obvious trend of changes among groups was selected,and the expression level of the cytokine/chemokine mRNA was further detected by quantitative real-time PCR(qRT-PCR).Results:1.Successfully established rat models of AKI-induced ALI:The sCr levels of rats in I/R groups showed a trend of increase and then decreased with the change of reperfusion time.It began to increase at I/R 3h,reached a peak at I/R 24h and decreased at I/R 48h.HE staining showed that there were obviously pathological injuries in the I/R 24h and 48h of kidney and lung tissues.Kidney and lung pathological injury scores in I/R 24h and 48h were significantly higher than those in Control group(P<0.001).The pathological scores of kidney I/R 48h group were lower than 24h group(P<0.05).But the lung I/R 48h group was more seriously damaged than 24h group,and the pathological scores between two groups were statistically different(P<0.001).2.The protein expression of inflammatory cytokines/chemokines in kidney and lung tissues and serum of renal I/R rats:Multifunctional liquid chip technology results showed that the expression levels of IL-10 and VEGF at I/R 3h were significantly increased compared with Control group(P<0.05);compared with Sham group,the expression levels of CXCL10,IFN-? and IL-6 were significantly increased(P<0.05);With the extension of reperfusion time,CXCL10 and IL-10 changed significantly among I/R groups(P<0.001).In lung tissues,EGF and MIP-1? at I/R 3h had a significantly higher level compared with Control group(P<0.05);IL-4 and VEGF at I/R 3h had a significantly higher level compared with Sham group(P<0.05);As the reperfusion time prolonged,CXCL10,IL-6,IL-10 and MCP-1 changed significantly among I/R groups(P<0.001).The expressions of CXCL10,Fractalkine,IL-5,IL-12p70,IL-18,LIX,MCP-1,RANTES could be detected in the serum.IL-5,IL-18,LIX,MCP-1 changed obviously among all groups(P<0.05).3.The protein expression of CXCL10 and IL-10 in kidney and lung tissues and serum:The expression levels of CXCL10 in kidney and lung tissues both increased first and then decreased.CXCL10 increased at I/R 3h,peaked at I/R 12h,and returned to control group level at I/R 24h.Compared with Control and sham groups,the expression levels of CXCL10 in two kinds of tissues significantly increased in the I/R 6h and 12h groups(P<0.05).The expression levels of CXCL10 in serum also increased first and then decreased.Serum CXCL10 increased at I/R 3h,peaked at I/R 6h,and then decreased to the level of Control group at I/R 12h.But there was no significant difference among all groups(P>0.05).The expression levels of IL-10 in kidney and lung tissues were highest at I/R 3h,as the reperfusion time extended,the expression level of IL-10 showed a downward trend,and returned to the level of Control group at I/R 24h.Compared with Control and Sham groups,the expression levels of IL-10 in kidney and lung tissues increased significantly in I/R 3h,6h and 12h groups(P<0.05);but the concentration of serum IL-10 was not detected.4.CXCL10 mRNA expression in kidney and lung tissues:The results of qRT-PCR showed that there was no significant difference in CXCL10 mRNA expression between Sham group and Control group in both kidney and lung tissues(P>0.05).Compared with Control group,the expression of CXCL10 mRNA was up-regulated at I/R 1h of kidney tissues(P<0.001),but at I/R 12h of lung tissues(P<0.05),and the expression levels of CXCL10 mRNA at I/R 24h were significantly higher than other I/R groups in both two kinds of tissues(P<0.001).Conclusion:1.Renal ischemia-reperfusion injury could induce acute lung injury in neonatal rats.2.The expression levels of chemokine CXCL10 were significantly increased early in kidney and lung tissues and serum in renal ischemia-reperfusion rats,and the expression of CXCL10 mRNA in kidney tissues were up-regulated earlier than that in lung tissues,suggesting that CXCL10 may play an important role in the process of acute lung injury induced by renal ischemia-reperfusion injury in neonatal rats.Part ? Correlation study of urinary CXCL10 with acute kidney injury,acute lung injury and mortality in critical ill childrenObjective:The results of the first part of animal experiments have suggested that the chemokine CXCL10 plays an important role in AKI and ALI in neonatal rats.The aims of this study were to investigate the association between uCXCL 10 and AKI,ALI and mortality in critically ill children.Methods:All patients admitted to PICU from September to December 2016 were eligible for this prospective study.We reviewed the medical records of eligible patients.The clinical status of critically ill children on the day of admission,medication,and therapies were recorded daily during PICU period.Acute respiratory distress syndrome(ARDS),sepsis,multiple organ dysfunction syndrome(MODS),shock,and disseminated intravascular coagulation(DIC)that developed during the PICU stay were diagnosed by the treating physicians.Length of PICU stay,duration of MV,length of hospital stay and mortality were also recorded.Pediatric Risk of Mortality ?(PRISM ?)score was calculated during the first 24h after PICU admission.Spot urine samples were collected during the first 24h after PICU admission and followed by serial collection every 48h during the PICU stay within the first week.The concentration of uCXCL10 was measured by using enzyme linked immunosorbent assay(ELISA).The initial and peak values of uCXCL10 within the first week after PICU admission were used for association analysis.The diagnosis of pediatric AKI was based on:?Serum creatinine(sCr)rise?26.5?mol/L within 48h;?sCr rise?1.5 times the baseline sCr within 7 days;?urine volume<0.5 ml/kg/h for 6h.AKI stage 1 was defined as mild AKI,and AKI stages 2 and 3 were defined as severe AKI.The diagnosis of ARDS was based on the criteria of the 2015 Pediatric Acute Respiratory Distress Syndrome:Consensus Recommendations of the Conference on Pediatric Acute Lung Injury.According to the occurrence and severity of AKI during PICU period,they were further divided into mild AKI group,severe AKI group and non-AKI group;according to the absence of ARDS and/or need for MV,they were divided into MV/ARDS(ARDS and/or MV)group and non-(MV/ARDS)(non ARDS and non MV)group;according to whether death occurred during hospitalization,they were divided into death group and survival group.Spearman analyses were performed to investigate factors potentially associated with the levels of uCXCL10.Generalized linear model analyses were performed to assess the association between uCXCL10 and AKI,ARDS,etc.Multivariate logistic regression analysis evaluated the relationship between uCXCL10 and mortality after adjustment for confounding factors.The predictive value of uCXCL10 for mortality was assessed by the receiver operating characteristic(ROC)curve and area under the curve(AUC)in critically ill children.Results:1.The prospective study involved 123 critically ill children.35(28.5%)developed AKI during the first week after admission,including 12(9.76%)who fulfilled the criteria for KDIGO stage 1,which is defined as mild AKI,and 14(18.70%)patients who fulfilled the criteria for KDIGO stages 2 and 3,which are defined as severe AKI.14(11.38%)developed ARDS,54(43.90%)required MV,and 57(46.34%)developed MV/ARDS during PICU stay.The mortality rate in the whole cohort was 15(12.20%)during hospitalization.The levels of uCXCL10 were no significant difference among the three groups of AKI in critically ill children(P>0.05).The peak uCXCL10 level in severe AKI group was higher than that in non-AKI group(P=0.026).The levels of uCXCL10 in MV/ARDS group were significantly higher than those of non-(MV/ARDS)group(initial:P=0.032;peak:P<0.001).Spearman correlation analysis showed that the initial and peak uCXCL10 levels were correlated with the age of critically ill children,PRISM ? score,sepsis,MODS,shock/DIC,MV,MV/ARDS,and mortality(P<0.05).In addition,the peak uCXCL10 level was also related to severe AKI,ARDS and duration of MV(P<0.05).The levels of uCXCL 10 in sepsis group were also significantly higher than those in non-sepsis(initial:P=0.001;peak:P<0.001).2.The results of generalized linear model analysis showed that the initial value of uCXCL10 was associated with sepsis(OR 1.478,95%CI 1.012-2.157,P=0.043)and MODS(OR 1.860,95%CI 1.193-2.900,P=0.006)after adjustment for confounding factors,and the peak value of uCXCL10 was associated with PRISM III score(OR 1.021,95%CI 1.001-1.040,P=0.039)and sepsis(OR 1.646,95%CI 1.127-2.406,P=0.010)after adjustment for confounding factors.3.The levels of uCXCL10 in the death group was significantly higher than that in the survival group in critically ill children(all P<0.001).Multivariate logistic regression analysis showed that the peak uCXCL10 level was still significantly correlated with mortality(OR=7.926,95%CI 1.706-36.817,P=0.008)after adjusting for age,weight and PRISM ? score.The initial and peak uCXCL 10 for predicting mortality displayed AUCs of 0.77(95%CI 0.66-0.88,P=0.001)and 0.86(95%CI 0.78-0.94,P<0.001),and PRISM ?score for predicting mortality displayed AUC of 0.856(95%CI 0.76-0.95,P<0.001)in critically ill children.Conclusion:1.The peak uCXCL10 was associated with severe AKI and ALI in critically ill children.After adjustment for confounding factors,the initial uCXCL10 was associated with MODS,the peak uCXCL10 was associated with sepsis and PRISM ? score.2.Both the initial and peak uCXCL10 had important values for predicting mortality in critically ill children.The predictive value of peak uCXCL10 for mortality was close to the PRISM ? score. |
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