The Study Of N-acytyl-seryl-aspartyl-lysyl-proline In Children With Common Types Of Acute Kidney Injury

N-acetyl-seryl-aspartyl-lysyl-proline(AcSDKP)is an endogenous tetrapeptide.Animal experiments showed that AcSDKP could be secreted by renal tissue,mainly distal renal tubules.Previous studies have found that endotoxin and hypoxia can reduce the activity of the angiotensin-converting enzyme(ACE)in renal tissue,and AcSDKP is mainly degraded through the N-terminal of ACE.The level of urinary AcSDKP/urinary creatinine(uAcSDKP)may be increased due to the catabolic disorder of AcSDKP in renal tissue,which can be used as a biomarker to monitor and predict clinical AKI.Clinical studies have found that the predictive effectiveness of uAcSDKP for predicting diabetic nephropathy in patients with normal proteinuria after six months is similar to that of urinary kidney factor-1,which is significantly better than plasma AcSDKP and urine β2-microglobulin.However,uAcSDKP as a biomarker of AKI has not been reported.Cardiopulmonary bypass cardiac surgeryrelated AKI and sepsis-related AKI are common types of AKI in the clinic.During cardiopulmonary bypass cardiac surgery,a series of non-physiological changes,including hemodynamic changes,will lead to renal injury,but the degree of renal injury is different.AKI is the most common organ injury in sepsis.Therefore,the clinical experiment part of this study will explore the feasibility of uAcSDKP in monitoring and predicting clinical AKI by analyzing the relationship between uAcSDKP and AKI in cardiac surgery cases and sepsis cases.The significance of the uAcSDKP elevation during AKI is still unclear.Previous studies have shown that excessive renal tubular epithelial cell apoptosis caused by various pathogenic factors is one of the pathogenesis of AKI.AcSDKP is involved in regulating cell proliferation and apoptosis,but its outcome varies with different diseases and cell types,and its effect on renal tubular epithelial cell apoptosis has not been reported.Fibroblast growth factor receptor 1(FGFR1)signal pathway is considered to be one of the ways for AcSDKP to exert its biological effects.In this study,we observed the changes of renal tubular epithelial cell apoptosis and FGFR1 expression after treatment with different concentrations of AcSDKP and discussed the effects of AcSDKP on renal tubular epithelial cell apoptosis and FGFR1 expression under normal culture conditions.Part One Early diagnostic value of uAcSDKP in acute kidney injury associated with cardiac surgery under cardiopulmonary bypassObjective:By monitoring the trend of changes in urinary N-acetyl-seryl-aspartyl-lysylproline/creatinine(uAcSDKP)before and after cardiopulmonary bypass heart surgery,it is explored whether uAcSDKP can effectively reflect the changes of renal function and its effectiveness in the early prediction of acute kidney injury(AKI).Methods:Continuously enrolled children undergoing cardiopulmonary bypass surgery in the surgical intensive care unit(SICU)of the author’s hospital from July 2018 to December 2019.Their urine was taken before the operation,immediately after the operation,24 hours and 48 hours after operation to detect the concentration of AcSDKP and urinary Nacetyl-β-D-glucosaminidase/creatinine(uNAG),and clinical data were collected.The changes of uAcSDKP,uNAG,serum cystatin C(sCysC)and serum creatinine(sCr)levels before and after operation were analyzed to evaluate the feasibility of uAcSDKP as a biomarker of renal injury.According to whether AKI occurred after the operation,the selected cases were divided into non-AKI group and AKI group,and the risk factors of AKI after operation were analyzed by logistic regression.The area under the curve(AUC)of the receiver operator characteristic curve(ROC)was used to evaluate the efficacy of uAcSDKP in early predicting postoperative AKI,and whether the combination with other markers of renal injury can improve the predictive performance was studied.And the risk factors of postoperative uAcSDKP increase were also analyzed.Result:Cardiopulmonary bypass is the key factor of AKI related to cardiopulmonary bypass cardiac surgery.The duration of cardiopulmonary bypass is an independent risk factor for the onset of AKI and the elevation of uAcSDKP immediately after operation.The immediately postoperative uAcSDKP was 117.85(57.74,211.29)nmol/gCr,which was significantly higher than the preoperative uAcSDKP level of 34.53(26.77,58.13)nmol/gCr(P<0.001).At 24 hours after the operation,uAcSDKP was 59.33(36.20,94.66)nmol/gCr,which was significantly lower than that of immediately after operation(P<0.001),but still higher than the preoperative level(P=0.010).The level of uAcSDKP decreased by 43.57(25.04,55.13)nmol/gCr 48 hours after the operation,which had no significant difference from the preoperative level(P=0.144).The level of uAcSDKP reached its peak immediately after cardiac surgery,which was more than 3 times the preoperative level,and the immediate postoperative uAcSDKP level in the AKI group was 6 times the preoperative level.The level of sCr and sCysC peaked at 24 hours after operation,which were 1.2 times and 1.3 times the preoperative level,respectively.While the level of sCr in the AKI group at 24 hours after operation was only 1.5 times the preoperative level.Multivariate logistic regression analysis showed that immediate postoperative uAcSDKP and sCysC were independent risk factors for AKI related to cardiopulmonary bypass.The efficacy of the immediate postoperative uAcSDKP in predicting cardiopulmonary bypass cardiac surgery-related AKI was 0.77(95%CI:0.667-0.864,P<0.001),which was similar to that of the immediate postoperative sCysC((95%CI:0.536-0.761,P=0.018)(P=0.211),but the specificity was higher(80.43%vs.62.22%).The area under the ROC curve of uAcSDKP and sCysC immediately after the operation in predicting AKI related to cardiopulmonary bypass was significantly increased to 0.828(95%CI:0.724-0.905,P<0.001).Conclusion:uAcSDKP can effectively monitor the changes of renal function after cardiopulmonary bypass.Compared with sCr and sCysC,uAcSDKP can reflect changes in renal function early.The predictive performance of the immediate postoperative uAcSDKP for cardiopulmonary bypass cardiac surgery-related AKI is similar to that of the immediate postoperative sCysC,but with higher specificity.The combination of sCysC and uAcSDKP immediately after surgery can significantly improve the prediction efficiency of AKI.Part Two Study on the predictive value of uAcSDKP for sepsis-related acute kidney injuryObjective:By analyzing the relationship between the level of uAcSDKP when children with sepsis enter the pediatric intensive care unit(PICU)and sepsis-related AKI,the predictive value of uAcSDKP for sepsis-related AKI is explored.Methods:Eligible children with sepsis admitted to the PICU of the author’s hospital from July 2018 to December 2019 were included continuously,and their clinical data were collected.The urine of children admitted to PICU was tested for AcSDKP concentration.The uAcSDKP level was assessed as a categorical variable according to quartiles,in which the lowest quartile was used as the reference value.Two models were used to adjust the dependent variables.Logistic regression analysis was used to analyze the relationship between uAcSDKP and sepsis-related AKI or severe AKI(SAKI).According to the occurrence of sepsis-related AKI or SAKI,the patients were divided into non-AKI group and AKI group,non-SAKI group and SAKI group.The risk factors if sepsis-related AKI and sepsis-related SAKI were analyzed by logistic regression.The area under the ROC curve was used to evaluate the effectiveness of uAcSDKP in predicting sepsis-related AKI and SAKI.Results:Of all the 131 cases selected,62(50.38%)developed sepsis-related AKI,including 19(30.65%)in stage 1,19(30.65%)in stage 2 and 24(38.70%)in stage 3.According to the quartile of uAcSDKP,the cases were divided into Q1 group,Q2 group,Q3 group and Q4 group.After adjusting for clinical characteristics(gender,age,shock,PRISMⅢ score)and baseline renal function(sCysC and sCr>baseline),the risk of sepsisrelated AKI in the Q3 and Q4 group was 5 and 8 times higher than that in the Q1 group,and the risk of sepsis-related SAKI was 3 and 19 times higher than that in Q1 group,respectively.Multivariate logistic regression analysis showed that uAcSDKP,uNAG and sCysC were the risk factors of sepsis-related AKI.The area under the ROC curve of uAcSDKP for predicting sepsis-related AKI was 0.791(95%CI:0.717-0.857,P<0.001),and the optimal critical value was 65.54 nmol//gCr.There was no significant difference between uAcSDKP and uNAG 0.739(95%CI:0.655-0.812,P<0.001)and sCysC(95%CI:0.736-0.877,P<0.001)(P>0.05):The area under the ROC curve of sepsis-related AKI predicted by uAcSDKP and sCysC combined was 0.891(95%CI:0.824-0.939,P<0.001),which was 0.10(95%CI:0.043-0.157,P<0.001)higher than that predicted by uAcSDKP alone.Multivariate logistic regression analysis showed that uAcSDKP,positive renal angina index and sCysC were the risk factors of sepsis-related SAKI.The area under the ROC curve of uAcSDKP for predicting sepsisrelated SAKI was 0.790(95%CI:0.759-0.856,P<0.001),and the optimal critical value was 115.20 nmol//gCr.There was no significant difference among uAcSDKP,sCysC 0.834(95%CI:0.759-0.893,P<0.001)and positive renal angina index 0.769(95%CI:0.688-0.838,P<0.001)in predicting sepsis-related SAKI(P>0.05).Combined with uAcSDKP,sCysC and renal angina index,the area under the ROC curve of sepsis-related SAKI can be increased to 0.914(95%CI:0.852-0.956,P<0.001).Conclusion:Elevated uAcSDKP at PICU admission is a risk factor for sepsis-related AKI and SAKI,and can effectively predict the onset of AKI and SAKI in children with sepsis.Part Three The effect of N-acetyl-seryl-aspartyl-lysyl-proline on the apoptosis of renal tubular epithelial cellsObjective:To study the effect of AcSDKP on the apoptosis and FGFR1 expression of renal tubular epithelial cells.Methods:Human renal tubular epithelial cell line(HK-2 cells)were cultured in the 6well/96-well culture plate for 24 hours,and about 80%of the cells were fused,followed by intervention.Serum-free mediums with different concentrations of AcSDKP(0、10、102、103、104、105nM)were added respectively.After 24 hours of incubation,the proliferation and apoptosis of renal tubular epithelial cells and the expression changes of fibroblast growth factor receptor 1(FGFR1),Bax and Bcl-2 were detected.Result:There was no significant difference in HK-2 cell activity between each group after treatment with different concentrations of AcSDKP(0、10、102、103、104、105nM)for 24 hours and 48 hours(P>0.05).After treatment with different concentrations of AcSDKP(0、10、102、103、104、105nM)for 24 hours,there was no significant difference in the proportion of HK-2 cell apoptosis and the protein expression levels of Bax and Bcl-2 between each group(P>0.05);however,the mRNA and protein expression levels of FGFR1 increased with the concentration of AcSDKP.When the concentration of AcSDKP was 104 nM,the mRNA and protein expression levels of FGFR1 increased significantly(P<0.05).Conclusion:AcSDKP did not affect the proliferation and apoptosis of renal tubular epithelial cells but induced the expression of FGFR1 in renal tubular epithelial cells in a concentration-dependent manner.When the concentration of AcSDKP was 104 nM,the expression of FGFR1 protein and mRNA in HK-2 cells increased significantly.