| Part one:Multiple biomarkers on early diagnosis of acute kidney injury after cardiovascular surgery and the effects of dexmedetomidine on early postoperative renal injuryObjection:To investigate multiple biomarkers on early diagnosis of acute kidney injury(AKI)after cardiopulmonary bypass(CPB)in patients undergoing cardiovascular surgery,and observe the effects of dexmedetomidine on renal injury during early period after surgery.Methods:Part 1:A total of 200 patients aged 22?86 yr weighting 46?87 kg scheduled forcardiovascular surgery under CPB were enrolled in this stdy.Patiets received routine anesthesia care.The occurrence of acute kidney injury(AKI)was recorded.The level of serum creatinine(Scr)and urine neutrophil gelatinase-associated lipocalin(NGAL),cystatin C(Cys C),tissue inhibitor of metalloproteinase-2(TIMP-2),insulin-like growth factor binding protein-7(IGFBP-7)were determined 1d before and 0,2,6,12,24,48,72h after operation.The value of[TIMP-2]×[IGFBP-7]was calculated.The scores of sequential organ failure assessment(SOFA)and acute physiology and chronic health evaluation scoring system ?(APACHE ?)were evaluated Id before and 0,24,48,72h after surgery.NGAL,Cys C,[TIMP-2]×[IGFBP-7]were assessed for it's predictive AKI using Receiver Operating Characteristic(ROC)curves.Part 2:A total of 60 ASA ? or ? patients aged 23?64 yr weighting 48?85 kg with New York Heart Association(NYHA)class ? or ? scheduled for cardiovascular surgery under CPB were randomly divided into 2 groups(n=30 each):control group and dexmedetomidine(DEX group).The patients in DEX group were intravenously administrated 0.5 ?g/kg dexmedetomidine 10 min before induction of general anestnesia and followed by the dose of 0.2?0.5?g·kg-1·h-1 till the end of operation.The patients in control group received equal volume of normal saline at the same way.The occurrence of AKI was recorded.The level of Scr and urine NGAL,Cys C,TIMP-2,IGFBP-7 were determined Id before and 0,2,6,12,24,48,72h after operation.The value of[TIMP-2]×[IGFBP-7]was calculated.Results:Part 1:According to AKI critieria of Kiney Deisease Improving Global Outment(KDIGO)clinical practice guidelines,the subjects were divided into the AKI group(41 cases)and non-AKI group(159 cases).There was no significant difference of emographic data between the two groups(P>0.05).CPB time,aortic occlusion time and operative time were longer in AKIgroup than those in non-AKI group(P<0.05).A total of 41 patients developed AKI with incidence of 20.5%,of which 21 cases developed stage 1 of AKI(the incidence was 51.2%),14 cases developed stage 2(34.1%),6 cases developed stage 3(14.6%).The scores of SOFA and APACHE ? in two groups elevated significantly Oh after surgery and declined graduately later.The scores of SOFA and APACHE ? in AKI group were significantly higher than those in non-AKI group 0,24,48,72h after operation(P<0.05).The level of urine NGAL,Cys C and[TIMP-2]×[IGFBP-7]elevated early until 72h after surgery in AKI group.The level of urine biomarkers elevated early after surgery,declined graduately,and dropped down to preoperrativelevels 24?48h after surgery.Compared with non-AKI group,the level of NGAL,Cys C and[TIMP-2]×[IGFBP-7]increased 0?72h after operation in AKI group(P<0.05).At 0,2,6,12h after operation,the area of under ROC curves(AUC)of urine NGAL were 0.689,0.709,0.713 and 0.803,respectively,the AUC of urine Cys C were 0.639,0.762,0.774 and 0.812,respectively,and the AUC of urine[TIMP-2]×[IGFBP-7]were 0.687,0.721,0.740 and 0.779,respectively.The AUC for parallel tests of these three indicators were 0.694,0.773 and 0.794 at 0,2,6h after operation,respectively.The AUC for serial tests of these three indicators were 0.610,0.631 and 0.667 at 0,2,6h after operation,respectively.There were siginificantly differences in the value of diagnosis for single or joint detection of these three indicators(P<0.05).Sensitivity of AKI diagonosis of parallel tests was higher than that of single indicator,which was higher than that of serial tests.Part 2:There was no significant difference of emographic data between the two groups(P>0.05).According to KDIGO guidelines on AKI,6 cases(20.0%)developed AKI in control group and 3 cases developed AKI in DEX group.There was no significant difference between AKI occurrence in the two groups(P>0.05).There was no significant difference in the level of Scr and urineNGAL,Cys C,[TIMP-2]×[IGFBP-7]1d before operation between the two groups(P>0.05).Compared with 1d before operation,the levels of Scr were no changes after operation in the two groups(P>0.05);the levels of urine NGAL,Cys C and[TIMP-2]×[IGFBP-7]increased 0?6h after operation,decerased graduately 12?24h after operation and dropped to preoperative levels 48?72h after operation.Compared with the control group,the levels of urine NGAL and Cys C decreased 2?24h after operation,and the levels of urine[TIMP-2]×[IGFBP-7]dereased 0?24h after operation in DEX group(P<0.05).Conclusion:Urine NGAL,Cys C and[TIMP-2]×[IGFBP-7]are sensitive in early predicting AKI after cardiovascular surgery under CPB,joint detection of these three index can improve the predictive value.Dexmedetomidine can reduce the level of urine NGAL,Cys C and[TIMP-2]×[IGFBP-7],inhibit renal injury early after cardiovascular surgery.Part two:The effects of dexmedetomidine on microcirculatory perfusion injury induced by renal I/R via quantification with contrast-enhanced ultrasoundObjective:To observe the changes of microcirculatory perfusion in renal ischemia/reperfusion(I/R)injury rabbit by quantitative analysis of contrast-enhanced ultrasound(CEUS),and investigate the effects of dexmedetomidine on microcirculatory perfusion.Methods:A total of 24 New Zealand rabbits were randomly divided into 3 groups(8 rabbits in each group):control group,renal ischemia/reperfusion injury group(group I/R)and dexmedetomidine group(group DEX).The right kidneys were resected and I/R injury model of the left kidneys were established in rabbits of group I/R and DEX.A dose of 10 ?g/kg dexmedetomidine was administrated by intraperitoneal injection 30 min before renal ischemia in the DEX group.At 24h after reperfusion,renal size was measured by two-dimensional ultrasound and renal artery resistance(RI)was measured by color Doppler flow imaging(CDFI).Renal cortex perfusion was observed by CEUS.The value of time to peak(TTP),peak signal intensity(PSI),gradient between start frame to peak frame(Grad)and area under the curve(AUC)were analyzed by the time-intensition curves(TIC).The level of serum creatinine(Scr),plasma urea,serum neutrophil gelatinase-associated lipocalin(NGAL)and cystatin C(Cys C),urine NGAL and Cys C were measured immediately After CEUS.Pathological changes of renal tissue were observed.The level of intercellular cell adhesion molecule-1(ICAM-1)and vascular cell adhesion molecule-1(VCAM-1)in renal cortex were measured.Linear correlation between TIC parameters and ICAM-1,VCAM-1 were analized.Results:Compared with the control group,renal size and the pathological morphology changed remarkably,RI,PPT and AUC increased,PSI and Grad decreased,the level of Scr,plasma urea,serum and urine NGAL,Cys C increased,the level of ICAM-1 and VCAM-1 in renal cortex increased significantly in group I/R and DEX(P<0.05).Compared with I/R group,I/R-induced changes of renal size and pathological morphology were smaller,RI,TTP and AUC decreased,PSI and Grad increased,the level of Scr,plasma urea,serum and urine NGAL,Cys C decreased,the level of ICAM-1 and VCAM-1 in renal cortex decreased significantly in group DEX(P<0.05).The value of TTP,as wells as the other TIC parmeters PSI and Grad,was found to correlated with renal cortical ICAM-1 and VCAM-1 level(P<0.05).Conclusion:CEUS combined with TIC parameters may quantitatively and dynamically analyze the changes of microcirculatory perfusion in renal I/R injury.Dexmedetomidine could protect against renal microcirculatory perfusion injury.Part three:Dexmedetomidine attenuates renal ischemia-reperfusion injury by regulating deacetylase sirtuin 3 Objective:To investigate the protective effects of dexmedetomidine against renal ischemia andreperfusion(I/R)injury by activating deacetylase sirtuin(SIRT)3 and its mechanisms.Methods:Human renal proximal tubular epithelial line HK2 cells were randomly devided into three groups:Control group,hypoxia/reoxygenation(H/R)group and dexmedetomidine(DEX)group(0.001nM,0.01nM and 0.1nM).Cells in H/R group was induced by 24 h of hypoxia(5%CO2,1%O2,and 94%N2)followed by 3h of reoxygenation(5%CO2,21%O2,and 74%N2).Cells in DEX group were incubated with dexmedetomidine 60 min prior to H/R exposure.Cells served as control that incubated under normoxic conditions.SIRT3-targeted Small interfering RNA(siRNA)and SIRT3 plasmid were delivered by transfection reagent to HK2 cells before hypoxia.C57BL/6J mice were randomly devided into three groups:Sham group,ischemia and reperfusion(I/R)group and dexmedetomidine(DEX)group.I/R model was induced by bilateral renal pedicle clamping for 45 min followed by 24 h of reperfusion.Animals in DEX group were treated with dexmedetomidine(25 ?g/kg,i.p.)30 min prior to renal I/R.Animals in Sham group served as control that underwent the open-abdominal surgery only.SIRT3-targeted siRNA was delivered by transfection reagent to animals 3d before renal ischemia.Cell death and mitochondrial membrane potential(??m)of HK2 were determined.The level of Scr,plasma urea,serum neutrophil gelatinase-associated lipocalin(NGAL)and Cystatin C(Cys C)of mice were measured.Histology and apoptosis of renal tissue from mices were examined,Mitochodrial SIRT3 activity of renal tissue was detected.Cells or renal tissue were prepared to assess expression of cyclophilin D(CypD),SIRT3 and cytochromeC(CytC),and the level of CypD acetylation.Result:H/R increased cell death,decreased ??m and SIRT3 expression and increased CytC expression and CypD acetylation in HK2 cells(P<0.05,compared to Control group).Dexmedetomidine(0.01nM and 0.1nM)inhibited these changes induced by H/R in vitro experiments(P<0.05,compared to H/R group),and the effects of 0.1nM dexmedetomidine was better than that of 0.01nM dexmedetomidine(P<0.05).SIRT3 plasmid enhanced these changes of dexmedetomidine.SIRT3 siRNA inhibited the effectes of dexmedetomidine.I/R increased the level of Scr,plasma urea,serum NGAL and Cys C,increased renal tubular necrosis score and the number of apoptotic cells,decreased SIRT3 expression and activity,inceased CytC expression and CypD acetylation(P<0.05,compared to Sham group).Dexmedetomidine inhibited these changes induced by I/R in vivo experiments.These protective effects of dexmedetomidine were not observed in SIRT3 knockdown mice.Conclusions:Dexmedetomidine appears to act at least in part by up-regulating SIRT3 to inhibit mitochondrial damage and cell apoptosis and thereby protect against renal I/R injury. |
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