The Clinical Study Of Acute Renal Injury Of Septic Patients And Cell Cycle Arrest During Acute Renal Injury In Sepsis

ObjectiveThe incidence of acute kidney injury(AKI)in ICU patients was 15%?20%,and the fatality rate of those patients was 11.0%.In addition to increasing the length of hospital stay,hospitalization cost and mortality,AKI can also increase the incidence of late chronic kidney disease(CKD)and long-term death risk.Sepsis is the main factors leading to AKI.In recent years,researchers conducted a lot of basic and clinical researches about biomarkers and pathological physiology relevant sepsis AKI aims to address the specific mechanisms of occurrence and development of diseases,they want to be able to apply the new indicators to predict or detection sepsis AKI more early.Unfortunately,although researchers have conducted a large number of studies on AKI with sepsis,the current diagnosis of AKI still relies on traditional indicators such as creatinine and urine volume,which is not conducive to the early detection and prevention of AKI,at the same time,after the occurrence of AKI,we still lack sufficient effective treatment measures.Based on this background,this study firstly studied the clinical index evaluation of blood flow to kidneys,combined use of resistance index(RI)of the kidney and urinary oxygen partial pressure change assessment to predict the occurrence of sepsis AKI.Our basic research study the cell cycle arrest of sepsis AKI,which may related with AMPK phosphorylation level within cell cycle arrest and cell regeneration.MethodsClinical part:septic shock patients admitted to the intensive care department of our hospital from 2018 August to November were selected.We Collected baseline data of those patients,we uesed bedside ultrasound to obtain renal resistance index(RI),and used gas analysis to detected the levels of urine oxygen partial pressure,and we analysis risk factors by logistics regression.At the same time,we predict the occurrence of acute kidney injury in patients with septic shock by the receiver-operating characteristic curve(ROC)to analysis RI,urine oxygen partial pressure,RI combined urine oxygen partial pressure.Basic part:human renal epithelial cell line hk-2 cells were selected in the cell experiment,and the sepsis-damaged cell model was generated by LPS stimulation.Compound C was used as AMPK phosphorylation inhibitor.After pre-stimulation,the effects of different levels of AMPK phosphorylation on cell cycle were studied.Cell cycle detection,AMPK phosphorylation level detection,and mRNA and protein expression levels of cyclin D1,cyclin E,CDK2 and CDK4 in G1/S phase were performed in each group.Cell activity was detected by cck-8 and MTT assays,and intracellular ATP,glucose and apoptosis-related protein caspase-3 and bcl-2 levels were also detected.During animal experiment we choose C57BL/6 mice,cecum ligation perforation in middle abdominal cavity to infection sepsis model.18 h after CLP,we detection the cell cycle protein expression,cell cycle changes,tissue protein levels detection by immunohistochemical.We also study the kidney tissue HE staining and TUNEL staining,apoptosis related proteins such as caspase-3,Bcl-2 protein,serum creatinine,blood urea nitrogen etc.Results:(1)clinical studies showed that RI(OR=1.139,95%ci 1.029?1.261,P=0.012)and urine oxygen partial pressure(OR=0.957,95%ci 0.923?0.991,P=0.014)were independent risk factors of AKI in patients with septic shock.The predictive value of RI,urine oxygen partial pressure,and RI combined with urine oxygen partial pressure in the occurrence of AKI:the sensitivity and specificity of RI>0.694 in the prediction of AKI in patients with septic shock were 96.6%and 30.2%,respectively,the sensitivity and specificity of AKI in patients with septic shock were 48.3%and 81.4%,as well,the sensitivity and specificity of RI combined with urine oxygen partial pressure in predicting AKI in patients with septic shock were 65.5%and 76.7%,respectively.Patients were divided into three groups by the cut-oof value of RI 0.70 and urine oxygen partial pressure 48mmHg.Lactic acid in patients with RI 0.70 and urine oxygen partial pressure 48mmHg was higher than that in other patients(P<0.0125).The incidence of AKI in the four groups was statistically significant(P=0.020).(2)basic studies found that p-AMPKa 1 levels were increased in HK-2 cells after LPS stimulation.Compound C pretreated before LPS stimulation reduced p-AMPK a 1 levels compared with LPS group.WB showed that the changes in each group were similar to immunofluorescence results.LPS significantly inhibited the cell cycle of HK-2 cells,primarily in the G1/S phase.Compound C pretreatment relieved cell cycle arrest and increased cell number and activity(P<0.05).LPS stimulation decreased the levels of cyclin E,cyclin D1,CDK2 and CDK4 in HK-2 cells.When Compound C stimulation was used,the level of these four kinds of protein was higher than that when LPS stimulation alone accompanied by cell cycle arrest relieved.Compound C pretreated cells subjected to LPS stimulation showed lower levels of caspase-3 protein in both groups than in the LPS group.The results of animal experiments were similar to the results of cell experiments,indicating that different levels of AMPK phosphorylation can regulate cell cycle arrest during sepsis AKI in animals,which is achieved by regulating the key regulatory protein of cell cycle in G1/S phase.Through tissue staining and overall organ function evaluation,we found that Compound C inhibited AMPK phosphorylation level and improved cell cycle arrest was more conducive to cell regeneration,organ function protection and recovery.Conclusion:(1)High RI and low urine oxygen partial pressure are independent risk factors for AKI in patients with septic shock.(2)Combined application of RI and urine oxygen partial pressure has a good value in the early prediction of AKI in patients with septic shock;(3)RI of 0.70 and urine oxygen partial pressure of 48mmHg were used as cut-off values,which could be used as early predictors of AKI in patients with septic shock and guide clinical treatment.(4)Cell cycle arrest in G1/S phase was observed in renal injury of sepsis,accompanied by increased AMPK phosphorylation level.(5)Inhibition of AMPK activation can relieve cell cycle arrest in sepsis and promote cell regeneration;(6)AMPK regulates cell cycle by affecting key regulatory proteins in G1/S phase.(7)Removing cell cycle arrest and promoting cell regeneration are more conducive to organ function protection and recovery in sepsis kidney injury.