Plasma Diamine Oxidase Is Associated With Different Stages During Chronic Hepatitis B Virus Infection

Part ?:Plasma diamine oxidase(DAO)in patients with chronic hepatitis B,liver cirrhosis and hepatocellular carcinomaBackgroundHepatitis B virus(HBV)chronic infection can lead to liver inflammation,long-term repeated inflammation can lead to cirrhosis and hepatocellular carcinoma,called the "trilogy of hepatitis B virus infection".Liver failure can occur in different stages of chronic HBV infection,with serious onset,high mortality,and lack of effective treatment in the clinic.The liver and intestinal anatomy are closely related and interact with each other through the " gut-liver axis".The intestine is an important barrier to maintain microecology and prevent pathogen invasion.The intestinal digestive substances are first transported to the liver through the portal vein.When the intestinal barrier function is impaired and the intestinal wall permeability is increased,the liver is more likely to be exposed to the intestinal tract.Microorganisms and their metabolites aggravate the original damage of the liver.The level of plasma diamine oxidase(DAO)is the most commonly used molecular index reflecting intestinal barrier function and intestinal wall permeability.However,we do not know the expression of DAO in different stages of chronic HBV infection,and the relationship between plasma DAO levels and disease progression and prognosis.Therefore,it is of great clinical significance to study the dynamic changes of plasma DAO levels in patients with different stages of chronic HBV infection.Aims1.To investigate the plasma DAO levels in patients with chronic hepatitis B,hepatitis B cirrhosis and hepatocellular carcinoma,and to study the correlation between plasma DAO levels and clinical indicators.2.Assessing the level of plasma DAO as a biomarker to evaluate the diagnostic value of readmission in patients with HBV-related decompensated cirrhosis within 6 months.MethodsThis study included 365 patients with chronic HBV infection,including 70 patients with chronic hepatitis B,255 patients with cirrhosis(120 patients with compensated cirrhosis,135 patients with decompensated cirrhosis),and 40 patients with hepatocellular carcinoma and 12 healthy controls.Patients with decompensated cirrhosis were followed up by telephone and inpatient system.The follow-up period was no less than 6 months.Plasma DAO levels were measured in all subjects by ELISA and statistical analysis was performed using SPSS 19.0 software and MedCalc 15.6 software.The categorical variables were tested by chi-square test and the continuous variables were analyzed by Mann-Whitney U test and rank sum test.Spearman correlation analysis was used to analyze the correlation between DAO and clinical and experimental data.Univariate Cox regression analysis was used to determine the risk factors for readmission in patients with HBV-related decompensated cirrhosis.P<0.05 covariates were included in multivariate Cox regression analysis to determine independent risk factors for readmission in patients with HBV-related decompens'ated cirrhosis..The ROC curve was used to evaluate the diagnostic value of DAO in patients with HBV-related decompensated cirrhosis.The Kaplan-Meier method was used to compare the readmission time at cut-off point.P<0.05 on both sides was statistically significant.Results1.The plasma DAO level of patients with chronic hepatitis B was significantly higher than that of healthy controls(P<0.001):the plasma DAO level of patients with liver cirrhosis was significantly higher than that of patients with chronic hepatitis B(P<0.05);the plasma DAO level of patients with hepatocellular carcinoma was significantly higher than patients with liver cirrhosis(P<0.001)and patients with chronic hepatitis B(P<0.001)2.In patients with chronic hepatitis B,there was no significant difference in DAO levels between HBeAg-positive and HBeAg-negative groups;there was no significant difference between the HBV-DNA positive group and the HBV-DNA negative group.Plasma DAO levels were positively correlated with ALT,AST,and TBIL(r=0.530,P<0.001;r=0.483,r<0.001;r=0.313,P<0.05).There is no significant correlation with indicators such as HBsAg,ALB,PTA,age,gender.3.In patients with hepatitis B cirrhosis,the plasma DAO level of patients with compensated cirrhosis was significantly lower than that of patients with decompensated cirrhosis(P<0.001);the plasma DAO level of patients with Child-pugh score A was significantly lower than that of Child-pugh B patients(P<0.001);Child-pugh score B patients had significantly lower plasma DAO levels than Child-pugh C patients(P<0.001).There was no significant difference in plasma DAO levels between ascites positive patients and ascites negative patients(P>0.05).The level of DAO in patients with gastrointestinal bleeding was significantly higher than that in patients without gastrointestinal bleeding(P<0.05).The level of plasma DAO in patients with spontaneous peritonitis was significantly higher than that in patients without spontaneous peritonitis(P<0.05).Plasma DAO levels in patients with hepatic encephalopathy were significantly higher than those without hepatic encephalopathy(P<0.05).Plasma DAO levels correlated with AST,TBIL,and ALB levels(r=0.14,P=0.026;r=0.574,P=0.000;r=-0.230,P=0.000).4.After 6 months of follow-up,36 of 135 patients with HBV-related decompensated cirrhosis were readmission for liver-related disease.Univariate COX analysis showed plasma DAO levels(HR=1.126,95%CI:1.068-1.188,P<0.001),Child-pugh score(HR=3.700,95%C1:1.665-8.223,P<0.05),gastrointestinal bleeding(HR=5.179,95%CI:2.113-12.690,P<0.001),hepatic encephalopathy(HR=4.375,95%Cl:1.567-12.215,P<0.001),spontaneous bacterial peritonitis(HR=3.077,95%CI:1.176-8.049,P<0.001)and hemoglobin(HR=0.979.95%CI:0.963-0.995,P<0.05)were risk factors for readmission with 6 months of HBV-related decompensated cirrhosis patients.Multivariate COX analysis showed plasma DAO levels(HR=1.102,95%CI:1.023-1.188,P<0.05)and hepatic encephalopathy(HR=5.01 8,95%CI:1.436-17.533,P<0.05)were independent risk factors for readmission of patients with HBV-related decompensated cirrhosis within 6 months.5.The operating characteristic curve area(AUROC)of plasma DAO levels for predicting HBV-related decompensated cirrhosis patients readmission within 6 months was 0.769(SE 0.0457,95%Cl 0.689-0.837),significantly better than HE[0.598(SE 0.0403,95%C10.511-0.682,P<0.05)]and Child-Pugh score[0.652(SE 0.0474,95%CI 0.565-0.731,P<0.05)1.When the plasma DAO was selected to predict readmission within 6 months of HBV-related decompensation cirrhosis patients,the optimal cut-off value of 19.7 ng/mL,sensitivity was 58.33%,specificity was 84.35%was selected,Readmission curve of the two groups has significant differences[1.75(SE 0.435,95%CI:0.898-2.602)vs.3.65(SE 0.386,95CI:2.893-4.407),P<0.05)].6.In patients with hepatocellular carcinoma,plasma DAO levels were significantly higher in HCC patients with tumor size>3 cm than tumor size<3cm(P<0.001).Plasma DAO levels in patients with TNM stage III/IV were significantly higher than those in ?/?(P<0.05);plasma DAO levels were positively correlated with AFP levels(r=0.582,P<0.001).There was no significant correlation between the presence or absence of cirrhosis,the number of tumors,and the HBeAg status and ALT,AST,TBIL,ALB,and PTA,gender and age.ConclusionThe level of plasma DAO in patients with chronic HBV infection was significantly higher than that in healthy controls.As the disease progressed,DAO levels gradually increased,suggesting that intestinal permeability is involved in the process of chronic HBV infection.Plasma DAO levels in patients with chronic hepatitis B were closely related to liver function indicators;plasma DAO levels in patients with cirrhosis were closely related to the severity and complications of the disease.Plasma DAO levels were independent risks of readmission with 6 months in patients with decompensated cirrhosis.DAO level can be used as molecular markers to predict readmission for patients with decompensated cirrhosis within 6 months;plasma DAO levels in patients with hepatocellular carcinoma were associated with tumor size and TNM staging.Part?:Plasma diamine oxidase(DAO)used as a marker for predicting 1-month mortality of patients with acute-on-chronic hepatitis B liver failureBackgroundAcute-on-Chronic Hepatits B liver failure(ACHBLF)refers to acute liver function decompensation based on chronic HBV infection.ACHBLF progresses rapidly and short-term mortality is high,with approximately 50-90%of ACHBLF patients dying within 1 month.At present,ACHBLF lacks specific and effective treatment in clinical practice.Liver transplantation is often the only effective treatment strategy for ACHBLF patients.However,due to the lack of liver donors,only a very small number of ACHBLF patients can successfully receive liver transplantation.Therefore,it is very important to establish an effective ACHBLF prognostic model to accurately identify patients with high mortality in ACHBLF.Model for end-stage liver diseases(MELD)is currently the most commonly used model for predicting the prognosis of liver failure,but the score is not based on ACHBLF,and MELD predicts short-term mortality rate of ACHBLF remains controversial.Because most patients with ACHBLF die within a month,a new indicator is urgently needed to effectively respond to the 1-month prognosis of ACHBLF patients.Diamine oxidase(DAO)is a highly active intracellular enzyme that has been shown to be a fast and sensitive biomarker for intestinal wall permeability and barrier function.Our previous studies have confirmed that plasma DAO levels are significantly elevated in chronic HBV infections including chronic hepatitis B,liver cirrhosis,and hepatocellular carcinoma,and plasma DAO levels and liver function in patients with chronic hepatitis B and complications in patients with liver cirrhosis As well as the tumor size and TNM staging of hepatocellular carcinoma patients,but there is currently no data on the potential value of plasma DAO as a biomarker for predicting the 1-month prognosis of ACHBLF patients.In our current study,we measured plasma DAO concentrations in patients with ACHBLF to determine whether DAO was associated with the condition and prognosis of ACHBLF,and to assess the diagnostic value of DAO as a biomarker for predicting the prognosis of ACHBLF within 1 month.Aims1.To determine the plasma DAO level in patients with ACHBLF and to explore the relationship between DAO and severity of ACHBLF;2.Assessing the diagnostic value of plasma DAO levels as a biomarker for predicting 1-month mortality in patients with ACHBLFMethodsThis study included 106 patients with ACH'BLF who were hospitalized in the Department of Hepatology,Qilu Hospital of Shandong University.The follow-up period was no less than one month.The end point of follow-up was a death event.Plasma DAO levels were measured in all subjects by ELISA and statistical analysis was performed using SPSS 19.0 software and MedCalc 15.6 software.The categorical variables were tested by chi-square test and the continuous variables were analyzed by Mann-Whitney U test Spearman correlation analysis was used to analyze the correlation between DAO and clinical and experimental data.Univariate logistic regression analysis was used to determine the risk factors affecting the 1-month prognosis of ACHBLF patients.Variates with P<0.05 were included in multivariate logistic regression analysis to determine independent risk factors for 1-month mortality in patients with ACHBLF.To assess the combined predictive value of DAO and MELD scores for 1-month mortality in ACHBLF patients,a new variable based on the binary logistic regression equation was established:variable(plasma DAO plus MELD score)=0.642 × MELD+0.056 × DAO-16.671.The ROC curve was used to evaluate the diagnostic value of DAO,MELD score,DAO plus MELD score for 1-month mortality of ACHBLF patients.The Kaplan-Meier method was used to compare the cumulative survival time of each cut-off point.P<0.05 on both sides considered the difference to be statistically significant.Results1.A total of 106 patients with ACHBLF were enrolled in the study.After one month of follow-up,66 patients died,with a mortality rate of 62.3%.We found that the group of nonsurvivals had higher median levels of TBIL[305(262,385)vs.196(151,248)?mol/L,P<0.001],MELD score[27.3(24.8,32.3)vs.22.0(20.7,26.5),P<0.001],and INR[2.5(2.1,3.7)vs.1.9(1.8,2.1),P<0.01]and and lower median levels ALB[32.3(30.0,34.7)vs.34.2(31.6,37.4)g/L,P<0.05],PTA[30(18,33)vs.38(32,9),P<0.05],AFP[56(39,88)vs.156(104,416)ng/mL,P<0.001],and PLT[129(100,150)vs.132(111,173)×109/L,P<0.05]than the survival group.In nonsurvival group,the incidences of ascites(60.6%vs.17.5%,P<0.001),SBP(54.5%vs.5%,P<0.001)and HE(57.6%vs.12.5%,P<0.001)were' higher than survival group.Between survival and nonsurvival group,age,HBV-DNA,HBsAg,HBeAg,ALT,AST,WBCs,HGB,Na and Cr were no significantly differences.2.Plasma DAO levels in the surviving patients group[14.0(7.1,26.5)ng/ml]were significantly lower than the death group[58.6(32.5,121.3)ng/ml,P<0,001].3.We used Spearman correlation analysis to correlate plasma DAO levels with clinical parameters of ACHBLF patients.The results showed that plasma DAO levels and TBIL(r=0.507,P<0.001),INR(r=0.518,P<0.001)MELD scores(r=0.578,P<0.001),PTA(r=-0.511,P<0.001)AFP(r=-0.412,P<0.001)and HGB(r=-0.231,P=0.017)were significantly correlated in ACHBLF patients.However,there was no significant correlation between DAO levels and age,HBsAg,HBeAg,HBV-DNA,ALT,AST,ALB,Cr,WBCs and PLTs(P>0.05).4.Univariate logistic regression analysis showed MELD score(OR=1.838,P<0.001),plasma DAO level(OR=1.055,P<0.001),ALB(OR=0.886,P<0.05),PLT(OR=0.985),P<0.05),ascites(OR=7.731,P<0.001),SBP(OR=22.800,P<0.05)and HE(OR=9.500,P<0.05)were significant factors affecting the 1-month prognosis of ACHBLF patients.5.Multivariate logistic regression analysis showed plasma DAO levels(OR=1.053,P<0.05),HE(OR=7.678,P<0.05),SBP(OR=15.91,P<0.05)and MELD score(OR=1.680,P<0.05)were independent risk factors for 1-month mortality in ACHBLF patients.6.The AUROC of plasma DAO was 0.880(SE 0.0329,95%CI 0.803-0.935),and the AUROC of MELD score was 0.843(SE 0.0391,95%Cl 0.760-0.907).There was no statistically significant difference between them(P=0.43).).The AUROC of the DAO plus MELD score was 0.916(SE 0.0256,95%Cl 0.846-0.961),which was significantly higher than the MELD score and DAO.7.The cutoff value of plasma DAO level was 15.2 ng/mL,with sensitivity 95.45%,specificity 62.5%,MELD score was 22.6,with sensitivity 90.91%,specificity 67.5%,DAO plus MELD score was 0.07,with sensitivity 87.88%,specificity 80%.The cumulative survival time of patients with ACHBLF with plasma DAO<15.2ng/mL and>15.2ng/mL was statistically significant(P<0.001).The cumulative survival time of ACHBLF patients with MELD<22.6 and MELD>22.6 was statistically significant(P<0.001).The cumulative survival time of DAO combined with MELD<0.07 and DAO combined with MELD score>0.07 was statistically significant(P<0.001).ConclusionPlasma DAO levels correlate with the severity of ACHBLF patients,and DAO combined with MELD scores can predict 1-month mortality of ACHBLF better.Part ?:Preliminary study on the relationship between plasma diamine oxidase and gut microbiota in patients with acute-on-chronic hepatitis B pre-liver failureBackgroundAcute-on-chronic hepatitis B liver failure(ACHBLF)is progressing rapidly,and the short-term mortality is high.There is a lack of specific and effective treatment in clinical practice.Therefore,early detection and treatment of patients with ACHBLF is very important.Therefore,domestic and foreign scholars have proposed the concept of early liver failure.Acute-on-Chronic hepatitis B pre-liver failure(pre-ACHBLF)is a pre-stage of CHB progression to liver failure with serum total bilirubin(TBIL)?85umol/L and prothrombin activity(PTA)>40%.Early diagnosis and identification of pre-ACHBLF patients,timely and effective treatment,can reduce the morbidity and mortality of patients with ACHBLF.At present,a number of studies have confirmed that ACHBLF has a disorder of intestinal flora structure and function.Our previous study found that intestinal diathermic index plasma diamine oxidase(DAO)can reflect the severity of ACHBLF,which is a independent risk factors for one-month prognosis of ACHBLF patients.It is unclear whether there is a disorder of intestinal flora in patients with pre-ACHBLF who have a potential risk of developing ACHBLF,and whether there is damage to the intestinal barrier function.The study aimd to explore the intestinal barrier function,intestinal flora and their relationship in pre-ACHBLF patients.Aims1.To explore the relationship between plasma DAO levels and clinical indicators in patients with pre-ACHBLF;2.To explore the changes of intestinal flora in patients with pre-ACHBLF;3.To explore the correlation between plasma DAO levels and intestinal flora in patients with pre-ACHBLF.MethodsThis study included 31 patients with pre-ACHBLF and 26 healthy adult volunteers.Plasma DAO levels were measured by ELISA,and Spearman correlation analysis was used to analyze the correlation between DAO levels and clinical indicators.The subject's stool was used as a specimen,and the intestinal flora of the study subject was detected by 16S rRNA sequencing.Through microbial diversity analysis,NMDS analysis and species composition difference analysis clarified the changes in intestinal flora of pre-ACHBLF patients.Spearman correlation analysis was used to study the correlation between plasma DAO levels and pre-ACHBLF differential flora.P<0.05 on both sides considered the difference to be statistically significant.Results1.Plasma DAO levels were significantly higher in patients with pre-ACHBLF than in HCs[31.7(22.1,43.2)vs.1.7(1.4,2.2)ng/mL,P<0.001].In patients with pre-ACHBLF,plasma DAO levels were positively correlated with TBIL levels(r=0.672,P=0.000),negatively correlated with ALB levels(r=-0.548,P=0.001),and positively correlated with AST levels(r=0.369,P=0.041),there was no significant correlation between ALT level,PTA level,MELD score,age,HBsAg,HBeAg,HBV-DNA,Cr,WBCs and PLTs in patients with pre-ACHBLF(P>0.05).2.57 stool samples obtained 5,101,887 sequencing sequences,all sequences were clustered by 97%similarity,and all sample copolymers obtained 916 OUT,of which 501 were common OTU,and the OTU of HCs was 358.The pre-ACHBLF unique OTU was 57.3.In pre-ACHBLF group,microbial diversity index:shannon index decreased significantly(P<0.01),simpson index decreased significantly(P<0.01);microbial abundance index:Chaol index decreased significantly(P<0.01),ACE index decreased significantly(P<0.01),suggesting that the intestinal flora diversity and the abundance decreased in pre-ACHBLF patients.4.NMDS analysis showed that the pre-ACHLBF group and the HCs group could be easily distinguished(stress=0.161).The Anosim analysis showed that the difference between the pre-ACHBLF group and the HCs group was greater than the intra-group difference(R=0.3873,P=0.001).It is suggested that the composition of the flora of pre-ACHBLF patients changes.5.At the level of the phylum,the relative abundance of the Firmicutes was reduced in the pre-ACHBLF group(0.544 vs.0.641,P=0.035);the relative abundance of the Proteobacteria increased(0.163 vs.0.069,P=0.008).6.At the genus level,we screened 19 strains with different abundances,of which 11 were significantly missing and 8 species of pre-ACHBLF abundance increase.7.The plasma DAO level was positively correlated with the relative abundance of Veillonella(r=0.665,P=0.000);positively correlated with the relative abundance of Streptococcus(r=0.436,P=0.014).ConclusionThe level of DAO in Pre-ACHBLF patients was significantly higher than that in HCs group,and the level of DAO was positively correlated with TBIL level and AST level.and negatively correlated with ALB level.Pre-ACHBLF patients had changes in intestinal flora structure and abundance.Pre-ACHBLF abnormally enriched Veillonella,Streptococcus and plasma DAO levels were positively correlated.