Analysis Of Liver Inflammation In Chronic Hepatitis B Patients With Detectable HBV DNA And Establishment Of A Non-invasive Index For Predicting Significant Liver Inflammation

Background:Chronic hepatitis B(CHB)virus infection is a worldwide public health burden that causes major adverse outcomes,including liver cirrhosis,liver failure,and hepatocellular carcinoma(HCC).Antiviral therapy may prevent the progress of the disease and improve the long-term prognosis of CHB patients by inhibiting HBV replication.Liver inflammation grades and fibrosis stages of chronic hepatitis B(CHB)patients are regarded as the indications for antiviral therapy,especially for chronic hepatitis B patients with normal ALT and detectable HBV DNA.Currently,there are many methods to evaluate liver fibrosis stage,whereas those for liver inflammatory activity are insufficient.Serum alanine aminotransferase(ALT)levels remain the most commonly used laboratory parameter to reflect liver inflammatory activity.However,the severity of liver inflammation is not always consistent with the levels of ALT in CHB patients.For the CHB patients with normal ALT and detectable HBV DNA in absence of liver fibrosis,liver inflammatory activity is of great significance to determine the initiation of antiviral therapy and to assess prognosis.We aimed to investigate the proportions and determinants of significant inflammation in CHB patients with detectable HBV DNA.Furthermore,we developed a novel noninvasive model comprised of clinical laboratory parameters for predicting significant liver inflammation in CHB patients with detectable HBV DNA and normal ALT.MethodsPart I Presence of liver inflammation in chronic hepatitis B patients with detectable HBV DNAWe conducted a cross-sectional study that retrospectively enrolled 581 CHB patients with detectable HBV DNA who had undergone liver biopsy in Nanjing Drum Tower Hospital and Huai’an No.4 People’s Hospital between Nov 2004 and Oct 2020.Clinical and laboratory data of the included patients,including age,gender,blood routine test,liver function and HBV serological markers were acquired from electronic medical records within two weeks prior to liver biopsy.The ULN of the ALT level was defined as 35 U/L for men and 25 U/L for women according to the AASLD 2018 hepatitis B guidance.Liver inflammation and fibrosis were staged by Scheuer’s classification.G0-G1,G2,and G3-G4 were defined as no or mild,moderate,and severe liver inflammation,respectively.Liver fibrosis was categorized into no significant fibrosis(S0-S1),moderate fibrosis(S2-S3),and cirrhosis(S4).Significant inflammation and significant fibrosis were defined as≥ G2 and≥ S2,respectively.The risk factors of significant liver inflammation for CHB patients with detectable HBV DNA were identified by logistic regression analysis.Part Ⅱ Development of a non-invasive model for significant liver inflammation in chronic hepatitis B patients with detectable HBV DNA and normal ALTIn this study,155 CHB inpatients with detectable HBV DNA and normal ALT who received liver biopsy in Nanjing Drum Tower Hospital and Huai’an No.4 People’s Hospital between November 2004 and October 2020 were included.The clinical laboratory data including gender,age,routine blood test,liver function,HBV serological markers and coagulation function were collected within two weeks before liver biopsy.The definition of ULN for ALT was 35/25 U/L for males/females.Liver inflammation and fibrosis were graded by the Scheuer’s classification.The binary logistic regression was utilized to identified the predictors for significant liver inflammation in CHB patients with detectable HBV DNA and normal ALT.The diagnostic accuracy of different predicting indices for significant liver inflammation was evaluated by the area under receiver operating characteristic curve(AUROC).ResultsPart Ⅰ Presence of liver inflammation in chronic hepatitis B patients with detectable HBV DNAOf 581 CHB patients with detectable HBV DNA,the median age was 38.0(IQR 30.0-46.0)years and 68.2%of the patients were males.A total of 265(45.6%)patients were hepatitis B envelope antigen(HBeAg)-positive.There were 179(30.8%)patients with ALT<1×ULN,205(35.3%)patients with ALT 1-2×ULN,and 197(33.9%)patients with ALT 2×ULN.The HBV DNA levels were below 3 log10 IU/ml in 63(10.8%)patients,3-5 log10 IU/ml in 190(32.7%)patients,5-7 log10 IU/ml in 165(28.4%)patients,and>7 log10 IU/ml in 163(28.1%)patients.A total of 397(68.3%)patients had significant inflammation(≥G2),and 340(58.5%)patients had significant fibrosis(≥ S2).The proportions of significant inflammation were 45.8%,65.9%and 91.4%in patients with ALT<1 ×ULN,ALT 1-2×ULN and ALT>2×ULNOf CHB patients with detectable HBV DNA and normal ALT,90.5%of patients with cirrhosis,59.7%of patients with moderate fibrosis,and 28.7%of patients without significant fibrosis had significant liver inflammation.In the different subgroups on HBeAg statues,HBV DNA levels and age categories,with serum ALT levels and liver fibrosis stages ascending,liver inflammatory activities were increased.At least 20%of CHB patients with detectable HBV DNA and normal ALT in absence of significant fibrosis had significant inflammation in different subgroups.Moreover,the proportion of significant inflammation was highest in patients with moderate HBV DNA levels(5-7 log10 IU/ml),with a decreasing trend in patients with higher and lower levels of HBV DNA.Compared to patients without significant fibrosis,age,ALT,AST,GGT levels and positive rate of HBeAg were higher in patients with significant fibrosis.Significant inflammation was found in 85.0%of patients with significant fibrosis and in 44.8%of patients without significant fibrosis.The multivariate logistic regression analysis revealed that HBV DNA levels[3-5 log10 IU/ml(OR 2.058,95%CI 1.059-4.001,P=0.033),5-7 log10 IU/ml(OR 6.929,95%CI 2.830-16.966,P<0.001),≥ 7 log10 IU/ml(OR 2.947,95%CI 1.278-6.795,P=0.011)],ALT≥2×ULN(OR 4.562,95%CI 2.209-9.423,P<0.001),GGT≥ 50 U/L(OR 2.829,95%Cl 1.499-5.339,P=0.001),and significant fibrosis(OR 4.955,95%CI 3.137-7.824,P<0.001)were independent risk factors for significant inflammation in CHB patients with detectable HBV DNA.Significant fibrosis(OR 4.088,95%CI 2.304-8.214;P<0.001)and moderate serum HBV DNA(5-7 log10 IU/ml)(OR 12.602,95%CI 2.257-70.370;P=0.004)were the independent risk factors for significant inflammation in CHB patients with detectable HBV DNA and normal ALT levels in the multivariate logistic regression analysis.HBV DNA levels[5-7 log10 IU/ml(OR 5.229,95%CI 1.445-18.930;P=0.012),≥ 7 log10 IU/ml(OR 3.350,95%CI 1.096-10.236;P=0.034)]and ALT≥ 2×ULN(OR 4.076,95%CI 1.694-9.807;P=0.002)were the independent risk factors for significant inflammation in CHB patients with detectable HBV DNA in the absence of clinically significant fibrosis in the multivariate logistic regression analysis.After adjusting for age,gender,and HBeAg status,moderate serum HBV DNA level(5-7 log10 IU/ml)(adjusted OR 13.161,95%CI 1.026-168.889,P=0.048)remained the independent risk factor for significant inflammation in CHB patients with detectable HBV DNA and normal ALT in the absence of significant fibrosis.Part Ⅱ Development of a noninvasive index for significant liver inflammation in chronic hepatitis B patients with detectable HBV DNA and normal ALTAmong 155 CHB patients with detectable HBV DNA and normal ALT,the median age was 40.0(IQR 31.0-49.0)years and 58.7%(91/155)of the patients were males.The percentage of HBeAg-positive patients was 32.9%(51/155).42.6%(66/155)of patients had significant liver inflammation,likewise 42.6%(66/155)with significant liver fibrosis.By the logistic regression analysis,we identified that hepatitis B core antibody(HBcAb)(OR 1.241,95%CI 1.042-1.478,P=0.015)and prothrombin time(PT)(OR 1.787,95%CI 1.276-2.503,P=0.001)were independent predictors for significant liver inflammation in CHB patients with detectable HBV DNA and normal ALT.Based on the two predictors,HBcAb-PT Index(HPI)was developed to predict significant liver inflammation(HPI =-10.312+0.216 ×HBcAb(S/CO)+0.581 × PT(s)).Of CHB patients with detectable HBV DNA and normal ALT,HPI value had significant positive correlation with liver inflammatory grades in CHB patients with detectable HBV DNA and normal ALT(r= 0.500,P<0.001).HPI value of patients with significant liver inflammation was significantly higher than that of patients in absence of significant liver inflammation(-1.354 vs.-0.381,P<0.001).The area under receiver operating characteristic curve(AUROC)of HPI for significant inflammation in CHB patients with detectable HBV DNA and normal ALT was 0.777(95%CI 0.701-0.853),significantly higher than that of ALT(AUROC 0.605 95%CI 0.516-0.694,P=0.004)Furthermore,the diagnostic accuracy of HPI for significant liver inflammation was estimated among CHB patients with detectable HBV DNA and normal ALT in absence of significant liver fibrosis.Among 89 patients with detectable HBV DNA and normal ALT in absence of significant liver fibrosis,the median age was 39.0(IQR 32.5-46.0)years and 56.2%(50/89)of the patients were males.32.6%(29/89)of patients was HBeAg-positive.25.8%(23/89)of patients had significant liver inflammation.Among CHB patients with detectable HBV DNA and normal ALT without significant liver fibrosis,the AUROC of HPI for significant inflammation was 0.812(95%CI 0.707-0.917),significantly higher compared to ALT(AUROC 0.599 95%CI 0.472-0.727,P=0.012).Conclusions1.A high proportion of CHB patients with detectable HBV DNA and normal ALT had significant liver inflammation,even though in patients in the absence of significant fibrosis.Liver biopsies are recommended to evaluate liver inflammation in such patients,especially for those with moderate HBV DNA.2.HPI,as a novel,convenient and non-invasive index for predicting significant liver inflammation in CHB patients with detectable HBV DNA and normal ALT,has high diagnostic accuracy.This index can also predict significant liver inflammation in patients with detectable HBV DNA and normal ALT in absence of significant liver fibrosis.