A Cross-Sectional Study On Vitamin D₃Metabolism In402Chronic HBV Patients In Different Progression Stages

Objective:To determine the prevalence of Vitamin D3deficiency in HB V population and the situation of Vitamin D3in different stages of HBV disease progression ranging from HBV carriers, chronic hepatitis B, compensated cirrhosis, decompensated cirrhosis and hepatocellular carcinoma. We have also supposed the imbalance of hydroxylases as the reason for Vitamin D3deficiency to reveal the exact mechanism in chronic hepatitis B liver diseases.Methods:This study was a cross-sectional study. Patients (n=1278) with positive HBsAg were recruited in this study at Beijing Friendship Hospital, Capital Medical University from August1st,2012to April30th,2013. We also enrolled408volunteers as healthy control from Physical Examination Center, Beijing Friendship Hospital from January1st,2013to April30th,2013. Totally, there were1686samples, stored at-80℃. The pertinent medical data including virological markers (HBsAg, HBeAg, HBeAb and HBV-DNA loading), biochemical parameters and image profiles were extensively reviewed. Finally, we gave up956samples and enrolled730qualified subjects into our study,328of them were healthy people as control, and402of them were patients infected by HBV, including HBV Carriers (n=75), CHB(n=142), CC(n=66), DC(n=59) and HCC(n=60). The measurement of25(OH) D3in730samples was achieved by High Performance Liquid Chromatography Tandem Mass Spectrometry method (HPLC-MS/MS). HPLC (Transcend, ThermoFisher Scientific, Pennsylvania, USA) and MS of API3000(Applied Biosystems/MDS Sciex, USA) with the chromatographic column Agilent ZORBAX SB-C18(Agilent, HP, California, USA) were used in the study. Part of730samples were detected the level of hydroxylases (n=135) including CYP27A1, CYP2R1and CYP24A1. We used Enzyme-linked Immunosorbent Assay to test serum CYP27A1(E93538Hu, Uscn, US), CYP2R1(CSB-EL006431HU, CUSABIO, China) and CYP24A1(CSB-EL006401RA, CUSABIO, China). The operation process was in accordance with manufactures’instructions.Results:(1)、The25(OH)D3status of HBV population was lower than healthy group (7.08±4.11vs8.22±1.33ng/mL; P<0.05). Among the HBV population, the25(OH)D3status of HBV carriers was lower than healthy group,but had no significance(8.15±4.47vs8.22±1.33ng/mL, P>0.05). There was a consistent trend for lower25(OH)D3levels with increasing severity of cirrhosis, while the DC had the lowest25(OH)D3status. The level of25(OH)D3in HCC group was lower than CHB, higher than DC group (7.75±4.34vs4.33±3.11ng/mL, P<0.05).(2)、Among the HBV population, the25(OH)D3level had positive correlation with CHOL (r=0.264, P<0.05), negative correlation with TBiL(r=-0.229, P<0.05), positive correlation with ALB(r=0.383, P<0.05) and negative correlation with AST(r=-0.118, P<0.05), but had no correlation with ALT(r=0.034, P>0.05).(3)、The95%reference range of healthy group were5.61-10.83ng/mL, and the degree of Vitamin D deficiency were defined as mild deficiency (4-5.61ng/mL), moderate deficiency (2-4ng/mL) and severe deficiency(=2ng/mL). Within the healthy control group,81.7%were normal level and there was no severe deficiency.With the progression of the liver disease:the normal proportion were reducing and mild-moderate deficiency were increasing.The severe deficiency appeared in CHB group firstly and DC group with the highest proportion.(4)、Compared with healthy control, three hydroxylases were overall increased. There was a consistent trend for lower25(OH)D3levels with decreasing CYP27A1and increasing trend of CYP24A1and CYP2R1.(5)、Among the HBV population, the25(OH)D3level had negative correlation with CYP2R1(r=-0.208, P<0.05) and CYP24A1(r=-0.266, P<0.05), and positive correlation with CYP27A1(r=0.201, P<0.05). CYP27A1had increasing tends with increasing CYP2R1, but there was no correlation between them. CYP2R1had positive correlation with CYP24A1(r=0.295, P<0.05). Conclusion:Vitamin D3deficiency is common in HBV population and the25(OH)D3levels are gradually decreasing with the progression of liver disease, especially in decompensated cirrhosis patients. The serum25(OH)D3reducing may be related with the following reasons:the lack of pre-vitamin D3materials (CHOL), jaundice affecting the synthesis of pre-vitamin D3in the skin, declining synthetic ability of liver, reducing expression of VDR, mitochondria damage and the mismatched hydroxylases, but may had nothing to be with HBV-DNA replication. There is a disorder of hydroxylase metabolism in HBV population.