Association of Cytokines with Alanine Aminotransferase, Hepatitis B Virus Surface Antigen and Hepatitis B Envelope Antigen Levels in Chronic Hepatitis B

Background： Cytokines play an important role in occorrence and recovery of hepatitis B virus （HBV） infection. The aim of this study was to investigate the changes ofcytokines concentration and its correlation to alanine aminotransferase （ALT）, HBV deoxyribonucleic acid （HBV-DNA）, hepatitis B envelope antigen （HBeAg）, and HBV surface antigen （HBsAg） in the development of chronic hepatitis B （CHB）. Methods： Thirteen healthy individuals （HI）, 30 chronic HBV-infected patients in immune tolerant （IT） phase, and 55 CHB patients were enrolled between August 2015 and May 2017. The peripheral blood samples were collected from all individuals. Tile levels of interferon （IFN）-α2, interleukin （IL）-10, transforming growth factor （TGF）-β1, HBV-DNA, HBsAg, and HBeAg and liver function were measured. The quantitative determinations of cytokines levels, including I FN-ct2, IL-10, and TGF-[31 were performed using Luminex multiplex technology. The correlation of cytokines to ALT, HBV-DNA, HBsAg, and HBeAg was analyzed by linear regression analysis. Results： IFN-ct2 levels were similar between HI and IT groups （15.35 [5.70, 67.65] pg/ml vs. 15.24 [4.07, 30.73] pg/ml, Z = -0.610, P - 0.542）, while it elevated significantly in CH B group （35.29 [ 15.94, 70.15] pg/ml vs. 15.24 [4.07, 30.73] pg/ml; Z = -2.522, P = 0.012）. Compared with HI group （3.73 [2.98, 11.92] pg/ml）, IL-10 concentrations in IT group （5.02 [2,98, 10.11] pg/ml）, and CHB group （7.48 [3. I 0, 18.00] pg/ml） slightly increased （X^2 = 2.015, P - 0.365）, and there was no significant difference between IT and CHB group （Z =- 1.419, P = 0.156）. The TGF-β1 levels among HI （3.59 ±0.20 pg/ml）, IT （3.62 ±0.55 pg/ml）, and CHB groups （3.64±0.30 pg/ml） were similar （X^2=2.739, P = 0.254）. In all chronic HBV-infected patients （including patients in IT and CHB groups）, the elevation of IFN-α2 level was significantly associated with ALT level （β = 0.389, t = 2.423, P = 0.018）, and was also negatively correlated to HBV-DNA load （β = -0.358, t=-2.308, P = 0.024）, HBsAg （β = -0.359, t = -2.288, P = 0.025）, and HBeAg contents （β = -0.355, t = -2.258, P = 0.027）. However, when both ALT level and cytokines were included as independent variable, HBV-DNA load, HBsAg, and HBeAg contents were only correlated to ALT level （[β= -0.459, t = -4.225, P = 0.000; β = -0.616, t = -6.334, P = 0.000; and β = -0.290, t = -2.433, P = 0.018; respectively）. Conclusions： IFN-α2 elevation was associated with ALT level in patients with chronic HBV infection. However, in CHB patients, only ALT level was correlated to HBV-DNA, HBsAg and HBeAg contents.

Plasmacytoid Dendritic Cell Function and Cytokine Network Profiles in Patients with Acute or Chronic Hepatitis B Virus Infection

Background： Plasmacytoid dendritic cells （pDCs） and cytokines play an important role in occurrence and recovery of hepatitis B virus （HBV） infection. The aim of this study was to explore the frequency and function ofpDC and serum cytokine network profiles in patients with acute or chronic HBV infection. Methods： The healthy individuals （HI group）, hepatitis B envelope antigen （HBeAg）-positive chronic HBV patients in immune tolerance （IT） phase （IT group）, HBeAg-positive chronic HBV patients （CHB group）, and acute HBV patients （AHB group） were enrolled in this study. The frequency of cluster of differentiation antigen 86 （CD86） ＋ pDC and the counts of CD86 molecular expressed on surface ofpDC were tested by flow cytometer. The quantitative determinations ofcytokines, including Fins-like tyrosine kinase 3 ligand （FIt-3L）, interferon （1FN）-α2, IFN-γ, interleukin （IL）-17A, IL-6, IL-10, transforming growth factor （TGF）-β1 and TGF-β2, were performed using Luminex multiplex technology. Results： In this study, there were 13 patients in HI group, 30 in IT group, 50 in CHB group, and 32 in AHB group. Compared with HI group, HBV infected group （including all patients in IT, CHB and AHB groups） had significantly higher counts of CD86 molecular expressed on the surface ofpDC （4596.5 ± 896.5 vs. 7097.7 ± 3124.6; P 〈 0.001）. The counts of CD86 molecular expressed on the surface of pDC in CriB group （7739.2 ±4125.4） was significantly higher than that of IT group （6393.4 ± 1653.6, P=0.043）. Compared with IT group, the profile of cytokines of FIt-3L, IFN-γ, and IL-17A was decreased, IFN-α2 was significantly increased （P =0.012） in CH B group. The contents of IL-10, TGF-{31, and TGF-132 in AHB group were significantly increased compared with IT and CHB groups （all P 〈 0.05）. Conclusions： This study demonstrated that the function of pDC was unaffected in HBV infection. The enhanced function of pDC and IFN-α2 might involve triggering the immune response from IT to hepatitis active phase in H BV infection. Acute patients mainly presented as down-regulation of the immune response by enhanced IL-10 and TGF-β.

Kinetics of Hepatitis B Surface Antigen Level in Chronic Hepatitis B Patients who Achieved Hepatitis B Surface Antigen Loss during Pegylated Interferon Alpha-2a Treatment

Background： Hepatitis B surfhce antigen （HBsAg） Ioss/seroconversion is considered to be the ideal endpoint of antiviral therapy and the ultimate treatment goal in chronic hepatitis B （CHB）. This study aimed to assess the patterns of ttBsAg kinetics in CHB patients who achieved HBsAg loss during the treatment of pegylated interferon （PEG-IFN） α-2a. Methods： A total of 150 patients were enrolled, composing of 83 hepatitis B envelope antigen （H BeAg）-positive and 67 HBeAg-negative patients. Patients were treated with PEG-IFN α-2a180 μg/week until HBsAg loss/seroconversion was achieved, which occurred within 96 weeks. Serum hepatitis B virus deoxyribonucleic acid and serological indicators （HBsAg, anti-HBs, HBeAg, and anti-HBe） were determined before and every 3 months during PEG-1FN α-2a treatment. Biochemical markers and peripheral blood neutrophil and platelet counts were tested every 1-3 months. Results： Baseline HBsAg levels were 2.5 ± 1.3 log IU/ml, and decreased rapidly at 12 and 24 weeks by 48.3%, and 88.3%, respectively. The mean time to HBsAg loss was 54.2 ± 30.4 weeks, though most patients needed extended treatment and 30.0% of HBsAg loss occurred during 72-96 weeks. Baseline HBsAg levels were significantly higher in HBeAg-positive patients （2.9 ± 1.1 log 1U/ml） compared with HBeAg-negative patients （2.0 ± 1.3 log I U/ml; t = 4.733, P 〈 0.001）, but the HBsAg kinetics were similar. Patients who achieved HBsAg loss within 48 weeks had significantly lower baseline HBsAg levels and had more rapid decline of HBsAg at 12 weeks compared to patients who needed extended treatment to achieve HBsAg loss. Conclusions： Patients with lower baseline HBsAg levels and more rapid decline during early treatment with PEG-IFN are more likely to achieve HBsAg loss during 96 weeks of treatment, and extended therapy longer than 48 weeks may be required to achieve HBsAg loss.

Ratios of T-helper 2 Cells to T-helper 1 Cells and Cytokine Levels in Patients with Hepatitis B

Background：Hepatitis B is an immune response-mediated disease.The aim of this study was to explore the differences of ratios of T-helper （Th） 2 cells to Thl cells and cytokine levels in acute hepatitis B （AHB） patients and chronic hepatitis B virus （HBV）-infected patients in immune-tolerance and immune-active phases.Methods：Thirty chronic HBV-infected patients in the immune-tolerant phase （IT group） and 50 chronic hepatitis B patients in the immuneactive （clearance） phase （IC group）,32 AHB patients （AHB group）,and 13 healthy individuals （HI group） were enrolled in the study.Th cell proportions in peripheral blood,cytokine levels in plasma,and serum levels of HBV DNA,hepatitis B surface antigen,and hepatitis B e antigen were detected.Results：The Th1 cell percentage and Th2/Th1 ratio in the HBV infection group （including IT,IC,and AHB groups） were significantly different from those in HI group （24.10% ± 8.66% and 1.72 ± 0.61 vs.15.16% ± 4.34% and 2.40 ± 0.74,respectively;all P 〈 0.001）.However,there were no differences in the Th1 cell percentages and Th2/Th1 ratios among the IT,IC,and AHB groups.In HBV infection group,the median levels of Flt3 ligand （Flt3L）,interferon （IFN）-γ,and interleukin （IL）-17A were significantly lower than those in HI group （29.26 pg/ml,33.72 pg/ml,and 12.27 pg/ml vs.108.54 pg/ml,66.48 pg/ml,and 35.96 pg/ml,respectively;all P 〈 0.05）.IFN-α2,IL-10,and transforming growth factor （TGF）-β2 median levels in hepatitis group （including patients in AHB and IC groups） were significantly higher than those in IT group （40.14 pg/ml,13.58 pg/ml,and 557.41 pg/ml vs.16.74 pg/ml,6.80 pg/ml,and 419.01 pg/ml,respectively;all P 〈 0.05）,while patients in hepatitis group had significant lower Flt3L level than IT patients （30.77 vs.59.96 pg/ml,P =0.021）.Compared with IC group,patients in AHB group had significant higher median level s of IL-1 0,TGF-β 1,and TGF-β2 （22.77 pg/ml,10,447.00 pg/ml,and 782.28 pg/ml vs.8.66 pg/ml,3755.50 pg/ml,and 482.87 pg/ml,respectively;all P 〈 0.05）.Conclusions：Compared with chronic HBV-infected patients in immune-tolerance phase,chronic HBV-infected patients in immune-active phase and AHB patients had similar Th2/Th 1 ratios,significantly higher levels of IFN-α2,IL-10,and TGF-β.AHB patients had significantly higher IL-10 and TGF-β levels than chronic HBV-infected patients in immune-active phase.

＂Onion Skin-liked Sign＂ in Thyroid Ultrasonography： A Characteristic Feature of Benign Thyroid Nodules

Background： Some ultrasonographic （US） signs overlap between benign and malignant nodules. The purpose of this study was to raise a special US sign of benign thyroid nodules, termed the ＂onion skin-liked sign.＂ Methods： Twenty-seven patients with 27 nodules who shrank naturally and the ＂onion skin-liked sign＂ appeared on the final US images were enrolled in the study. The ultrasound characters and risk stratifications at the start and end of observation were compared. Then, thirty goiters with fibrosis and thirty papillary thyroid carcinomas （PTC） were randomly selected from the database of our hospital, matched the sizes of 27 nodules at the end point of observation. The differences of ＂onion skin-liked sign＂ between the two groups were analyzed. Results： The average duration of follow-up of 27 nodules was 24.0 ± 12.2 months （range, 12-65 months）. At the end of the follow-up, the size of the nodules decreased on average by 1.26 ± 0.82 cm （range, 0.3-3.4 cm） and calcification was found in 21 nodules, compared with only 2 nodules with calcification at the start of the follow-up. In addition, only negligible or no blood flow signal could be detected at the periphery of all the nodules and 100% （27/27） were high suspicion at the end of observation. In matched groups, all PTC showed high suspicion of malignancy, 18/30 （60%） goiters with fibrosis were high suspicion and 11/30 （37%） were intermediate suspicion. Twenty-two patients in the group of nodular fibrosis presented ＂onion skin-liked sign,＂ which was not shown in any patient of PTC group. The sensitivity, specificity, positive predictive value, and negative predictive value of ＂onion skin-liked sign＂ in predicting nodular goiter with fibrosis were 73.3%, 100%, 100%, and 78.9%, respectively. Conclusions： The ＂onion skin-liked sign＂ was a characteristic US feature of benign thyroid nodules detected in the follow-up of thyroid nodules. It is useful to differentiate PTCs and nodular goiters with fibrosis.

Negative Correlation of Serum Hepatitis B Surface Antigen and Hepatitis B e Antigen Levels with the Severity of Liver Inflammation in Treatment-naive Patients with Chronic Hepatitis B Virus Infection

Background： Estimating the grades of liver inflammation is critical in the determination ofantiviral therapy in patients chronically infected with hepatitis B virus （HBV）. The aim of this study was to investigate the correlation ofserum levels of hepatitis B surface antigen （HBsAg） and hepatitis B e antigen （HBeAg） with the liver inflammation grades in treatment-naTve patients with chronic HBV infection. Methods： We retrospectively enrolled 584 treatment-na＇l＇ve HBeAg-positive patients who underwent liver biopsy in Ditan Hospital from January 2008 to January 2016. Based on the severity of liver inflammation, the patients were divided into minimal, mild, and moderate groups. SPSS software was used lbr statistical analysis of all relevant data. Results： The liver histological examinations showed that 324, 194, and 66 patients had minimal, mild, and moderate liver inflammation, respectively. The median age of the three groups was 30, 33, and 38 years, respectively （X2 =26.00, P 〈 0.001 ）. The median HBsAg levels in minimal, mild, and moderate inflammation groups were 4.40, 4.16, and 3.67 log U/ml, respectively, and the median HBeAg levels in the three groups were 3.12, 2.99, and 1.86 log sample/cutoff. respectively; both antigens tended to decrease as the grade of inflammation increased （X2 = 99.68 and X2 =99.23, respectively; both P 〈 0.001 ）. The cutoff values of receiver operating characteristic curve in the age, HBsAg and HBeAg levels were 36 years, 4.31 log U/ml, and 2.86 Iog S/CO, respectively, 1 to distinguish minimal grade and other grades of treatment-naTve HBeAg-positive patients with chronic HBV infection. Conclusions： Serum HBsAg and HBeAg quantitation might gradually decrease with aggravated liver inflammation and the corresponding cutoff values rnight help us to distinguish rninimal grades and other grades and detect those who do not need antiviral therapy in treatment-naive HBeAg-positive patients with chronic HBV infection.