抗原抗体复合物型治疗性疫苗(乙克)临床研究中患者血清乙型肝炎病毒表面抗原下降的分析与启示

近年来全球慢性乙型肝炎(chronic hepatitis B,CHB)防治指南提出了“功能性治愈”(functional cure)的概念,即患者经过治疗达到血清乙型肝炎病毒表面抗原(hepatitis B virus surface antigen,HBsAg)消失,但现有抗病毒治疗很难实现这一目标。本研究对既往临床试验中经抗原抗体复合物型治疗性疫苗(乙克)治疗后的CHB患者HBsAg下降情况进行了归纳分析,结果显示,经乙克治疗随访后达到乙型肝炎e抗原(hepatitis B e antigen,HBeAg)血清学转换者的HBsAg下降高达0.95log₁₀IU/mL,显著高于未达到HBeAg血清学转换者的0.32log₁₀IU/mL(P<0.01),而经氢氧化铝佐剂治疗随访后发生HBeAg血清学转换(0.49log₁₀IU/mL)者与未发生HBeAg血清学转换者(0.36log₁₀IU/mL)之间HBsAg下降无统计学差异。乙克组治疗过程中,丙氨酸氨基转移酶(alanine aminotransferase,ALT)骤升(ALT flare)在HBsAg下降>1.0log₁₀IU/mL者中较多见,氢氧化铝组未观察到此现象。回归分析显示,乙克治疗后HBsAg下降的影响因素有患者出现HBeAg血清学转换、感染的HBV为B基因型、治疗过程中ALT出现10倍增高,以及基线血清HBsAg为高水平。结果提示,乙克诱导的特异性免疫对降低CHB患者血清HBsAg水平有一定效果,采用“抗病毒药物治疗＋针对HBsAg的中和性抗体被动免疫＋乙克主动免疫”的“三明治”治疗策略可能会提高“功能性治愈”率。     

干扰素-α治疗HBeAg阴性慢性乙型肝炎患者的早期疗效及影响因素分析

目的探讨HBeAg阴性慢性乙型肝炎(CHB)患者干扰素-α(IFN-α)早期疗效的影响因素。方法收集40例HBeAg阴性CHB患者,检测IFN-α治疗前及治疗12周时的ALT、HBV DNA、HBsAg、HBsAb、HBeAg、HBeAb、HBcAb;分析年龄、性别、ALT基线水平、HBV DNA基线水平对IFN-α疗效的影响。结果性别组间、不同年龄段组间、不同ALT基线水平组间、不同HBV DNA基线水平组间比较IFN疗效差异无统计学意义(P>0.05)。结论性别、年龄、ALT基线水平、HBV DNA基线水平、年龄不可以作为预测IFN-α治疗HBeAg阴性慢性乙型肝炎患者的早期疗效指标。     

聚乙二醇干扰素α 2a治疗HBeAg阳性慢性乙型肝炎患者的基线评分预测模型建立

目的构建一个基线评分预测模型,用于预测HBeAg阳性慢性乙型肝炎(CHB)患者接受聚乙二醇干扰素α 2a(Peg IFN α 2a)的治疗效果。方法收集2013年1月至2017年2月在温州医科大学附属第一医院治疗的242例初治HBeAg阳性CHB患者为研究对象,入选患者均给予足量Peg IFN α 2a治疗,回顾性分析患者治疗前的基线临床数据,根据患者治疗48周后是否实现HBeAg转阴分为应答组和非应答组,筛选两组间差异指标,利用受试者工作特征曲线确定2个截断值并赋分,然后使用Logistic回归确定建立模型的指标,并根据OR值调整分值并建立模型。结果最终纳入170例HBeAg阳性CHB患者,其中应答组64例,非应答组106例。通过Logistic回归最终筛选得到5个指标,分别为HBeAg、HBV DNA、TSH、ALT和AFP。3区段HBeAg OR值赋分结果为0、1.5、3分,其余4个指标均为0、1、2分,据此建立一个基线评分预测模型,其曲线下面积为0.759。结论本研究成功建立了一个基线评分预测模型,其中包含HBeAg、HBV DNA、ALT、AFP和TSH这5个指标,可较好预测HBeAg阳性CHB患者接受Peg IFN治疗的应答情况。     

聚乙二醇干扰素α-2b治疗61例e抗原阳性慢性乙型肝炎临床分析

目的探讨聚乙二醇化干扰素α-2b治疗e抗原(HBeAg)阳性慢性乙型肝炎的临床疗效和不良反应,同时探讨影响应答的相关因素。方法用聚乙二醇干扰素α-2b治疗61例HBeAg阳性慢性乙型肝炎,治疗期间定期监测血常规、生物化学指标、病毒学标志、甲状腺功能等。结果 48周,表面抗原(HBsAg)和HBeAg血清学转换率分别为5.26%、31.2%;血清HBV DNA阴转率为59.6%;ALT复常率为64.9%。随访半年,HBV DNA复发率为17.4%;HBeAg血清学转换者维持应答。治疗前ALT>5 ULN时,48周时HBeAg血清学转换率和HBV DNA阴转率明显高于ALT<5 ULN时,二者比较差异有统计学意义;而血清HBV DNA水平与其无明显相关性。结论聚乙二醇化干扰素α-2b具有免疫调节和抗病毒双重作用,48周时HBeAg血清学转换率和HBV DNA阴转率与治疗前血清ALT水平与相关,停药后具有持续应答效应。     

阿德福韦酯治疗乙型肝炎的疗效及安全性分析

目的探讨阿德福韦酯治疗慢性乙型病毒性肝炎患者的临床疗效及安全性。方法将我院2010年2月~2014年4月收治的90例慢性乙型肝炎患者给予阿德福韦酯治疗,并定期监测(丙氨酸氨基酸转移酶)ALT,HBV DNA,HBeAg和抗-HBe,观察其病毒学应答率、血清学转换率、病毒学突破率和复发率;分析抗病毒治疗基线ALT(u/L)与血清学转换率。结果 (1)6个月累计病毒学应答率91.11%(82/90),1年为100%;(2)HBeAg阳性者1、2、3年累计HBeAg血清学转换率分别为27.91%、34.69%、40.33%;(3)基线ALT≥5×正常值上限(u/L)的患者3年发生HBeAg血清学转换率高于基线ALT5×正常值上限(u/L)的患者(50.00%、42.22%,χ2=5.527,P=0.013)。结论基线ALT水平可作为3年HBeAg血清学转换率预测指标;抗病毒治疗达我国《指南》终点标准仅仅是基本的停药标准,部分病人达标后仍有复发风险;阿德福韦酯治疗慢性乙型肝炎疗效确切,安全性高,值得临床推广应用。     

肝细胞癌患者血清外泌体microRNA表达鉴定

目的:探讨肝癌细胞外泌体中差异表达的microRNAs(miRNAs)在肝细胞癌(HCC)诊断中的应用价值。方法:通过高通量测序筛选肝癌细胞外泌体中差异表达的miRNAs。实时定量PCR验证差异表达分子;检测差异表达的miRNAs在健康人(Health)、慢性乙型肝炎患者(CHB)、肝硬化患者(LC)及乙型肝炎病毒阳性的肝细胞癌患者(HCC)血清外泌体中的表达。结果:高通量测序筛选到肝癌细胞外泌体中差异表达的miRNA共88种,其中58种表达上调,30种表达下调。选择其中8种差异表达的miRNAs进行q RT-PCR验证,结果显示,此8种miRNAs在细胞上清外泌体、细胞内、癌与癌旁组织中的表达趋势与测序结果一致。miR-221-3p和miR-224-5p在HCC组外泌体中的表达水平显著高于Health组、CHB组和LC组(P0.01),miR-124-3p和let-7a-5p在HCC组外泌体中的表达水平显著低于其他各组(P0.05)。四个组中,miR-21-5p、miR-191-5p、miR-34a-5p和miR-122-5p的表达水平不存在显著性差异(P0.05)。结论:血清外泌体中的miR-221-3p、miR-224-5p、miR-124-3p和let-7a-5p可能成为肝细胞癌的候选标志物。     

不同血清量标本外泌体提取及微小RNA的检测结果比较

本文旨在探讨Qiagen exoRNeasy Serum/Plasma试剂盒提取血清标本中外泌体所需的最适血清量。采用Qiagen exoRNeasy Serum/Plasma 试剂盒分别对250、500、1 000 μL血清中的外泌体进行抽提,使用透射电子显微镜检测分离的外泌体大小和形态,蛋白质免疫印迹法检测外泌体蛋白标记CD63和TSG101的表达,实时荧光定量聚合酶链反应(polymerase chain reaction,PCR)检测外泌体中微小RNA-122(microRNA-122,miR-122)的表达。结果显示,透射电子显微镜下可见血清外泌体呈圆形或椭圆形,直径30～150 nm,有完整的膜结构。蛋白免疫印迹法检测外泌体CD63和TSG101阳性。实时荧光定量PCR检测慢性乙型肝炎患者250、500、1 000 μL血清外泌体中miR-122表达量,与正常人相比,分别上调22.44、21.48、20.69倍(P＝0.42)。结果提示,在临床血清样本体积有限的情况下,采用 Qiagen exoRNeasy Serum/Plasma 试剂盒提取血清中外泌体,减少血清量至250 μL也可达到所需实验目的。     

苦参碱联合恩替卡韦抗乙型肝炎病毒感染的作用及其机制

目的:探讨苦参碱联合恩替卡韦治疗乙型肝炎病毒(HBV)感染的疗效并初步探讨其作用机制。方法:选择2014年9月-2016年8月我院收治的HBeAg阳性乙肝患者98例,按治疗方法分为对照组和观察组,每组49例。对照组患者给予恩替卡韦治疗,观察组患者在对照组基础上应用苦参碱联合治疗。分别在治疗后12、24、48周观察和比较两组患者的HBeAg转阴率、ALT复常率、HBV DNA转阴率、血清球蛋白水平。结果:观察组在治疗后24、48周的HBeAg转阴率、ALT复常率、HBV DNA转阴率均显著高于对照组,差异均具有统计学意义(P0.05)。观察组血清球蛋白水平明显高于对照组,差异有统计学意义(P0.05)。两组间不良反应发生情况比较差异无统计学意义(P0.05)。结论:苦参碱联合恩替卡韦对于HBV具有较好的协同治疗作用,且安全性较好,可能与苦参碱降低血清转氨酶水平,增强患者免疫功能有关。     

慢性乙型肝炎潜在治疗靶点和新药研发进展

尽管预防性疫苗显著减少了乙型肝炎病毒(hepatitis B virus,HBV)新发感染,但目前全球仍有超过2.4亿慢性HBV感染者,其中每年因HBV感染相关的终末肝病和肝癌引起的死亡人数高达68万。目前用于慢性乙型肝炎(chronic hepatitis B,CHB)治疗的抗病毒药物包括干扰素和核苷/核苷酸类似物两大类,但均难以实现理想的临床治疗终点,即乙肝表面抗原(HBsAg)阴转或血清学转换。针对CHB患者尚未被满足的巨大医疗需求,国内外团队正在针对HBV生活周期的各个关键步骤以及潜在的宿主因子,尝试研发更为有效的CHB治疗药物,本文简要综述了当前处于临床开发阶段以及部分临床前阶段的CHB候选药物研发进展。     

13039例乙型肝炎患者血清HBV DNA水平流行病学分析

目的了解本地区近四年乙型肝炎患者HBV DNA阳性率与病毒载量分布特点。方法采用荧光定量聚合酶链反应(PCR)检测2012年至2015年间13 039例HBsAg阳性患者HBV DNA载量,并进一步分析24例HBeAg阳性、31例HBeAg阴性初诊乙肝患者4年间接受正规治疗后HBV DNA变化趋势。结果本地区近4年间血清HBV DNA阳性率依次为48.6%、46.18%、38.69%、37.66%,呈逐年下降趋势,差异具有统计学意义(P0.01);各年度病毒载量均值的对数值分别为(5.48±1.56),(5.27±1.72),(5.29±1.68),(5.35±1.85)拷贝/mL。2013年、2014年、2015年分别与2012年相比,差异均有统计学意义(P0.01、P0.01、P0.05),而低病毒载量(即病毒拷贝数的对数值小于5拷贝/mL)患者比例2012年明显大于其他年份,2012年分别与2013年、2014年、2015年相比,差异具有统计学意义(P0.01,P0.01,P0.05);HBeAg阳性/阴性乙肝患者接受正规治疗后,其HBV DNA载量均呈逐年下降趋势,其中第一年下降显著(P0.01)。结论近4年本地区血清HBV DNA阳性率呈逐年下降趋势,且低病毒载量比例明显增加。     

Pegylated-Interferon Alpha Therapy for Treatment-Experienced Chronic Hepatitis B Patients

Background

Studies are limited on pegylated interferon (Peg-IFN) therapy for chronic hepatitis B (CHB) patients who failed or relapsed on previous antiviral therapy.

Objectives

We aimed to investigate the effect of Peg-IFN therapy in treatment-experienced CHB patients.

Study Design

A total of 57 treatment-experienced CHB patients at two medical centers were enrolled. All of the patients were treated with Peg-IFN α-2a at 180 μg weekly for 24 or 48 weeks. The hepatitis B serological markers and viral loads were tested every 3 months until 1 year after stopping Peg-IFN therapy. The endpoints were HBV DNA <2000IU/mL, hepatitis B e antigen (HBeAg) seroconversion, and a hepatitis B surface antigen (HBsAg) loss at 12 months post-treatment.

Results

In HBeAg-positive patients, 25.0%, 29.2%, and 12.5% of the patients achieved HBeAg seroconversion, HBV DNA <2000 IU/mL and a combined response, respectively, at 12 months post-treatment. Prior IFN therapy, a high baseline ALT level, a low creatinine level, undetectable HBV DNA at 12 weeks and a decline in HBV DNA >2 log₁₀ IU/mL at 12 weeks of therapy were factors associated with treatment response. In HBeAg-negative patients, 9.1%, 15.2%, and 6.1% of the patients achieved undetectable HBV DNA, HBV DNA <2000 IU/mL, and an HBsAg loss, respectively, at 12 months post-treatment. No factor was significantly associated with the treatment response in the HBeAg-negative patients. The median HBsAg level declined from 3.4 to 2.6 log₁₀ IU/mL in all the patients, and the 5-year cumulative rate of the HBsAg loss was 9.8% in the HBeAg-negative patients. Overall, none of the patients prematurely discontinued the Peg-IFN therapy.

Conclusions

Peg-IFN re-treatment is effective for a proportion of HBeAg-positive treatment-experienced patients; it has limited efficacy for HBeAg-negative treatment-experienced patients. Peg-IFN might facilitate HBsAg loss in HBeAg-negative treatment-experienced patients.     

Effects of Entecavir on Hepatitis B Virus Covalently Closed Circular DNA in Hepatitis B e Antigen-Positive Patients with Hepatitis B

The aim of this study was to assess the effect of 48-week entecavir therapy on serum and intrahepatic hepatitis B virus, covalently closed circular DNA (HBV cccDNA) levels in hepatitis B e antigen (HBeAg)-positive patients. A total of 120 patients with HBeAg-positive chronic hepatitis were treated with entecavir for 48 weeks. Serum HBV markers, total HBV DNA, and HBV cccDNA levels were measured at baseline and week 48. Biopsies from 20 patients were available for both intrahepatic total HBV DNA and cccDNA testing at these timepoints. HBV cccDNA levels were decreased from a median level of 5.1×10⁶ copies/mL at baseline to a median level of 2.4×10³ copies/mL at week 48. Reduction magnitudes of HBV cccDNA in patients with normalized alanine aminotransferase levels and those undergoing HBeAg seroconversion were significantly greater than those in alanine aminotransferase-abnormal and HBeAg positive patients. Intrahepatic HBV cccDNA was decreased significantly after 48 weeks of treatment, but could not be eradicated. In conclusion, treatment of HBeAg-positive hepatitis B patients with entecavir for 48 weeks decreased serum and intrahepatic HBV cccDNA significantly, and the magnitude of HBV cccDNA reduction was related to total HBV DNA decrease, alanine aminotransferase normalization, and HBeAg seroconversion.     

基线HBsAg和ALT水平是替比夫定治疗HBeAg阳性慢性乙型肝炎患者e抗原血清学转换的重要预测因素

目的评价初治、单药使用替比夫定治疗HBeAg阳性的慢性乙型肝炎（CHB）患者48周的e抗原血清学转换的基线预测因素。方法97例HBeAg阳性CHB患者分别以基线HBsAg、ALT和HBVDNA水平高低分组,对比两组治疗48周时生化学、病毒学和血清学应答情况。结果基线HBsAg-101500IU／mL组e抗原阴转率和血清学转换率均为42．3％,基线HBsAg〉1500IU／mL组分别为20％和17．8％,两组比较差异均有统计学意义（P〈0．05）;基线ALT〉5ULN组e抗原阴转率和血清学转换率均为45．1％,基线ALT05ULN组分别17．4％和15．2％,两组比较差异均有统计学意义（P〈0．01）;基线HBVDNA〈8．0log-10copies／mL组和基线HBVDNA≥8．0log-10copies／mL组e抗原阴转率和血清学转换率相比,差异无统计学意义（P〉0．05）;基线水平HBsAg≤1500IU／mL且ALT〉5ULN的CHB患者共40例作为观察组,其余57例患者作为对照组,治疗48周时观察组e抗原阴转率和血清学转换率均为45％,对照组分别为22．8％和21．1％,两组比较差异均有统计学意义（P〈0．05）。结论基线HBsAg水平≤1500IU／mL和ALT水平〉5ULN的HBeAg阳性慢性乙型肝炎患者,在接受替比夫定治疗48周时,有较高的e抗原转阴率和血清学转换率;基线HBsAg和ALT水平是替比夫定治疗e抗原血清学转换的重要预测因素。     

实时定量RT-PCR对乙型肝炎病毒全长RNA(fRNA)的定量检测

目的:建立一种简便的定量检测慢性乙型肝炎患者血清中终止于聚腺苷酸化位点的乙型肝炎病毒全长RNA(f RNA)的方法。方法:选取53例未治疗的乙型肝炎患者及22例HBs Ag阴性的健康者为研究对象,使用锚定oligo-d T的引物对其血清中f RNA进行实时定量反转录PCR检测,统计分析其与HBV DNA、HBcr Ag和HBe Ag的相关性。结果:对f RNA进行实时荧光定量RT-PCR检测的下线为2.3 log copies/ml,标准曲线的相关系数为0.99(P0.0001)。53例乙型肝炎患者中,29例(54.7%)可以检测到f RNA,22例正常对照中没有检测到f RNA。27例HBe Ag阳性和/或高水平HBV DNA的患者全部检测到f RNA,26例HBe Ag阴性并且低水平HBV DNA的乙型肝炎患者中有2例(7.7%)检测到f RNA(P0.0001)。HBe Ag阳性患者血清中f RNA水平高于HBe Ag阴性患者(5.0±0.3 vs.2.9±0.4 log copies/ml,P0.001)。f RNA与HBV DNA/HBcr Ag具有显著相关性(r=0.905、0.881,P0.0001)。Hayashi's定量分析法I显示f RNA与HBV DNA相关性强于其与HBcr Ag的相关性。结论:与HBV DNA和HBe Ag一样,f RNA可作为常规检测判断HBV的复制水平并指导用药。     

CXCL9 Associated with Sustained Virological Response in Chronic Hepatitis B Patients Receiving Peginterferon Alfa-2a Therapy: A Pilot Study

Background and Aims

There is lack of a practical biomarker to predict sustained virological response (SVR) in chronic hepatitis B (CHB) patients undergoing peginterferon alfa-2a (PEG-IFN). The aim of this pilot study was to identify immunological features associated with SVR.

Methods

Consecutive 74 CHB patients receiving 24 weeks (for hepatitis B e antigen (HBeAg)-positive) or 48 weeks (for HBeAg-negative) PEG-IFN, were prospectively enrolled. Serum HBV viral loads, hepatitis B surface antigen (HBsAg), CXCL9, IFN-γ-inducible protein 10 (IP-10), interferon-gamma (IFN-γ) and transforming growth factor beta (TGF-β) were measured at baseline and week 12. SVR was defined as HBeAg seroconversion combined with viral load <2000 IU/mL in HBeAg-positive (n=36), and viral load <2000 IU/mL in HBeAg-negative patients (n=38) at 48 weeks after the end of treatment.

Results

Nineteen patients (25.7%), 7 in HBeAg-positive and 12 in HBeAg-negative, achieved SVR. There were significant declines of HBV DNA, HBsAg, IP-10 and IFN-γ levels at week 12. In multivariate analysis, pre-treatment CXCL9 >80 pg/mL, HBV DNA <2.5 x 10⁷ IU/mL and on-treatment HBV viral load, HBsAg decline >10% at week 12 were predictors of SVR. The performance of CXCL9 in predicting SVR was good in patients with HBV DNA <2.5 x 10⁷ IU/mL, particularly in HBeAg-negative CHB cases (positive predictive value, PPV= 64.3%).

Conclusions

Pre-treatment CXCL9 level has the potential to select CHB patients who can respond to PEG-IFN, especially in HBeAg-negative patients with low viral loads.     

Consolidation treatment needed for sustained HBsAg-negative response induced by interferon-alpha in HBeAg positive chronic hepatitis B patients

Hepatitis B surface antigen (HBsAg) clearance is considered as functional cure in patients with chronic hepatitis B (CHB). This study aimed to assess the durability of HBsAg clearance achieved by interferon-based therapies in patients with CHB who were originally positive for hepatitis B envelope antigen (HBeAg). In this prospective study, HBeAg-positive CHB patients with confirmed HBsAg loss under interferon-based therapies were enrolled within 12 weeks from end of treatment and followed up for 48 weeks. Virological markers, biochemical indicators, and liver imaging examinations were observed every 3-6 months. Sustained functional cure was analysed as primary outcome. Factor associated with sustained HBsAg loss or reversion was also investigated. The rate of HBsAg loss sustainability was 91.8% (212/231). Patients receiving consolidation treatment for 12-24 weeks or ≥ 24 weeks had higher rates of sustained HBsAg negativity than those receiving consolidation treatment for < 12 weeks (98.3% and 91.2% vs. 86.7%, P=0.068), and the former groups had significantly higher anti-HBs levels than the later (P < 0.05). The cumulative incidence of HBsAg reversion and HBV DNA reversion was 8.2% and 3.9%, respectively. Consolidation treatment of ≥ 12 weeks[odd ratio (OR) 3.318, 95% confidence interval (CI) 1.077-10.224, P=0.037) was a predictor of sustained functional cure, and HBeAg-positivity at cessation of treatment (OR 12.271, 95% CI 1.076-139.919, P=0.043) was a predictor of HBsAg reversion. Interferon-alpha induced functional cure was durable and a consolidation treatment of ≥ 12-24 weeks was needed after HBsAg loss in HBeAg-positive CHB patients.     

α-2-HS-glycoprotein is a potential marker predicting hepatitis B e antigen seroconversion in patients with chronic hepatitis B during treatment with pegylated interferon alfa-2b

The efficacy of interferon (IFN) is limited in about 1/3 of patients with chronic hepatitis B (CHB). We used two-dimensional electrophoresis (2-DE)-based proteomic strategies to identify potential serum markers predicting hepatitis B e antigen (HBeAg) seroconversion in these patients during IFN therapy. Two groups of patients were enrolled: training and validation. In the training group, 2-DE experiments and subsequent identification of altered levels of proteins showed that α-2-HS-glycoprotein, leucine-rich α-2-glycoprotein, and haptoglobin were significantly upregulated as compared with baseline levels in the HBeAg seroconversion group, whereas apolipoprotein C-III precursor, leucine-rich α-2-glycoprotein, and α-albumin were downregulated in the non-seroconversion group. For patients with HBeAg seroconversion in the training group, Western blot analyses showed that α-2-HS-glycoprotein levels in 75% of patients were significantly upregulated at the end of the treatment as compared with baseline levels. Subsequent experiments in the validation group showed that α-2-HS-glycoprotein levels were significantly increased at week 4 in 83.33% of patients in the HBeAg seroconversion group. Dynamic changes in the serum level of α-2-HS-glycoprotein may be a potential early marker for predicting HBeAg seroconversion during IFN treatment for CHB.     

Levels of hepatitis B virus replicative intermediate in serum samples of chronic hepatitis B patients

Hepatitis B virus (HBV) cccDNA levels is an absolute marker of HBV replication in the liver of HBV infected patients. This study aimed to quantify the HBV cccDNA levels in sera and liver tissue samples of treatment naïve patients with chronic hepatitis B. Eighty one chronic hepatitis B (CHB) treatment naïve patients were enrolled from January 2009 to June 2011. Total HBV DNA and HBV cccDNA levels were quantified using sensitive real time PCR assay. The mean age of recruited patients was 34 ± 11.5 years. Fifty four (66.7 %) patients were HBeAg negative. Liver tissue samples were available from 2 HBeAg positive and 21 HBeAg negative CHB patients. The amount of total intrahepatic HBV DNA ranged from 0.09 to 1508.92 copies/cell. The median intrahepatic HBV cccDNA was 0.31 and 0.20 copies/cell in HBeAg positive and HBeAg negative cases, respectively. Serum HBV cccDNA was detectable in 85.2 % HBeAg positive and 48.1 % HBeAg negative CHB patients. Median serum HBV cccDNA was 46,000 and 26,350 copies/mL in HBeAg positive and HBeAg negative subjects, respectively. There was a significant positive correlation between the levels of intrahepatic total HBV DNA and intrahepatic HBV cccDNA (r = 0.533, p = 0.009). A positive correlation was also seen between serum HBV cccDNA levels and serum HBV DNA levels (r = 0.871, p < 0.001). It was concluded that serum HBV cccDNA could be detectable in higher proportion of HBeAg positive patients compared to HBeAg negative patients. Moreover, the median level of serum HBV cccDNA was significantly higher in HBeAg positive patients in contrast to HBeAg negative subjects.     

Downregulation of exosome-encapsulated miR-548c-5p is associated with poor prognosis in colorectal cancer

The role of circulating exosomal microRNAs (miRNAs) in colorectal cancer (CRC) has drawn more and more attention during the past few years. Previously, we have identified several specific miRNAs in serum exosomes as potential CRC biomarkers. However, little is known about the association between exosome-encapsulated miR-548c-5p and outcomes of patients with CRC. In the current study, the expression of serum exosomal miR-548c-5p was investigated by quantitative real-time polymerase chain reaction. Its correlation with CRC prognosis was estimated by Kaplan-Meier survival and log-rank tests. Cox regression analysis based on uni- and multivariate analyses was performed to estimate the relationship of exosome-encapsulated miR-548c-5p with the clinicopathological factors of patients with CRC. Reduced levels of serum exosomal miR-548c-5p were more significant in CRC patients with liver metastasis and at later TNM stage (III/IV tumor stages). Serum exosomal miR-548c-5p could inhibit the proliferation of CRC cells, while the precise molecular mechanisms warranted further elucidation. In addition, decreased levels of serum exosomal miR-548c-5p were independently associated with shorter overall survival in CRC adjusted by age, sex, tumor grade vascular infiltration, TNM stage (III/IV tumor stages) and metastasis (hazard ratio = 3.40, 95% confidence interval 1.02-11.27; P = 0.046). The downregulation of exosomal miR-548c-5p in serum predicts poor prognosis in patients with CRC. Exosomal miR-548c-5p may be a critical biomarker for CRC diagnosis and prognosis.