乙型肝炎自然进程中血清HBV RNA的动态变化及其与肝组织HBV cccDNA的关系  

The Dynamic Changes of Serum HBV RNA in the Natural Course of Hepatitis B and Its Relationship with HBV cccDNA in Liver Tissues

作　　者：李韦杰 徐东平 韩佳琪 李亚楠 周淳[1] 焦炳欣[1] 周茹[1] 黄鸿鑫 王雅杰 LI Wei-jie;XU Dong-ping;HAN Jia-qi;LI Ya-nan;ZHOU Chun;JIAO Bing-xin;ZHOU Ru;HUANG Hong-xin;WANG Ya-jie(Department of Clinical Laboratory,Beijing Ditan Hospital,Capital Medical University,Beijing 100015,China;The Fifth Medical Center of the Chinese People′s Liberation Army,Beijing 100039,China;ICU,First Affiliated Hospital of Tsinghua University,Beijing 100016,China;Department of Microbiology,Peking University,Beijing 100191,China)

机构地区：[1]首都医科大学附属北京地坛医院检验科,北京100015 [2]中国人民解放军第五医学中心,北京100039 [3]清华大学第一附属医院ICU,北京100016 [4]北京大学医学部微生物教研室,北京100191

出　　处：《标记免疫分析与临床》2019年第9期1445-1451,1460,共8页Labeled Immunoassays and Clinical Medicine

基　　金：首都医科大学附属北京地坛医院院内课题(编号:DTYM201604);吴阶平医学基金会临床科研专项资助基金(编号:320.6750.19019)

摘　　要：目的定量检测慢性乙型肝炎患者血清乙肝病毒RNA和肝组织乙肝病毒共价闭合环状DNA(HBV cccDNA),分析两种定量指标动态变化及其相关性。方法应用PSAD酶消化+滚环扩增+跨缺口实时荧光PCR方法,定量检测患者甲醛固定石蜡包埋肝组织HBV cccDNA(cccDNA copies/cell),用qPCR定量检测血清样本中HBV RNA的表达水平。结果在慢性乙型肝炎免疫耐受期、免疫清除期、低复制期、HBeAg阴性期患者的血清HBV RNA水平中位数分别是1.01×10^8 copies/mL、3.18×10^6 copies/mL、5.56×10^3 copies/mL和3.8×10^5 copies/mL。在慢性乙型肝炎中,患者血清RNA水平只在免疫清除期与肝组织cccDNA具有相关性(r=0.328,P=0.016),两者在免疫耐受期、低复制期、HBeAg阴性期均没有相关性(r=-0.003,P=0.995;r=-0.581,P=0.171;r=0.041,P=0.870)。HBV RNA复制效率,定义为血清中乙肝病毒RNA载量与肝组织中HBV cccDNA的比值(所有的统计量都经过了对数值转换),其在急性期肝炎患者中明显地低于慢性期患者(0.37 vs.0.99,P<0.05)。在慢性乙型肝炎的免疫耐受期、e抗原阴性期HBV RNA复制效率显著高于低复制期(1.005 vs.0.66,P=0.025;0.90 vs.0.66,P=0.048),在其他几期中差异没有统计学意义。结论在未经治疗的乙型肝炎患者中,血清HBV RNA水平可反映处于慢性乙肝免疫清除期患者肝组织内HBV cccDNA水平,但不能反映慢性乙肝免疫耐受期、低复制期和HBeAg阴性期患者肝组织内HBV cccDNA水平;急性乙肝免疫清除期患者的HBV RNA复制效率明显低于慢性期患者。Objective To quantitate the levels of serum hepatitis B virus RNA and intrahepatic covalently closed circular DNA(cccDNA)in patients with hepatitis B infection,then analyze the dynamic changes and correlation between the levels of serum HBV RNA and intrahepatic HBV cccDNA.Methods The quantitation of intrahepatic HBV cccDNA was measured using real-time PCR with PSAD digestion+rolling circle amplification.Formaldehyde-fixed paraffin-embedded liver tissues were used for HBV cccDNA detection.The levels of HBV RNA was measured by qPCR using serum samples.Results The median level of serum HBV RNA were 1.01×10^8 copies/mL,3.18×10^6 copies/mL,5.56×10^3 copies/mL,and 3.8×10^5 copies/mL for individuals with chronic hepatitis B infection during immune tolerance,immune clearance,low replication,and HBeAg-negative phases,respectively.In patients with chronic hepatitis B infection,serum RNA levels were only correlated with intrahepatic cccDNA during immune clearance period(r=0.328,P=0.016).There was no correlation of them at the phases of immune tolerance,low replication,nor in HBeAg-negative patients(r=-0.003,P=0.995;r=0.581,P=0.171;r=0.041,P=0.870).The replication efficiency of HBV RNA was defined as the ratio of serum HBV RNA level to hepatic HBV cccDNA level(all statistics are log-transformed),which was significantly lower in patients with acute hepatitis B virus infection than in patients with chronic hepatitis B infection(0.37 vs.0.99,P<0.05).Patients during immune tolerance and HBeAg-negative phase showed significantly higher replication efficiency of HBV RNA than the low replication phase of chronic hepatitis B infection(1.005 vs.0.66,P=0.025;0.90 vs.0.66,P=0.048),and no significant difference was found in other phases.Conclusion In treatment-na ve patients with hepatitis B infection,the levels of serum HBV RNA are correlated with intrahepatic HBV cccDNA levels during immune clearance phase,but not in acute or chronic hepatitis B patients during phases of immune tolerance,low replication,nor HBeAg-negative.The repl

关 键 词：乙型肝炎病毒 HBV RNA 共价闭合环状DNA 乙肝病毒E抗原 乙肝病毒自然感染进程 

分 类 号：R51[医药卫生—内科学]