慢性重型乙型肝炎患者HBV DNA前C/BCP区突变基因分析

Analysis of genetic mutation of HBV precore/basal core promoter in patients with chronic severe hepatitis B

目的分析慢性重型乙型肝炎(慢重乙肝)患者HBV DNA前C区和基本核心启动子(前C/BCP)区突变特点与意义。方法收集87例慢重乙肝和196例慢性乙型肝炎(慢乙肝)患者血清,提取HBV DNA,用巢式PCR扩增HBV DNA前C/BCP区基因,PCR产物进行DNA测序,用NBI软件比对结果,重点分析G1896、G1862、G1899、A1762、G1764、T17536个位点突变。结果慢重乙肝组和慢乙肝组6个位点突变全阴率分别为3.4%和28.1%(P<0.01);慢重乙肝组在其中5个位点上的突变检出率显著高于慢乙肝组。此外,慢重乙肝组和慢乙肝组≥三联突变检出率分别为56.3%和35.2%(P<0.01),≥四联突变检出率分别为25.3%和8.7%(P<0.01),插入/缺失突变检出率分别为10.3%和1.0%(P<0.01)。结论HBV DNA前C/BCP区基因突变发生频率的增加与慢乙肝发生重症化相关,结合临床资料分析突变的意义将有助于认识慢乙肝重症化的发生机制。

Objective To analyze the characterization and significance of genetic mutation of HBV precore/basal core promoter （BCP） in patients with chronic severe hepatitis B （CSHB）. Methods Sera from 87 patients with CSHB and 196 patients with chronic hepatitis B （CHB） were collected for extraction of HBV DNA. The precore / BCP gene fragment was amplified by nested PCR and analyzed by direct DNA sequencing. The data were processed by NBI software. Substitutions at G1896, G1862, G1899, A1762, G1764 and T1753 were specially focused on. Results Only 3.4% of CSHB patients were mutation-negative at all 6 sites, markedly lower than CHB patients （28.1%, P〈0.01）. At five of the 6 sites, the increase in mutation occurrence was statistically significant in CSHB patients compared with CHB patients, with 56.3% vs 35.2% （P〈0.01） for ≥three-site substitution, 25.3% vs 8.7% （P〈0.01） for ≥four-site substitution, and 10.3% vs 1.0% （P〈0.01） for insertion/deletion. Conclusions Mutation accumulation in HBV precore/BCP regions is associated with CSHB. Screening the mutations combined with clinical data analysis is valuable for understanding the pathogenesis of CSHB.