我国慢性乙型肝炎及肝癌患者中乙型肝炎病毒核心基因内缺失突变株的研究

Studies on HBV core internal deletion mutants in chronic hepatitis B and hepatocellular cancer patients

目的 研究乙型肝炎（ 乙肝） 病毒核心基因内缺失突变株（ Ｃ Ｉ Ｄ） 在慢性乙肝及肝癌患者中存在的状态及意义。方法 提取标本 Ｄ Ｎ Ａ 作 Ｓｏｕｔｈｅｒｎ 印迹及杂交，或用 Ｐ Ｃ Ｒ 扩增核心基因，分析片段大小，部分作克隆及测定。结果 ３５ 例慢性乙肝患者血清中３０ 例（８５ ％ ） 核心基因为阳性。３０例中３ 例（１０ ％ ） 除有正常大小片段外，还有相对小分子质量片段。１５ 例肝癌患者肝癌组织 Ｐ Ｃ Ｒ Ｃ 基因阳性为９ 例（６０ ％ ） ，未见小片段。肝癌组织中２ 份存在 Ｃ Ｉ Ｄ（２／９ ，２２ ％ ） 。克隆测序证明有大小不等的缺失突变，自６ ～３２４ 个核苷酸不等的符合框架的缺失，位于 Ｃ 基因的中段。结论 我国慢性乙肝患者中 Ｃ Ｉ Ｄ 株阳性率仅１０ ％ ，不是构成我国多数乙肝患者持续性病毒感染的主要原因。肝癌组织中 Ｃ Ｉ Ｄ 的阳性率高于乙肝患者血清的阳性率， Ｃ Ｉ Ｄ 在肝癌中的作用值得进一步研究。

Objective To study the status and significance of hepatitis B virus core internal deletion (CID) mutants in chronic hepatitis B and hepatocellular carcinoma patients. Methods DNA was extracted from specimens and studied by Southern blotting and hybridization to HBV core probe. DNA was also stu died by PCR and the core gene analyzed by checking for low molecularweight fragments, followed by sequencing.Results Thirty out of 35(85%) serum samples from chronic hepatitis B patients were core gene positive by PCR, and three showed an extra low molecularweight band (3/30, 10%). Nine out of the 15 sera from hepatocellular carcinoma (HCC) patients were core gene positive by PCR, however, no low molecular weight bands were detected. In two HCC tissues, CID mutants were detected (2/9,22%), and by cloning and sequencing, CID mutants were identified with various length of deletions, which ranged from 6 to 324 nucleotides. The deleted fragments were located in the middle of the core gene.Conclusion The incidence of HBV CID in chronic hepatitis B patients studied was 10%, which showed that CID did not seem to be the main mechanism leading to viral persistence in Chinese patients. The incidence of CID is higher in HCC tissues than that in serum samples from hepatitis B and HCC patients, suggesting that implications of CID in HCC merit further study.(Shanghai Med J, 1999,22∶595 597)