乙型肝炎病毒C基因变异对人白细胞抗原表达的影响

Influence of HBV/C mutation on HLA expression of host cells

目的阐明我国常见的 ＨＢＶ／Ｃ变异对宿主细胞人类白细胞抗原（ｈｕｍａｎ ｌｅｕｋｏｃｙｔｅ ａｎｔｉｇｅｎ， ＨＬＡ）表达的影响。方法构建携带野生型和变异型ＨＢＶ／Ｃ基因的真核细胞表达载体，转染ＨｅｐＧ_２细胞，鉴定目的基因在宿主细胞中的表达，检测ＨＬＡ－ Ｉ、 ＨＬＡ－ＤＲ在宿主细胞膜上的表达。结果 ＰＣＲ和ｗｅｓｔｅｒｎ ｂｌｏｔ分析能分别检测到目的ＤＮＡ片段和ＨＢｃＡｇ的表达。约１００％的ＨｅｐＧ_２细胞表达ＨＬＡ—Ｉ，但表达的密度不同： ＨｅｐＧ_２的平均荧光密度为５７．８，空载体宿主细胞５４．３与ＨｅｐＧ_２细胞没有显著差别；野生株宿主细胞明显下降为３１．２；变异株 Ｖ_６０、 Ｇ_８７和 Ｌ_９７较野生株升高，分别为 ４３．０、 ５４．０和 ６９．４，尤以 Ｌ_９７，最为显著，较 ＨｅｐＧ_２高 １１．６。 ＨＬＡ－ＤＲ在各个宿主细胞均无显著表达。结论ＨＢＶ／Ｃ基因在宿主细胞内的复制和表达能够直接影响宿主细胞ＨＬＡ—Ｉ的表达密度。变异株对宿主细胞ＨＬＡ－Ｉ的影响与野生株不同，可能与病毒变异导致病变活动相关。

Objective To study the influence of HBV/C mutation on host cellular HLA expression. Methods HBV expression vectors carrying wild or variant HBV/C gene were constructed and transferred into HepG2 cells. HBV/C gene expression on the host cells was identified. The expression of HLA-I and -DR on the host cell membrane was detected. Results DNA segments similar with HBV/C gene size and HBcAg were detected by PCR and Western blot analysis, respectively. Almost all HepG_2 cells expressed HLA--I but not HLA- DR. The fluorescence intensity of HLA- I expression on host cells was different: HepG_2 was 57.8 and wild vector 54.3. The wild HBV/C gene declined significantly to 31.2. While V60, G87 and L97 increased to 43.0, 54.0 and 69.4, respectively. L_97 was greatly increased with 11.6 higher than HepG2. HLA-DR had no significant expression. Conclusion Replication and expression of HBV/C gene can influence HLA-I expression on the host cells directly. Influence of variant gene on HLA-I expression is different from the wild gene. This might relate with the exacerbation of diseases resulting from HBV mutation.