巨噬细胞与乙型肝炎相关性肝病肝移植后乙型肝炎复发的联系

Relationship between macrophages and hepatitis B recurrence in hepatitis B-related patients after liver transplantation

背景:有文献报道外周血单个核细胞存在HBV-CCCDNA,那么复发的乙型肝炎病毒是否可能来自被巨噬细胞吞噬但未完全清除的乙型肝炎病毒DNA颗粒?目的:探讨人巨噬细胞与乙型肝炎相关性肝病肝移植后乙型肝炎复发的联系。方法:采集13例因乙型肝炎致肝硬化、肝癌等终末期肝病行同种异体肝移植后乙型肝炎复发患者的抗凝静脉血,另采集14例肝移植后乙型肝炎未复发患者外周血作为对照。Ficoll密度梯度离心法分离上述标本得外周血单个核淋巴细胞,含体积分数10%小牛血清的RPMI1640培养基和重组人粒-巨噬细胞集落刺激因子体外诱导培养7d,得贴壁细胞。流式细胞技术检测细胞表面特征性标志抗体CD14以鉴定贴壁细胞性质。荧光定量PCR检测培养所得巨噬细胞乙型肝炎病毒共价、环状、闭合DNA(HBV-CCCDNA)。结果与结论:13例乙型肝炎相关性肝病肝移植后复发患者巨噬细胞内HBV-CCCDNA阳性率69.23%,14例未复发患者中发现1例阳性,阳性率7.14%,两组比较差异有显著性意义(P<0.01)。提示肝移植后乙型肝炎复发可能与其巨噬细胞中的HBV-CCCDNA有关。

BACKGROUND：HBV-CCCDNA existing in peripheral blood mononuclear cell has been reported in literatures, then, whether the recurrent hepatitis B （HBV） possibly comes from the DNA grains which are phagocytized by macrophages but not clear wholly. OBJECTIVE：To investigate the relationship between macrophages and HBV recurrence in HBV-related patients after liver transplantation. METHODS：Venous blood with anticoagulation was extracted from 13 different HBV-related patients whose hepatitis B recurred after liver transplantation, while, we collected 14 examples from patients whose HBV was negative after liver transplantation for comparison. These examples were separated though Ficoll-Hypaque density centrifugation to get PBMC, then PBMC was cultured for 7 days with rhGM-CSF and RPMI-1640 which contained 10% calf serum to get adherent cells. Special antibody CD-14 was detected with flow cytometry to assay the adherent cells nature. HBV-CCCDNA within the cells was detected though fluorescent-quantitative PCR. RESULTS AND CONCLUSION：Of 13 patients who had HBV again after liver transplantation, 69.23% were HBV-CCCDNA positive within macrophages. Only 1 patient of 14 whose HBV did not recur, and its HBV- CCCDNA was positive, that was 7.14% （P 0.01）. There is probably a relationship between HBV recurrence after liver transplantation and HBV CCCDNA in macrophages.