Brind A, Jiang J, Samuel D, Gigou M, Feray C, Bréchot C, Kremsdorf D
INSERM U370, CHU Necker, Paris, France.
J Hepatol. 1997 Feb;26(2):228-35. doi: 10.1016/s0168-8278(97)80035-9.
BACKGROUND/AIMS: Despite anti-HBs immunoglobulin therapy, hepatitis B virus (HBV) infection recurs in a high proportion of patients transplanted for HBsAg positive and serum HBV DNA negative chronic liver disease. The contribution of HBV genetic variability to disease recurrence has not been yet thoroughly addressed. We have therefore undertaken a detailed comparison of preS/S and preC/C sequences in two selected patients with recurrence of HBsAg and HBV DNA after transplantation.
PreS/S and preC/C regions were amplified by PCR from the serum, peripheral blood mononuclear cell (PBMC) and liver tissues of two patients transplanted for end stage HBV-related cirrhosis. Samples were taken both pre- and post-transplantation. HBV-sequences from four to nine clones were determined and compared.
A mixing of different HBV DNA molecules was observed within and between serum, liver and PBMC samples. Sequences from both patients showed mutations in the preC region which abolished HBeAg secretion, and in the preS2 initiation codon which prevented preS2 envelope protein production. In addition, for both patients, deletions in the preS2 domain (3 and 21 base pairs) led to the expression of modified preS1 envelope protein. For one patient, the predominant HBs protein sequence found in the PBMC before transplantation showed four specific mutations. One of these mutations was in the "a" determinant (codon 144, asparagine to glycine change) of the major envelope protein. These mutations were not detected, as predominant mutations, in the liver and serum pre-orthotopic liver transplant samples. In contrast, after liver transplantation, this was the major form identified in serum, liver and PBMC.
Our results have shown the selection of different HBV DNA molecules in liver and mononuclear cells. In addition, they provide direct evidence for the role of PBMC in the infection of liver grafts and support the hypothesis that infection of PBMC might lead to selection of HBV variants which would escape immune therapy. Finally, we provide in vivo evidence for reinfection of the liver by HBV particles lacking preS2 envelope protein expression.
背景/目的：尽管采用了抗-HBs免疫球蛋白治疗，但在因HBsAg阳性且血清HBV DNA阴性的慢性肝病而接受移植的患者中，相当一部分会出现乙型肝炎病毒（HBV）感染复发。HBV基因变异性对疾病复发的影响尚未得到充分研究。因此，我们对两名移植后出现HBsAg和HBV DNA复发的患者的前S/S和前C/C序列进行了详细比较。
通过PCR从两名因终末期HBV相关肝硬化接受移植患者的血清、外周血单核细胞（PBMC）和肝组织中扩增前S/S和前C/C区域。在移植前后均采集样本。测定并比较4至9个克隆的HBV序列。
在血清、肝脏和PBMC样本内部及之间均观察到不同HBV DNA分子的混合。两名患者的序列均显示前C区域发生突变，导致HBeAg分泌缺失，前S2起始密码子发生突变，阻止前S2包膜蛋白产生。此外，两名患者的前S2结构域均出现缺失（3和21个碱基对），导致修饰的前S1包膜蛋白表达。对于一名患者，移植前在PBMC中发现的主要HBs蛋白序列显示有四个特定突变。其中一个突变位于主要包膜蛋白的“a”决定簇（第144密码子，天冬酰胺变为甘氨酸）。在原位肝移植前的肝脏和血清样本中，这些突变并非主要突变。相反，肝移植后，这是在血清、肝脏和PBMC中鉴定出的主要形式。
我们的结果显示了肝脏和单核细胞中不同HBV DNA分子的选择。此外，它们为PBMC在肝移植感染中的作用提供了直接证据，并支持PBMC感染可能导致逃避免疫治疗的HBV变异体选择这一假说。最后，我们为缺乏前S2包膜蛋白表达的HBV颗粒再次感染肝脏提供了体内证据。
