The BTLA/HVEM inhibitory pathway appears to be particularly important for balancing immune response and tolerance in autoimmune diseases, tumor development, transplantation, allergy and, in particular, the pathogenesis of viral infection
[16–18]. Recent studies have suggested that the BTLA/HVEM interaction functionally regulates CMV-specific T cell functions
. Our previous work has also demonstrated that BTLA signaling can maintain the survival of macrophages that are infected with MHV-3, resulting in virus distribution and tissue damage following MHV-3 infection
. ACLF is a severe clinical syndrome characterized by coagulopathy, jaundice and hepatic encephalopathy [19, 20]. Most ACLF cases in China are associated with HBV infection. In this study, we extended our research and examined the expression of BTLA/HVEM in four cases of HBV-ACLF patients. Immunohistochemical analysis revealed that the expression of BTLA was dramatically enhanced in liver samples from these HBV-ACLF patients, and the positive cells were distributed throughout the entire tissue. However, no positive cells were observed in the liver tissue of CHB patients or healthy individuals. The phenotypes of BTLA+ cells were further analyzed by immunofluorescence double staining, and the results demonstrated that BTLA was expressed on CK-18+ epithelial cells, CD31+ endothelial cells, CD68+ macrophages, CD56+ NK cells, CD16+ monocytes, CD3+ CD8+ T cells, and Foxp3+ regulatory T cells (Tregs). Although the limitation of HBV-ACLF cases was included here, the expression of BTLA was observed on all of these four cases and these data strongly indicates that BTLA may actively participate in regulating the pathogenesis of this disease.
HVEM, the ligand of BTLA, is a novel member of the TNF receptor superfamily, and its mRNA is broadly expressed in both lymphoid and non-lymphoid tissues
. The expression and distribution of HVEM in liver biopsies from HBV and HBV-ACLF patients has not previously been reported. In this study, our data indicated that HVEM was found in sections from HBV-ACLF and was restricted to the damaged bile ducts and certain infiltrating cells, whereas HVEM was absent in sections from normal individuals. Assessments using immunofluorescence double staining further determined that HVEM was expressed on CK-18+ epithelial cells and CD68+ macrophages but completely absent on CD31+ endothelial cells, CD16+ monocytes, and CD3+ CD8+ T cells. These results demonstrated that HVEM was induced in certain antigen-presenting cells (APCs) in these patients.
Our previous work has revealed that the expression of several members of the B7 superfamily, including PD-L1 (B7-H1), PD-L2 (B7-DC), B7-H3 and B7-H4, was found in liver sections from HBV-ACLF
[14, 15]. PD-L1 and PD-L2 are two immunoregulatory molecules belonging to the B7 superfamily that were identified as ligands for PD-1. The binding of PD-L1 or PD-L2 with PD-1 activates a signaling pathway that inhibits T cell responses
[21, 22]. B7-H3 is a type I transmembrane protein that is expressed on activated macrophages, dendritic cells, monocytes and several types of cells in non-lymphoid tissues
. B7-H3 promotes the proliferation, cytotoxicity and IFN-γ production of T cells by crosslinking with its hypothetical receptor and triggering the receptor-mediated expression of myeloid cell-like transcript 2 (TLT-2) on activated T cells
[23, 24]. B7-H4 is a GPI-linked protein, and mRNA encoding B7-H4 is widely distributed in murine and human peripheral tissues
[25, 26]. B7-H4−/− mice exhibited augmented Th1 responses and displayed lowered parasite burdens upon Leishmania major and Listeria monocytogenes infection compared with wild type mice, suggesting an inhibitory role for B7-H4 in T cell responses
[27, 28]. In this study, we also detected the relationships between HVEM and these members of the B7 superfamily in sections from HBV-ACLF, and the study results demonstrated that HVEM was co-expressed with all of these molecules.
FGL2 is a critical molecule that promotes fibrinogen deposition, which, in turn, activates the coagulation cascades and thereby induces PCA, causing hepatocellular necrosis during the development of fulminant virus hepatitis
. High levels of FGL2+ cells that are distributed throughout the liver have also been observed in HBV-ALCF patients. Phenotypic analyses indicated that the expression of FGL2 was mainly localized to infiltrating CD68+ macrophages, CK-18+ bile ducts, and CD31+ capillaries. Moreover, both BTLA and HVEM were co-expressed with FGL2, as assessed by immunofluorescence double staining.
To the best of our knowledge, this report is the first to investigate the expression of BTLA and its ligand, HVEM, in liver sections from HBV-ACLF patients. Our results indicate that these molecules might be useful diagnostic biomarkers for HBV-ACLF, and an understanding of the functional roles of these molecules could aid in the development of novel strategies for disease diagnosis or immunotherapy.