Introduction Animal studies suggest that sinusoidal endothelial cells and not hepatocytes are the site of cytomegalovirus (CMV) latency and reactivation in the liver and the source of secondary viral spread. Furthermore, murine CMV infection of sinusoidal endothelium is able to break immunotolerance and induce a strong T cell effector response. The aim of this study was to investigate, whether CMV infection of human hepatic sinusoidal endothelial cells (HSEC) modulates the ability of the liver to recruit and activate lymphocyte.

Methods Recombinant endotheliotropic eGFP-labelled CMV was propagated in RPE-1 cells and purified by ultracentrifugation in tartrate/glycerol gradients. Primary HSEC were isolated from explanted livers, grown to confluence and infected with CMV over 2 h. Infection was confirmed by fluorescence microscopy and plaque assay on fibroblasts. Chemokines and adhesion molecules were quantified by ELISA. Isolated primary lymphocytes and CMV-specific CD4 T cell clones were perfused over HSEC monolayers under constant flow simulating physiological shear stress and adhesion and transmigration recorded using phase contrast microscopy. Trans-well assays were used to study the phenotype of transmigrated cells using flow cytometry.

Results Human sinusoidal endothelial cells were permissive to CMV infection. CMV infection induced secretion of CXCL10 and CCL5 as well as an up-regulation of VCAM-1 and ICAM-1 surface expression. Early CMV infection resulted in a fourfold increase in the adhesion of allogeneic lymphocytes to infected HSEC monolayers compared with mock-infected endothelium. Under flow, transendothelial migration of CMV-reactive CD4 T cell clones was increased through CMV-infected endothelium and could be significantly reduced by the use of anti-CXCL10 antibodies. Transmigrated allogeneic CD4 CD45RO+ T cells and CMV-reactive T cell clones displayed increased expression of the early activation marker CD69 after transendothelial migration through CMV-infected HSEC.

Conclusion CMV infection of HSEC facilitates the up-regulation of cell-adhesion molecules and chemokines resulting in increased adhesion, transmigration and activation of CD4 T cells. This may explain how human CMV infection not only provokes significant hepatitis but also increases hepatic immune activation in graft rejection.

Competing interests None declared.