Distinct dynamics and kinetics determine efficient antigen-presentation by LSEC and support IL-2 dependent CD8 T cell activation

Abstract

Liver sinusoidal endothelial cells (LSEC) have important clearance and immune regulatory functions, acting as unique organ resident APC. Soluble antigens are rapidly taken up by LSEC and can be cross-presented within 1h to CD8 T cell. We investigate the mechanism of efficient antigen uptake and cross-presentation by LSEC which are distinct from DC. We found LSEC to show more pronounced antigen uptake and cross-presentation than CD8a⁺ DC on a per cell basis <i>ex vivo</i> and <i>in vitro</i>. However, whereas DC cross-presented antigen for a prolonged period of time, antigen taken up by LSEC in vivo was rapidly cleared (t_1/2= 5h) and cross-presentation declined accordingly within 24h. <br /> It has recently been shown in DC, that cross-presentation of ovalbumin (OVA) is exclusively dependent on uptake by the mannose receptor (MR) which routes antigen into stable early endosomes (EEA1⁺), while antigen taken up via scavenger receptors (SR) was routed into lysosomal compartments and was not cross-presented. In contrast, MR-deficient LSEC did not show a significant reduction in cross-presentation. Furthermore there was no spatial separation of antigen taken up via the MR or SR receptor, which colocalized in transiently EEA1⁺ endosomal sorting compartments in LSEC. Receptor-mediated endocytosis did not always lead to cross-presentation, because immune-complexed antigen taken up via Fc-receptors was badly cross-presented by LSEC indicating that induction of CD8 T cell tolerance by LSEC was impaired in the presence of preexisting immunity. Our results provide a mechanistic explanation how organ-resident LSEC accommodate continuous scavenger function as well as cross-presentation of circulating antigens, utilizing distinct kinetics and dynamics for antigen-uptake, routing and cross-presentation compared to DC. <br /> The functional outcome of cross-presentation of exogenous soluble antigen by LSEC on MHC-I-molecules to naïve CD8 T cells is the induction of T cell tolerance which depends on mutual up-regulation of co-inhibitory B7-H1 on LSEC and PD1 on CD8 T cells during antigen-specific interaction. <br /> We further addressed the question whether the highly efficient cross-presentation by LSEC had an influence on tolerance induction over a range of antigen-concentrations. The concentration of OVA used to pulse LSEC directly correlated with the expression-levels of peptide-loaded MHC-I-molecules and thus strength of initial T cell-receptor stimulation. Surprisingly, LSEC induced tolerance only at low but not at high antigen-concentrations. T cell-differentiation into effector T lymphocytes (CTL) was caused by early release of IL-2 by naïve CD8 T cells. IL-2 expression and subsequent CTL-differentiation was influenced by B7-H1 mediated signals from cross-presenting LSEC and strength of T cell receptor (TCR) avidity. Dynamic expression of B7-H1 on LSEC correlated with the strength of TCR-signaling at low but not at high antigen-concentrations, indicating that the balance between TCR- and co-inhibitory signals controlled IL-2 expression. Exogenously added IL-2 abrogated LSEC mediated tolerance induction resulting in full CTL-differentiation. Our results indicate that T cell tolerance mediated by LSEC can be broken in situations where T cells with high-avidity TCR encounter LSEC cross-presenting high numbers of cognate MHC-I-peptide molecules, such as during viral infection of the liver. Furthermore, IL-2 produced by T cells during priming has a strong co-stimulatory function which might promote local induction of immunity in the liver.