Endothelial cells as targets for antigen-specific cytotoxic T lymphocytes

BACKGROUND: In this study we have examined human vascular endothelial cells as targets
for antigen-specific cytotoxic T lymphocytes (CTL). Vascular endothelial cells (EC) act as the
major interface between blood and tissues and are crucial for the maintenance of vascular
integrity. It has been shown previously that EC are poor targets for immunodominant antigenspecific
CTL. This feature was due to the impaired capacity of EC to present the cognate
antigenic peptides to CTL. These findings fueled the hypothesis that EC present a
substantially different repertoire of MHC class I ligands compared with syngeneic leukocyte
derived cells and that they might be protected from CTL-mediated lysis by this mechanism. It
was the object of this thesis to confirm or rule out this hypothesis.
METHODS: Our study consisted of three parts. First, the peptide repertoire from endothelial
cells was compared with syngeneic leukocyte-derived cells. Cell type-specific HLA-A*02
restricted peptides were identified and characterized for several biochemical features like
expression of source proteins, binding properties and decay that would explain preferential
presentation by either one of the two cells. Next, a method for the induction of self-reactive
CTL was established by using peptide-pulsed dendritic cells as antigen-presenting cells and
the generation of CTL specific for the HLA-A*02 restricted endothelial self-peptides PTRF(56-
64) and CD59(106-114) was attempted. In the third part, we modified the endothelial peptide
repertoire and interfered with the peptide presentation pathway to test the hypothesis that
abundant endothelial peptides indeed compete with immunodominant antigens for
presentation on the surface of EC. The surface antigenic profile was altered by two
approaches: a) by transfection of influenza virus A matrix protein 1 gene to make EC
recognizable for Flu(58-66)-specific CTL and b) by siRNA knockdown of the abundant ECspecific
peptides. RESULTS: We show for the first time that EC present a quantitatively different peptide
repertoire with abundance of certain peptides, compared with leukocytes. The abundance of
endothelial peptides is mainly caused by the preferential expression of the source proteins.
This feature is immunologically interesting since it contributes twofold protection of EC from
CTL-mediated lysis: a) by an extraordinary strong tolerance that exists against EC peptides
and b) by competing with immunodominant peptides for the MHC class I binding site. Under
conditions that were sufficient to generate CTL specific for certain immunodominant low
abundant self-peptides induction of CTL against PTRF(56-64) and CD59(106-114) failed.
Improvement of culture conditions by stabilization of the pMHC I complex or interference with
inhibitory pathways affected again only the induction of CTL against low abundant selfpeptides
but not against EC-specific peptides indicating that these peptides are particularly
tolerogenic. After modification of the endothelial peptide repertoire and knockdown of
abundant EC peptides, EC became better targets for immunodominant CTL by increased presentation of endogenously processed peptides indicating that the selective presentation
of PTRF(56-64) and CD59(106-114) can protect EC by hiding them from CTL-mediated lysis.
CONCLUSION: Our in vitro findings are in accordance with the view that EC are protected
from CTL-mediated lysis by presentation of a quantitatively different peptide repertoire. This
protection results from peptides that compete with immunodominant peptides for the MHC
class I binding site and, in addition, evoke extraordinary strong tolerance.