Silencing the expression of platelet endothelial cell adhesion molecule-1 prevents allogeneic T-cell cytotoxicity.
BACKGROUND
After solid organ transplantation, the endothelium is the first allorecognition checkpoint of the recipient's immune system. A major player at this interface is the platelet endothelial cell adhesion molecule-1 (PECAM-1), which is an immunoglobulin-like glycoprotein, involved in white blood cell migration, cellular adhesion, and signal transduction.


METHODS
The potential of preventing allorecognition at this interface was explored by knocking down PECAM-1 expression using RNA interference. Lentiviral-based vectors encoding short-hairpin RNA sequences specific for PECAM-1 transcripts were used for the transduction of monocytic and endothelial cell lines.


RESULTS
Expression of PECAM-1 decreased by up to 80% at mRNA and protein levels on monocytic and endothelial cell lines. Antigen-binding capacity assays likewise showed up to 80% reduction of PECAM-1 expression on cell surfaces. In allogeneic T-cell stimulation assays, T cells stimulated with PECAM-1-silenced monocytes had granzyme B mRNA levels up to 85% lower than those in T cells stimulated with monocytes expressing nonspecific shRNA. Also, T-cell cytotoxicity showed to decrease significantly against PECAM-1-silenced monocytes versus those nonsilenced for PECAM-1. Moreover, the former T cells did not secrete interferon-γ and exhibited reduced proliferation.


CONCLUSIONS
These findings suggest that permanent silencing of PECAM-1 expression is feasible and contributes to efficiently inhibiting the cytotoxic T-cell response to allogeneic target cells. Thus, targeting both the immunologic synapse at the regulatory interface and the lymphocyte migration might provide a new strategy to overcome allogeneic transplant rejection.
Jaimes
Yarúa
Y
Institute for Transfusion Medicine, Hannover Medical School, Hannover, Germany.
Figueiredo
Constança
C
Blasczyk
Rainer
R
D015153:Blotting, Western; D018821:CD18 Antigens / Q000378:metabolism; D002460:Cell Line; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D053804:Granzymes / Q000235:genetics; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism; D016086:Lentivirus / Q000235:genetics; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000235:genetics / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D013601:T-Lymphocytes / Q000166:cytology* / Q000502:physiology*
D018821:CD18 Antigens; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D007371:Interferon-gamma; D053804:Granzymes