FK778 attenuates lymphocyte-endothelium interaction after cardiac transplantation: in vivo and in vitro studies.
BACKGROUND
The malononitrilamide FK778 is a novel derivate of leflunomide and interacts with T- and B-cell function by inhibiting de novo pyrimidine synthesis. We investigated the effects of FK778 upon acute cardiac allograft rejection and upon adhesion molecule upregulation in experimental transplantation and by using in vitro cell culture.


METHODS
Heterotopic, abdominal cardiac transplantations were performed in the Brown Norway (BN) to Lewis (Lew) rat model. The study groups received daily low- or high-dose FK778 immunosuppression. FK778 plasma levels were quantified by HPLC. Grafts were harvested on the fifth postoperative day for histologic and immunohistologic examinations using computerized morphometry. Purified BN aortic endothelial cell cultures were pretreated with low- or high-dose FK778 according to FK778 plasma levels and were stimulated with tumor necrosis factor (TNF)-alpha. Adhesion molecule expression was quantified by immunofluorescence, FACS analysis, and Western blotting. Lymphocyte-endothelium adhesion assays were performed using purified Lew lymphocytes and radiolabeled TNF-alpha was used for receptor binding assays.


RESULTS
FK778 treatment dose-dependently reduced graft mononuclear infiltration of CD4(+), CD8(+), and ED1(+) cells, but only high-dose FK778 treatment significantly reduced early upregulation of ICAM-1 and VCAM-1 in vivo. FK778 also dose-dependently reduced TNF-alpha-stimulated endothelial adhesion molecule upregulation in vitro, whereas the effect on VCAM-1 was more dominant. We did not find evidence that FK778 interferes with surface receptor binding of TNF-alpha. Lymphocyte adhesion to endothelial cell monolayers was significantly attenuated by FK778.


CONCLUSIONS
Besides its inhibitory effect on pyrimidine synthesis, FK778 directly reduces endothelial adhesion molecule upregulation and attenuates lymphocyte-endothelium interaction, which is a critical step in graft rejection.
Deuse
Tobias
T
Department of Cardiovascular Surgery, University Hospital Hamburg-Eppendorf, Hamburg, Germany.
Schrepfer
Sonja
S
Schäfer
Hansjörg
H
Koch-Nolte
Friedrich
F
Schwedhelm
Edzard
E
Böger
Rainer H
RH
Reichenspurner
Hermann
H
D000208:Acute Disease; D000480:Alkynes; D000818:Animals; D000970:Antineoplastic Agents / Q000493:pharmacokinetics; D002448:Cell Adhesion / Q000187:drug effects* / Q000276:immunology; D042783:Endothelial Cells / Q000166:cytology* / Q000378:metabolism; D006084:Graft Rejection / Q000188:drug therapy* / Q000276:immunology / Q000378:metabolism; D016027:Heart Transplantation; D066298:In Vitro Techniques; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism; D007457:Iodine Radioisotopes; D007555:Isoxazoles / Q000097:blood / Q000494:pharmacology*; D008214:Lymphocytes / Q000166:cytology* / Q000378:metabolism; D009570:Nitriles; D051381:Rats; D011914:Rats, Inbred BN; D011917:Rats, Inbred Lew; D018124:Receptors, Tumor Necrosis Factor / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000493:pharmacokinetics; D019010:Vascular Cell Adhesion Molecule-1 / Q000378:metabolism
D000480:Alkynes; D000970:Antineoplastic Agents; D007457:Iodine Radioisotopes; D007555:Isoxazoles; D009570:Nitriles; D018124:Receptors, Tumor Necrosis Factor; D014409:Tumor Necrosis Factor-alpha; D019010:Vascular Cell Adhesion Molecule-1; D018799:Intercellular Adhesion Molecule-1; C115284:2-cyano-3-hydroxy-N-(4-(trifluoromethyl)phenyl)-2-hepten-6-ynamide