Monocyte influx secondary to ischemia-reperfusion conditions the renal allograft to rejection by presentation of antigens and production of cytokines. Monocyte influx depends on NFkappaB-dependent transcription of genes encoding adhesion molecules and chemokines. Here we demonstrate that cationic liposomes containing phosphorothioated oligodeoxynucleotides (ODN) with the kappaB binding site serving as competitive binding decoy, can prevent TNF-alpha-induced NFkappaB activity in endothelial cells in vitro. In an allogenic rat kidney transplantation model (BN to LEW), we show that perfusing the renal allograft with this decoy prior to transplantation abolishes nuclear NFkappaB activity in vivo and inhibits VCAM-1 expression in the donor endothelium (P<0.05). At 24 h postreperfusion, periarterial infiltration of monocytes/macrophages was significantly reduced in decoy ODN-treated allografts compared to control allografts (3.7+/-0.7 vs. 9.2+/-1.2 macrophages/vessel; P<0.01). At 72 h, there was a reduction of tubulointerstitial macrophage infiltration in decoy ODN-treated kidneys compared to controls (75.6+/-13.9 vs. 120.0+/-11.2 macrophages/tubulointerstitial area; P<0.05). In conclusion, perfusion of the renal allograft with NFkappaB decoy ODN prior to transplantation decreases the initial inflammatory response in a stringent, nonimmunosuppressed allogenic transplantation model. Therefore, the NFkappaB decoy approach may be useful to explore the role of endothelium and macrophages in graft rejection and may be developed into a graft-specific immunosuppressive strategy allowing reduction of systemic immunosuppression on organ transplantation.

In these studies, we find that the vascular endothelial growth factor (VEGF) receptor KDR is expressed on subsets of mitogenactivated CD4 ؉ and CD8 ؉ T cells in vitro. We also found that KDR colocalizes with CD3 on mitogen-activated T cells in vitro and on infiltrates within rejecting human allografts in vivo. To evaluate whether VEGF and KDR mediate lymphocyte migration across endothelial cells (ECs), we used an in vitro live-time transmigration model and observed that both anti-VEGF and anti-KDR antibodies inhibit the transmigration of both CD4 ؉ and CD8 ؉ T cells across tumor necrosis factor␣ (TNF␣)-activated, but not unactivated ECs. In addition, we found that interactions among CD4 ؉ or CD8 ؉ T cells and TNF␣-activated ECs result in the induction of KDR on each T cell subset, and that KDRexpressing lymphocytes preferentially transmigrate across TNF␣-activated ECs. Finally, using a humanized severe combined immunodeficient mouse model of lymphocyte trafficking, we found that KDRexpressing lymphocytes migrate into human skin in vivo, and that migration is reduced in mice treated with a blocking anti-VEGF antibody. IntroductionVascular endothelial growth factor (VEGF), an angiogenesis factor, is established to function in the migration, proliferation, and survival of endothelial cells (ECs). 1,2 VEGF is well known to function in wound healing, organ development, and tumor growth and it serves to promote tissue protection and repair after acute injury. 3 VEGF is also expressed in association with cell-mediated immune inflammation and acute and chronic inflammatory reactions. 4 In chronic inflammatory disease processes, VEGF fails to elicit effective tissue repair, and rather may induce a pathologic form of angiogenesis that has been proposed to augment disease activity. 4,5 Indeed, several studies have demonstrated that blockade of VEGF may attenuate the progression of chronic diseases such as arthritis, atherosclerosis, and allograft rejection. [6][7][8] Although relatively underappreciated, VEGF has potent proinflammatory properties including an ability to mediate leukocyte trafficking into sites of cell-mediated immunity. [7][8][9][10][11][12][13] The proinflammatory properties of VEGF are reported to be dependent on its ability to interact directly with monocytes resulting in chemotaxis, 10 its ability to induce the expression of endothelial adhesion molecules 9,11 and chemokine production, 8,12,14 and its ability to enhance vascular permeability. 2 Furthermore, VEGF has been reported to have direct chemoattractant effects on murine and human T cells, 13,15 and blockade of VEGF in vivo has been found to inhibit lymphocyte trafficking into skin and rejecting cardiac allografts. 8,16,17 However, the mechanism underlying the ability of VEGF to interact with T cells is not known, and the molecular basis for its ability to facilitate lymphocyte chemotaxis in vitro or in vivo is poorly understood.Several recent studies have determined that the VEGF receptors Flt-1 (VEGF receptor 1), KDR (VEGF receptor 2) and neu...

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