Antibodies eluted from acutely rejected renal allografts bind to and activate human endothelial cells

This study was designed to investigate how antiendothelial antibodies (EAbs) are involved in acute irreversible renal graft rejection. Eluates from 25 renal allografts, lost by irreversible rejection (n = 22) and by renal vein thrombosis (controls n = 3), were tested against a panel of cultured human umbilical vein endothelial cells (HUVEC). All patients were under immunosuppression at the time of nephrectomy. EAbs binding and membrane expression of adhesion molecules ELAM-1 and VCAM-1 were analyzed by flow cytometry (FACS) and by semiquantitative RT-PCR for mRNAs coding for those molecules. The absence of anti-HLA antibodies against the donor was ascertained at transplant, and before and after nephrectomy by the negativity of specific crossmatches performed using the most sensitive techniques. EAbs eluted from eight rejected kidneys bound to HUVEC. They did not induce any cytotoxicity, but their incubation with HUVEC (4 h at 37 degrees C; 2.5 mg/ml) led to upregulation of mRNAs coding for VCAM-1 (35- to 60-fold increases) and ICAM-1 (8- to 12-fold increases) as compared with control EAbs. Membrane expression of adhesion molecules was also strikingly increased, with 80% of the cells expressing VCAM-1 and 65% expressing ELAM-1 upon incubation. EAbs were detected in eight out of nine (88.8%) eluates from kidneys lost from acute vascular rejection, but in none of the 13 (0.0%) kidneys lost from other types of rejection (p < 0.0001). We conclude that EAbs, capable of activating human endothelial cells, can be recovered from acutely rejected kidneys and may play a direct role in the pathogenesis of acute rejection.