Fig. 5
Effect of anti-ICAM-1 mAbs on apical-to-basolaterally directed PMN transepithelial migration
Transmigration assays were performed in the apical-to-basolateral direction on both untreated (A) and IFNγ-pretreated (1000 U/ml, 48 hr; B) T84 monolayers cultured on 0.33-cm2 permeable supports. As described in Materials and Methods, saturating concentrations of the antibodies (10 µg/ml) indicated were added to the apical side of confluent T84 monolayers and preincubated for 20 min (20°C) followed by addition of 1 × 106 PMN. The monolayers were transferred to 24-well tissue culture wells, each containing 1 ml of 1 µM fMLP in HBSS to initiate PMN transmigration which was allowed to proceed for 110 min (37°C). PMN which had transmigrated to the opposite reservoir or PMN within monolayers were quantitated by myeloperoxidase assay as described in Materials and Methods. Transmigration is normalized with respect to a control binding antibody, anti-MHC I W6/32 and expressed as a percentage of control. As can be seen in Panel A, none of the anti-ICAM-1 antibodies had any effect on transmigration in the control unstimulated monolayers. However, as shown in Panel B, following IFNγ stimulation mAbs R6.5, CBRIC1/7, and CBRIC1/11 significantly inhibited (>60%) transmigration. In data not shown, anti-CD11b (44a) but not anti-CD11a (TS1/22) markedly inhibited transmigration in such assays as previously reported (16,18). Data represents the mean ± SD of four monolayers for each condition; one of three experiments.