Figure 3.

Rap1 can be activated with the drug 8CPT-2′OMe-cAMP in ARPE-19, and this increases electrical impedance. (A) Confluent ARPE-19 cells were treated with vehicle (PBS) or with the Rap1-activating drug 8CPT-2′OMe-cAMP before harvesting lysates for determination of Rap1 activity. Active (GTP-bound) Rap1 is shown in top row, and bottom row confirms equivalent loading using total Rap1 from whole cell lysates. (B) ARPE-19 cells were treated with vehicle (PBS) or drug, and electrical impedance was measured using RTCA. Graph shows a typical RTCA trace from one representative experiment. (C) Combined data from 3 independent experiments showing the fold change in Cell Index ± SD, 22 hours after 8CPT-2′OMe-cAMP treatment, normalized to untreated control. Rap1 activation increases Cell Index, indicating enhanced barrier properties (*P = 0.01, n = 3 independent experiments).