(A) The interaction between PECAM-1 isoforms and eNOS was assessed by immunoprecipitation (IP) analysis of lysates prepared from PECAM-1 −/− bEND cells expressing a specific isoform of PECAM-1under normal cell culture condition. The amount of total protein for lysates was 1/20 of that applied for IP analysis. (B) Intracellular NO levels in PECAM-1 −/− bEND cells expressing a specific isoform of PECAM-1 was measured using the DAF-FM assay as previously described [185]. Please note that all the PECAM-1 isoforms interacted with PECAM-1. The isoforms lacking exona 14, 15 or both showed higher levels of eNOS compared to full length PECAM-1 or wild type cells. However, expression of none of these isoforms resulted in activation of eNOS and NO production as seen in wild type cells. Thus, simple expression of PECAM-1 and its interaction with eNOS is not sufficient to restore NO levels in null cells. (S.Park, CM Sorenson and N. Sheibani, unpublished work)