Influence of antisense oligonucleotides to PRCP to block PK activation or antigen on HUVECs. (A) Eighty percent to 90% confluent monolayers of HUVECs in microtiter plates were incubated with various oligonucleotides for 4 hours, as described in “Materials and methods.” After incubation and washing, the cells were incubated with standard culture media for 16 to 20 hours before assay for formed plasma kallikrein (“Materials and methods”). Results presented are the mean ± SEM of 5 separate experiments. HK + PK lane: Nontreated cells incubated with these proteins. Sense, inverted, and antisense lanes: Wells incubated with modified sense, inverted, or antisense morpholino oligonucleotides, respectively. (B) The figure is a representative laser scanning confocal micrograph of untreated HUVECs (control) or antisense treated (Antisense) using a goat anti-PRCP antibody.9 Original magnification × 60. (C) The influence of antisense oligonucleotides to PRCP on PK antigen expression on HUVECs. HUVECs were grown on microscope slides and prepared for morpholino treatment, as described in “Materials and methods.” Data presented for each condition are the mean ± SEM of 32 to 34 fields of view with an average of 14 to 15 cells/field from 5 independent experiments. Results are expressed as arbitrary fluorescent units per cell. Control lane: untreated cells. Sense, inverted, and antisense lanes: HUVECs incubated with modified sense, inverted, or antisense morpholino oligonucleotides.
