Fig. 3.

Expression of MCPIP induces upregulation of EC phenotypic markers and uptake of acLDL in BMNCs. BMNCs were transfected with Flag-tagged MCPIP expression vector or Flag-tagged empty vector and cultured in EC cultured medium. At day 7 after transfection, expression of monocytic and endothelial cell phenotypic markers was evaluated by RT-PCR (A). Relative mRNA expression of these markers is represented in bar graphics (B and C). *P < 0.05 versus cells transfected with control vector. (D) Representative photomicrographs of indirect immunofluorescence using fluorescein isothiocyanate–labeled secondary antibody (green) for EC surface markers CD31 and CD144. Nuclei were counterstained with DAPI in blue. Numbers of cells expressing CD31 and CD144 are presented in a bar graph (E). *P < 0.05 versus cells transfected with control vector. (F) Representative photomicrographs of cells uptaking DiI-acLDL under a phase-contrast/fluorescent microscopy. The red fluorescence indicates intracellular DiI-acLDL. Numbers of cells uptaking DiI-acLDL are presented in a bar graph (G). *P < 0.05 versus cells transfected with control vector. HUVECs and DiI-acLDL-labeled BMNCs were cocultured in EBM medium on 24-well plate coated with Matrigel. (H) Representative photomicrographs of DIC images and DiI-acLDL-loaded images at 24 hours after coculture. The red fluorescence indicates intracellular acLDL. Arrows indicate DiI-acLDL-labeled cells incorporating into tube-like structure (original magnification, ×400). (I) Cells incorporating into tube-like structure were calculated and averaged by five randomly selected high power fields. *P < 0.01 versus cells transfected with control vector.