PLPs stimulate proliferation of and are internalized by HepG2 cells. (A) Activated platelets or PLPs were added to HepG2 cells and incubated under serum-free conditions. After 48 hours, cell proliferation rate was estimated by quantifying BrdU incorporation. *P < .05. Data represent the mean of 3 independent triplicate experiments. Error bars indicate SD. (B) PLPs (green) were labeled with CMFDA and were incubated under serum-free conditions for 4 hours with HepG2 cells. Original magnification ×400. Scale bar, 10 µm. (C) Quantification of EU incorporation into PLP RNA by flow cytometry. PLPs were generated from MEG01 cells that had been exposed to EU. EU in PLPs was visualized using Click-iT RNA Alexa Fluor 488. Histogram plot overlay shows control PLPs (red line) and PLPs generated from EU-treated MEG01 cells (blue line). (D) EU-labeled PLPs were incubated under serum-free conditions for 2, 4, or 8 hours with HepG2 cells. Dashed lines represent the HepG2 cell membrane. Structures resembling PLPs (arrows) as well as parts of the cytoplasm of the HepG2 cell are EU positive (green, arrowheads). Original magnification ×400. Scale bar, 10 µm.