Annexin A5 inhibits platelet uptake and platelet-mediated hepatocyte proliferation. (A) Activated platelets were added to HepG2 cells in presence of Annexin A5 (AV), OSE, or asialofetuin (Asf). Internalized platelets were quantified based on fluorescence microscopy. Platelets were manually counted in at least 5 high-power fields and expressed as number of platelets/100 cells. ***P < .001. Data represent the mean of 3 independent triplicate experiments. Error bars indicate SD. (B) Activated platelets were added to HepG2 cells in the presence or absence of various inhibitors and incubated under serum-free conditions. After 48 hours, the cell proliferation rate was estimated by quantifying BrdU incorporation. Control groups represent HepG2 cells cultured in the presence or absence of FCS. In addition, the effect of Annexin A5 on BrdU incorporation in the absence of platelets, but in the presence of FCS, is shown. *P < .05 compared with −FCS. Data represent the mean of 3 independent triplicate experiments. Error bars indicate SD. (C) Representative fluorescence microscopy image from activated platelets incubated with HepG2 cells in the absence (i) or presence (ii) of Annexin A5 for 2 hours. HepG2 cells were stained for actin (red). Platelets were labeled with CMFDA (green). Original magnification ×200. Scale bar, 20 µm.