Figure 3. MVs from senescent HUVEC cells induced calcification in HASMC. Cells were cultured with 50,000 MVs/mL from young (Y MV) or senescent (S MV) HUVEC for different times. (A) Qualitative calcification was stained with alizarin red. A representative experiment from six different experiments per duplicate is show. (B) Calcium content was determined by spectrophotometer by phenolsulphonephthalein dye at 6 and 9 days of treatment. The graphs present calcium content of the cells expressed as µg/mg protein. The data represent means ± SD of 3 independent experiments. *** p<0.001 vs control and Y MV at the same time.