BrdU assay using rat aortic smooth muscle cells. (A) After rat aortic smooth muscle cells were plated, the cells were incubated in DMEM containing 0.1% FBS with or without 10 nM linagliptin for 48 h. Subsequently, cells were stimulated with PDGF (25 ng/ml) for 24 h. (B) Cells were incubated in DMEM with 0.1% FBS for 24 h. Subsequently, they were stimulated with DPP-4 (50 ng/ml) with or without 10 nM linagliptin for 24 h. BrdU solution was added during the last 2 h, and cells were harvested for measurement of DNA synthesis using a microplate reader at 450-620 nm. Mean data are expressed as the ratio of the control cell proliferation. Unpaired t-tests were performed to calculate statistical significance. Data are mean ± SEM. **p <0.01 vs. control PDGF(+), ##p <0.01 vs. control PDGF(+), *p <0.05 vs. control.