Snail deficiency blocks nicotine-induced endothelial to mesenchymal transition (EndMT). (A) The protein levels of Snail in human aortic endothelial cells (HAECs) were assessed by western blot. n = 4-5. (B) The mRNA levels of Snail were determined by RT-PCR. n = 5. (C-D) Effects of α-BTX and PD98059 on Snail levels in nicotine-treated HAECs. +α-BTX indicates the co-application of α-BTX with nicotine. +PD98059 indicates the co-application of PD98059 with nicotine. n = 3-5. (E) Verification of silencing efficiency of Snail by siRNA in nicotine-treated HAECs. Ctl indicates negative control. n = 5. (F) The protein levels of Snail, CD31, VE-cadherin, α-SMA and FSP1 in nicotine-treated HAECs. n = 5. (G) Immunofluorescence staining for EndMT-related markers and F-actin staining in nicotine-treated HAECs. Scale bar indicates 50 µm. (H) Morphological changes of nicotine-treated HAECs after silencing Snail. Scale bar indicates 200 µm. (I) mRNA levels of different EndMT-related markers and stem cell markers in nicotine-treated HAECs after silencing Snail. n = 5. *P < 0.05, **P < 0.01, ***P < 0.001.