FXII stimulates ERK1/2 or Akt phosphorylation on HUVECs cultured on gelatin. (A) Immunoblots that used an antibody to phospho-ERK1/2 (pERK1/2) and antibody to total ERK1/2 (ERK1/2). (B) Immunoblots that used an antibody to phospho-Akt [pAkt (S473)] and total Akt (Akt). (A) ERK1/2 phosphorylation: lane 1, the control (cells) is untreated serum-starved cells. In lanes 2 and 3, 62nM FXII-treated cells in the absence or presence of 50μM Zn²⁺ are shown. The effect of PD98059, Wortmannin, LY294002, 0.6μM Mab 3B10, or 300μM peptide YHK9 on 62nM FXII plus 50μM Zn²⁺-treated cells is shown in lanes 4-8, respectively. (B) Akt phosphorylation: lane 1 is serum-starved cells (cells). In lanes 2 and 3, 10% serum-treated cells (serum) alone with Wortmannin. In lanes 4-7, 62nM FXII plus 50μM Zn²⁺-treated cells in the absence or presence of Wortmannin, LY294002, or PD98059, respectively, is shown. (A-B) The figures are representative immunoblots of 3 or more experiments. (C) Bar graphs of the means ± SD of scans of 3 or more immunoblots shown in panels A and B for both pERK1/2 and pAkt. (C) An asterisk over a bar graph indicates P ≤ .05 compared with FXII-stimulated HUVECs alone. (D) A single immunoblot is shown with SIN1^−/− MEFs treated with FXII plus 50μM Zn²⁺ in the absence or presence of PD98059, Wortmannin, or 50μM U1026 immunoblotted for pERK1/2, total ERK, pAkt, or total Akt. (E) Bar graphs of the means ± SD of scans of 4 immunoblots shown in panel D for pERK1/2. (E) An asterisk over a bar graph indicates P ≤ .05 compared with FXII-stimulated HUVECs alone.