E2 treatment impacts on the platelet proteome. (A) Platelets from 3 or 4 mice treated or not with E2 were pooled, and proteins were labeled with CyDye DIGE Fluor Minimal Dyes as described in supplemental Methods. Results from mass spectrometry analysis are summarized in a table showing accession numbers, names, Mascot score, sequence coverage, and number of peptides for each of the 5 excised spots. The experiment was repeated 3 times. Spots displaying a ≥ 1.5 increase or decrease in abundance with a P value < .05 were selected for protein identification. (B) Level of β1 tubulin in platelets from ovariectomized mice treated or not with E2 (n = 7 and n = 8, respectively) quantified by a specific ELISA test. Results are mean values ± SEM. **Significant difference (P < .01). (C) Average ratios (−E2/+E2) of β1 tubulin expression in megakaryocytes from mice ERα^+/+ or ERα^−/−. Progenitor cells isolated from bone marrow of mice treated with E2 (n = 4 in each group) or not (n = 3 in each group) were incubated or not with the anti-estrogen ICI 182780 (10⁻⁶M) along the differentiation process. Four or 5 days after addition of thrombopoietin (100 ng/mL), Megakaryocytes (MKs) were lysed and their β1 tubulin content was analyzed using the ELISA test. *Significant difference (P < .1) between mice treated or not with E2 in each group (ERα^+/+ or ERα^−/−). (D) Representative transmission electron microscopy images of a resting platelet from E2-treated mice showing the microtubule coils in the marginal band (scale bar: 0.1 μm).