IL-1α is released from activated platelets and potently induces brain endothelial CXCL1 release and ICAM-1 and VCAM-1 expression. (A) The effects of CM from resting (vehicle/ethanol-treated) or PMA-activated platelets on the release of CXCL1 from MBECs. The effects of IL-1RA (B) or coapplication of neutralizing antibodies to IL-1α or IL-1β on activated platelet CM-induced CXCL1 release from MBECs (C). Rat brain endothelial cell line GPNT (D), or primary cultures of MBECs (E-G) were treated with vehicle, IL-1α, or IL-1β (0-100 U/mL). Twenty-four hours later (D-F), the CM was assayed for CXCL1 (D-E) or the cells lysed and assayed for ICAM-1 and VCAM-1 (F) by ELISA. (G) Four hours after treatment, MBECs were immunostained for CXCL1 and PECAM-1 (scale bar represents 20 μm). Images were acquired on a Delta Vision RT restoration microscope using a 40×/1.3 Plan Apo objective and the Sedat filter set (G). The images were collected using a CoolSNAP HQ camera with a Z optical spacing of 0.2 μm, raw images were deconvolved using softWoRx software, and maximum intensity projections of these deconvolved images are shown (G). A, from left to right: n = 4, 3, 3, 3, 3, 3; B, n = 4, 3; C, n = 5, 5, 6, 6; D, n = 2; E, n = 5, 3, 3; F, ICAM-1 n = 5, 5, 5, 6; VCAM-1, n = 3, 3, 3, 4. Data are mean ± SEM. ***P < .001. **P < .01. *P < .05.