Inflammation does not play a role in BBB leakage after tPA overexpression. All experiments were performed 48 hours after hydrodynamic transfection of pLIVE-tPA or pLIVE-0 plasmids. (A) Immunofluorescence images of pLIVE-0- and pLIVE-tPA-transfected animals, showing that microglial cells (expressing ionized calcium binding adaptor molecule 1 [Iba-1], red) remain ramified (ie, not activated) in mice presenting a hyperfibrinolytic state. (B) Quantification of Iba-1–positive cells (n = 5 per group). (C) Interleukin 1-β expression levels (mRNA) in brain lysates in pLIVE-0- and tPA-transfected mice (n = 5 per group). (D) Tumor necrosis factor expression levels in brain lysates in pLIVE-0- and tPA-transfected mice (n = 5 per group). (E) Fluorescence images of pLIVE 0- and tPA-transfected animals showing that brain VCAM-1 expression (green) levels are not modified after plasmatic tPA overexpression. (F) Quantification of VCAM-1–positive endothelial cells in E (n = 5 per group). (G) VCAM-1 mRNA expression levels in brain lysates in pLIVE-0- and pLIVE-tPA-transfected mice (n = 5 per group). (H) ICAM-1 mRNA expression levels in brain lysates in pLIVE-0- and tPA-transfected mice (n = 5 per group). (I) Experimental design performed to study VCAM-1 expression using ultrasensitive molecular magnetic resonance imaging technique. Imaging was performed 20 minutes after iron oxide microparticles-α–VCAM-1 injection. (J) Representative magnetic resonance image after iron oxide microparticles–VCAM-1 administration in control and tPA-transfected mice. (K) Mean signal void iron oxide microparticles–VCAM-1 levels of J (n = 5 per group). (L) Using Fluoro-Jade C staining (green), no neuronal cell-death was detected 48 hours after pLIVE-tPA injection (left) compared with a positive control (transient bilateral occlusion of the common carotid arteries model; right), arguing against the implication of neuronal damage in tPA-induced BBB leakage. 4′,6-diamidino-2-phenylindole (DAPI; blue) was used as a nuclear marker, collagen IV (Coll IV; red) as an endothelial marker, and NeuN (green) as a neuronal marker. FJC, fluoro-jade C; Iba+, ; IL, interleukin; MPIO, iron oxide microparticles; MRI, magnetic resonance imaging; ns, nonsignificant differences.