Fig. 1

Experimental design & ultrasound apparatus. A Experimental design of ICH animal model: Exp.1) Expressions of Iba1 (microglia) and GFAP (astrocyte) were measured at 3 h, 6 h, 1 day, and 3 days after ICH; Exp.2) LIPUS treatment (red dash) was performed daily from D0 to D2 in the brain, and mice were sacrificed on D3; Exp.3) 2-h-delayed LIPUS treatment post-ICH. B Schematic diagram of LIPUS for ICH animal model. The ultrasound transducer was coupled with a metal collimator to target on the ICH lesion. C Experimental design of cell cultures: Exp.1) The study involved the following cell types: BV2 (microglia), TNA2 (astrocyte), and N2A (neuron). BV2 cells were stimulated with thrombin (10 U/mL), TNA2 cells were stimulated with either thrombin (10 U/mL) or microglia-conditioned media (MCM), and N2A cells were stimulated with either MCM or astrocyte-conditioned media (ACM). After the induction, cells received LIPUS treatment, and cell viability was measured at 24 h or 48 h. Additionally, conditioned media were collected for 24 h following the induction. Exp.2) Primary cell cultures were subjected to thrombin stimulation (10 U/mL) and either treated or not treated with LIPUS. The cells were then harvested at 1 h, 3 h, or 24 h following the stimulation. Exp.3) The LY294002 (PI3K inhibitor) was administered 2 h prior to inducing thrombin stimulation (10 U/mL) with or without LIPUS treatment. The cells were then harvested after 24 h. D Schematic diagram of LIPUS treatment of cell cultures. Acoustic wave was sonicated from the bottom of culture plate to stimulate cells. (IF immunofluorescence staining, WB western blots, CV cresyl violet staining, mNSS modified neurological severity score, BWC brain water content, NO nitrite oxide, MTT 3-(4,5-dimethyl-2-thiazolyl)- 2,5-diphenyl-2H-tetrazolium bromide assay)