Quantification of Ventricle and Lesion Volumes

On day 3 after ICH, five mice in each group were anesthetized and perfused intracardially with 4% paraformaldehyde in 0.1 mol/L phosphate-buffered saline (pH 7.4). The brains were removed, kept in 4% paraformaldehyde for 24 h, and then immersed in 30% sucrose for 72 h at 4°C. The entire brain of each mouse was cut into 50-μm-thick sections with a cryostat. All sections except for those from the IVH group were stained with Luxol fast blue (for myelin) and Cresyl violet (for Nissl bodies). Sections from the IVH group were stained only with Cresyl violet. SigmaScan Pro software (version 5.0.0 for Windows; Systat, Port Richmond, CA) was used to quantify volumes of the ventricle and lesion. The volume of the ventricles in cubic millimeters was calculated according to a published method [26] (link). The volume of the lesion in cubic millimeters was calculated by multiplying the thickness by the sum of the damaged areas of each section [14] (link). Sections were analyzed by an investigator blinded to the experimental cohort.