Here we applied a 3D analytical method with the CUBIC tissue clearing technique (Cell Rep 2018) for visualizing the structure of vessel networks in the brains of patients with SVD.

We evaluated the post-mortem brains of two patients with CADASIL (a woman aged 69 years and a man aged 62 years), two patients with heterozygous HTRA1 mutations (heterozygous CARASIL: men aged 66 and 61 years), and two controls (a woman aged 34 years and a man aged 89 years). For 3D analysis, formalin-fixed, 1-cm³ tissue blocks of the inferior frontal gyrus were prepared. The blocks were highly cleared using the CUBIC protocol and subjected to immunohistochemistry with an anti-smooth muscle actin (SMA) antibody. Then, 3D images of SMA-positive structures were captured using light-sheet fluorescence microscopy.