Cell Reports: Methods (Feb 2023)
A versatile vessel casting method for fine mapping of vascular networks using a hydrogel-based lipophilic dye solution
Abstract
Summary: Efficient labeling of the vasculature is important for understanding the organization of vascular networks. Here, we propose VALID, a vessel-labeling method that enables visualization of vascular networks with tissue clearing and light-sheet microscopy. VALID transforms traditional lipophilic dye solution into hydrogel by introducing gelatin and restrains the dye aggregation, resulting in complete and uniform vessel-labeling patterns with high signal-to-background ratios. VALID also enhances the compatibility of lipophilic dyes with solvent-based tissue-clearing protocols, which was hard to achieve previously. Using VALID, we combined lipophilic dyes with solvent-based tissue-clearing techniques to perform 3D reconstructions of vasculature within mouse brain and spinal cord. We also employed VALID for 3D visualization and quantification of microvascular damage in a middle cerebral artery occlusion mouse model. VALID should provide a simple, cost-effective vessel-labeling protocol that would significantly widen the applications of lipophilic dyes in research on cerebrovascular complications. Motivation: 3D reconstruction of the entire vasculature is fundamental to understanding vascular lesions during the progression of various neurovascular diseases. The combination of vessel-labeling, tissue-clearing, and optical imaging techniques allow 3D mapping of the vasculature of different tissues/organs. Current vessel-specific marker-based labeling methods can provide biological ground truth for blood vessels but are restricted by either insufficient signal-to-background ratios, incomplete labeling for large vessels, or high cost. Perfusion-based methods can serve as useful alternatives with relative high signal-to-background ratios; however, dye leakage has greatly limited their applications. Additionally, the tissue-clearing compatibility of these methods still needs to be improved. Here, we developed VALID, an efficient, easy-handling, and cost-effective protocol that enables complete and uniform labeling of different vasculatures with bright fluorescence.
