Loss of col4a1 in zebrafish recapitulates the cerebrovascular phenotypes associated with monogenic cerebral small vessel disease

Abstract

Cerebral small vessel disease (cSVD) is a major cause of vascular dementia and stroke. Our understanding of cSVD pathophysiology is incomplete and our ability to treat patients is limited. Pathogenic variants in type IV collagen alpha 1 (COL4A1) cause a monogenic form of cSVD with variable age of onset, via disturbance of cerebrovascular basement membranes. Zebrafish larvae are a powerful model organism for studying cerebrovascular disease due to their optical clarity and applicability for live imaging. In this study, we characterised a zebrafish crispant model for loss-of-function COL4A1-associated cSVD that successfully recapitulates key disease features, including spontaneous intracerebral haemorrhage and cerebrovascular abnormalities. We also identified evidence for abnormal cerebrovascular basement membranes and elevated matrix metalloproteinase 9 (mmp9) transcription associated with loss of col4a1. Pharmacological inhibition of mmp9 was sufficient to ameliorate some cerebrovascular phenotypes. Finally, we describe the generation of a mutant line carrying a germline-transmissible 20 bp deletion in zebrafish col4a1 (col4a1^Δ20) which is associated with cerebrovascular abnormalities, swimming defects and increased susceptibility to pharmacologically induced brain haemorrhages during larval stages. In adulthood, mutant col4a1^Δ20 animals developed spontaneous brain haemorrhages that were observable in free-swimming fish. Overall, this study validates the use of zebrafish disease modelling for preclinical COL4A1-associated cSVD research and highlights its potential for further understanding disease pathophysiology and future drug discovery projects.

Keywords: Cerebral small vessel disease; Cerebrovasculature; Col4a1; Intracerebral haemorrhage; MMP9; Type IV collagen; Zebrafish.