Arteriosclerosis, Thrombosis, and Vascular Biology | ANGPT2 Gene Expression Associated with Enlarged Schlemm's Canal Area

This study successfully established an Angpt2-miR-145 mouse model carrying the rs76020419 (G>T) mutation, confirming this variant elevates ANGPT2 expression while correlating with expanded Schlemm's canal dimensions. The model provides a critical platform for investigating ANGPT2's functional roles in ocular hypertension and retinal neurodegeneration.

 

Literature Overview
This article, 'A Glaucoma-Protective Human SNP in the Angpt2 Locus', published in Arteriosclerosis, Thrombosis, and Vascular Biology, reviews functional validation studies of rs76020419 (G>T), a single nucleotide polymorphism located in the miR-145 binding region of ANGPT2 3’UTR. This mutation demonstrates genome-wide significance in GWAS for reduced intraocular pressure (IOP) and primary open-angle glaucoma (POAG) risk, and functions as an expression quantitative trait locus (eQTL) in GTEx database analyses showing ANGPT2 upregulation. The research team employed CRISPR/Cas9 technology to develop this SNP-carrying mouse model, systematically evaluating ANGPT2 expression patterns and Schlemm's canal morphological changes across ocular and systemic tissues.

Background Knowledge
Glaucoma constitutes a major cause of irreversible blindness worldwide, affecting approximately 60 million individuals. Maintaining aqueous humor homeostasis through balanced production and drainage remains essential for IOP regulation, with major outflow facilitated via the trabecular meshwork (TM) and Schlemm's canal (SC). Previous studies identified ANGPT1 and TEK gene loss-of-function mutations in congenital glaucoma pathogenesis, while ANGPT2 demonstrates partial functional compensation in Angpt1-deficient contexts. miR-145 functions as an established ANGPT2 suppressor; the rs76020419 (G>T) SNP disrupts this regulatory interaction, increasing ANGPT2 expression that may expand SC diameter and reduce IOP. Prior absence of an appropriate in vivo model hindered genetic validation, making this study the first to provide direct evidence for ANGPT2's protective role in SC homeostasis and glaucoma prevention.

 

 

Research Methods and Experiments
The research team introduced rs76020419 (G>T) mutation into C57BL/6 mice using CRISPR/Cas9 technology to construct the Angpt2-miR-145 mouse model. ANGPT2 protein levels were quantified in plasma and tissues (eye, lung, kidney) through qPCR and ELISA. Schlemm's canal area was analyzed via immunofluorescence staining combined with confocal microscopy. Intraocular pressure (IOP) was measured using Tonolab rebound tonometry. Fluorescent angiography assessed vascular leakage patterns to confirm model stability.

Key Conclusions and Perspectives

• Mice carrying rs76020419 [T] allele exhibit significantly elevated ANGPT2 protein levels in plasma and ocular/lung/kidney tissues.
• Mutant mice demonstrate markedly increased Schlemm's canal cross-sectional area compared to wild-type (WT) controls.
• Mutant mice display significantly reduced IOP, suggesting SC expansion enhances aqueous humor drainage efficiency.
• Absence of retinal vascular leakage in fluorescent angiography indicates preserved vascular stability despite ANGPT2 upregulation.
• This work provides first in vivo evidence for miR-145 regulation of ANGPT2 expression and its impact on SC development.

Research Significance and Prospects
The study establishes a functional relationship between ANGPT2 overexpression and SC structural modifications, supporting its role as a genetic protective factor against glaucoma. Future investigations should integrate multiple IOP elevation models to comprehensively evaluate this mutation's protective effects against ocular hypertension and retinal neurodegeneration, providing foundational insights for developing ANGPT2/miR-145-targeted glaucoma therapies.

 

 

Conclusion
This study represents the first validation in mice of human ANGPT2 gene mutation rs76020419 (G>T) effects on IOP regulation and Schlemm's canal morphology. Through CRISPR/Cas9 genome editing, the research team successfully generated a mouse model exhibiting ANGPT2 upregulation across multiple tissues. The mutation correlates with expanded SC dimensions and reduced IOP, providing in vivo evidence for miR-145-mediated ANGPT2 regulation. This model establishes a foundation for investigating ANGPT2's protective mechanisms in glaucoma and related neurodegenerative diseases, while identifying novel molecular targets for gene therapy.

 

Naoki Kiyota, Tuncer Onay, Phoebe Leeaw, Janey L Wiggs, and Susan E Quaggin. A Glaucoma-Protective Human SNP in the Angpt2 Locus Increased ANGPT2 Expression and Schlemm’s Canal Area in Mice. Arteriosclerosis, thrombosis, and vascular biology.