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

This study successfully constructed an Angpt2-miR-145 mouse model carrying the rs76020419 (G>T) mutation, validating that this mutation elevates ANGPT2 expression and enlarges Schlemm's canal area, suggesting its potential role in glaucoma protection mechanisms. This model provides a critical tool for future exploration of ANGPT2's functions in ocular hypertension and retinal neurodegeneration.

 

Literature Overview
This article titled 'A Glaucoma-Protective Human SNP in the Angpt2 Locus' published in Arteriosclerosis, Thrombosis, and Vascular Biology reviews functional validation of rs76020419 (G>T), a human SNP located in the miR-145 binding site within the ANGPT2 3'UTR region. The mutation has been associated with reduced intraocular pressure (IOP) and decreased risk of primary open-angle glaucoma in GWAS studies, and functions as an eQTL in the GTEx database linked to ANGPT2 upregulation. The research team employed CRISPR/Cas9 technology to develop a murine model carrying this SNP, evaluating ANGPT2 expression and Schlemm's canal morphological changes across ocular and systemic tissues.

Background Knowledge
Glaucoma represents a leading cause of blindness globally, affecting approximately 60 million individuals. Maintaining aqueous humor production and drainage equilibrium is critical for IOP regulation, with fluid primarily drained through the trabecular meshwork (TM) and Schlemm's canal (SC). Previous studies identified loss-of-function mutations in ANGPT1 and TEK genes associated with congenital glaucoma, while ANGPT2 demonstrates partial functional compensation in Angpt1-deficient contexts. miR-145, a known ANGPT2 inhibitor, binds this specific site; however, the rs76020419 (G>T) SNP disrupts miR-145 binding, elevating ANGPT2 expression which may dilate SC and reduce IOP. This study fills a critical gap by providing the first in vivo mouse model for investigating this genetic mechanism.

 

 

Research Methods and Experiments
The research team introduced rs76020419 (G>T) mutation into C57BL/6 mice using CRISPR/Cas9 technology. ANGPT2 protein expression levels were quantified in plasma and ocular, pulmonary, and renal tissues through quantitative PCR (qPCR) and ELISA. Schlemm's canal morphological changes were analyzed via immunofluorescence staining and confocal microscopy. Intraocular pressure (IOP) was measured using Tonolab rebound tonometry. Fluorescein angiography assessed vascular leakage to confirm model stability.

Key Conclusions and Perspectives

• Mice carrying the rs76020419 [T] allele exhibited significantly elevated ANGPT2 protein levels in plasma and ocular, pulmonary, and renal tissues.
• Mutant mice demonstrated markedly expanded Schlemm's canal cross-sectional area compared to wild-type (WT) controls.
• Mutant mice showed significant IOP reduction, suggesting canal expansion enhances aqueous drainage efficiency.
• Absence of retinal vascular leakage in fluorescein angiography indicates preserved vascular stability despite ANGPT2 elevation.
• This study provides the first in vivo evidence demonstrating miR-145 regulation of ANGPT2 expression and its role in SC development.

Research Significance and Prospects
The study establishes a functional relationship between ANGPT2 upregulation and structural modifications in Schlemm's canal, supporting its potential as a glaucoma-protective genetic factor. Future investigations should integrate multiple ocular hypertension models to evaluate this mutation's neuroprotective effects and therapeutic potential for targeting ANGPT2 or miR-145 pathways in glaucoma treatment.

 

 

Conclusion
This study represents the first validation in a murine model of human ANGPT2 gene rs76020419 (G>T) mutation's impact on IOP regulation and Schlemm's canal morphology. Using CRISPR/Cas9 gene editing, the research team successfully developed a mouse model carrying this specific mutation, confirming ANGPT2 protein upregulation across multiple tissues. Mutant mice exhibited both Schlemm's canal expansion and reduced IOP, providing in vivo evidence for miR-145's regulatory role in ANGPT2 expression. This model establishes a foundation for investigating ANGPT2's protective mechanisms in glaucoma and related neurodegenerative diseases, while identifying novel molecular targets for genetic therapies.

 

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.