AI Article Synopsis

  • Primary open angle glaucoma (POAG) is a major global cause of blindness, linked to the degeneration of retinal cells, with raised intraocular pressure (IOP) being a significant risk factor.
  • The study involved knocking out 62 genes in human trabecular meshwork cells and used advanced techniques like single-cell RNA sequencing and fluorescence analysis to investigate changes in gene expression and cell morphology.
  • Results highlighted key genes (ANGPTL2, LMX1B, CAV1, KREMEN1) affecting POAG, revealing potential genetic networks and cellular changes that could help understand the disease's pathogenesis and explore similar genetic disorders.

Article Abstract

Introduction: Primary open angle glaucoma (POAG) is a leading cause of blindness globally. Characterized by progressive retinal ganglion cell degeneration, the precise pathogenesis remains unknown. Genome-wide association studies (GWAS) have uncovered many genetic variants associated with elevated intraocular pressure (IOP), one of the key risk factors for POAG. We aimed to identify genetic and morphological variation that can be attributed to trabecular meshwork cell (TMC) dysfunction and raised IOP in POAG.

Methods: 62 genes across 55 loci were knocked-out in a primary human TMC line. Each knockout group, including five non-targeting control groups, underwent single-cell RNA-sequencing (scRNA-seq) for differentially-expressed gene (DEG) analysis. Multiplexed fluorescence coupled with CellProfiler image analysis allowed for single-cell morphological profiling.

Results: Many gene knockouts invoked DEGs relating to matrix metalloproteinases and interferon-induced proteins. We have prioritized genes at four loci of interest to identify gene knockouts that may contribute to the pathogenesis of POAG, including ANGPTL2, LMX1B, CAV1, and KREMEN1. Three genetic networks of gene knockouts with similar transcriptomic profiles were identified, suggesting a synergistic function in trabecular meshwork cell physiology. TEK knockout caused significant upregulation of nuclear granularity on morphological analysis, while knockout of TRIOBP, TMCO1 and PLEKHA7 increased granularity and intensity of actin and the cell-membrane.

Conclusion: High-throughput analysis of cellular structure and function through multiplex fluorescent single-cell analysis and scRNA-seq assays enabled the direct study of genetic perturbations at the single-cell resolution. This work provides a framework for investigating the role of genes in the pathogenesis of glaucoma and heterogenous diseases with a strong genetic basis.

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Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC11031357PMC
http://dx.doi.org/10.1093/hmg/ddae003DOI Listing

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