RNA-seq transcriptomic profiling of TGF-β2-exposed human trabecular meshwork explants: Advancing insights beyond conventional cell culture models

Exp Cell Res. 2024 Oct 1;442(2):114220. doi: 10.1016/j.yexcr.2024.114220. Epub 2024 Aug 28.

Abstract

Primary open-angle glaucoma (POAG), a leading cause of irreversible vision loss, is closely linked to increased intraocular pressure (IOP), with the trabecular meshwork (TM) playing a critical role in its regulation. The TM, located at the iridocorneal angle, acts as a sieve, filtering the aqueous humor from the eye into the collecting ducts, thus maintaining proper IOP levels. The transforming growth factor-beta 2 (TGF-β2) signaling pathway has been implicated in the pathophysiology of primary open-angle glaucoma POAG particularly, in the dysfunction of the TM. This study utilizes human TM explants to closely mimic in vivo conditions, thereby minimizing transcriptional changes that could arise from cell culture enabling an exploration of the transcriptomic impacts of TGF-β2. Through bulk RNA sequencing and immunohistological analysis, we identified distinct gene expression patterns and morphological changes induced by TGF-β2 exposure (5 ng/ml for 48 h). Bulk RNA sequencing identified significant upregulation in genes linked to extracellular matrix (ECM) regulation and fibrotic signaling. Immunohistological analysis further elucidated the morphological alterations, including cytoskeletal rearrangements and ECM deposition, providing a visual confirmation of the transcriptomic data. Notably, the enrichment analysis unveils TGF-β2's influence on both bone morphogenic protein (BMP) and Wnt signaling pathways, suggesting a complex interplay of molecular mechanisms contributing to TM dysfunction in glaucoma. This characterization of the transcriptomic modifications on an explant model of TM obtained under the effect of this profibrotic cytokine involved in glaucoma is crucial in order to develop and test new molecules that can block their signaling pathways.

Keywords: Fibrosis; Glaucoma; RNA-Sequencing; Trabecular meshwork; Transforming growth factor-beta 2.

MeSH terms

  • Cell Culture Techniques
  • Extracellular Matrix / genetics
  • Extracellular Matrix / metabolism
  • Gene Expression Profiling / methods
  • Glaucoma, Open-Angle* / genetics
  • Glaucoma, Open-Angle* / metabolism
  • Glaucoma, Open-Angle* / pathology
  • Humans
  • Intraocular Pressure
  • RNA-Seq* / methods
  • Signal Transduction / drug effects
  • Trabecular Meshwork* / drug effects
  • Trabecular Meshwork* / metabolism
  • Trabecular Meshwork* / pathology
  • Transcriptome / drug effects
  • Transcriptome / genetics
  • Transforming Growth Factor beta2* / genetics
  • Transforming Growth Factor beta2* / metabolism
  • Transforming Growth Factor beta2* / pharmacology

Substances

  • Transforming Growth Factor beta2
  • TGFB2 protein, human