June 2020
Volume 61, Issue 7
Open Access
ARVO Annual Meeting Abstract  |   June 2020
LOXL1-AS1 regulates extracellular matrix remodeling and mechanotransduction in human primary trabecular meshwork and Schlemm’s canal cells
Author Affiliations & Notes
  • Heather Schmitt
    Duke University, Durham, North Carolina, United States
  • William M. Johnson
    Duke University, Durham, North Carolina, United States
  • Maria Gomez-Caraballo
    Duke University, Durham, North Carolina, United States
  • Kristin Marie Perkumas
    Duke University, Durham, North Carolina, United States
  • Michael A Hauser
    Duke University, Durham, North Carolina, United States
  • Daniel Stamer
    Duke University, Durham, North Carolina, United States
  • Footnotes
    Commercial Relationships   Heather Schmitt, None; William Johnson, None; Maria Gomez-Caraballo, None; Kristin Perkumas, None; Michael Hauser, None; Daniel Stamer, None
  • Footnotes
    Support  Research to Prevent Blindness Unrestricted Grant, 5P30EY005722, Pseudo exfoliation Hudson Glaucoma Project Gift
Investigative Ophthalmology & Visual Science June 2020, Vol.61, 4629. doi:
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      Heather Schmitt, William M. Johnson, Maria Gomez-Caraballo, Kristin Marie Perkumas, Michael A Hauser, Daniel Stamer; LOXL1-AS1 regulates extracellular matrix remodeling and mechanotransduction in human primary trabecular meshwork and Schlemm’s canal cells. Invest. Ophthalmol. Vis. Sci. 2020;61(7):4629.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Pseudoexfoliation glaucoma (PEXG) is the most common secondary form of open angle glaucoma, and it is clinically characterized by ocular hypertension and the visible accumulation of fibrillary material on tissues in the anterior segment. SNPs located in the LOXL1-AS1 lncRNA transcript are highly associated with risk of PEXG. When expression of LOXL1-AS1 is altered in cultured lens epithelial cells, expression of >300 genes significantly change, including extracellular matrix (ECM) remodeling genes. Unknown is the regulatory role of LOXL1-AS1 in the cells that control aqueous outflow resistance and thus set intraocular pressure. The goal of the present study is to test whether LOXL1-AS1 regulates gene sets responsible for ECM remodeling and mechanotransduction signaling in primary human trabecular meshwork (TM) and Schlemm’s canal (SC) cells.

Methods : Adenoviruses encoding shRNA targeted to LOXL1-AS1 were used to knock down expression in TM and SC cells. At 2-4 days post transduction, cells were processed for qPCR array and Western blot analysis of proteins involved in regulating ECM remodeling and mechanotransduction. Since cell-cell adhesion and morphology are linked with ECM remodeling and mechanotransduction, these metrics were assessed by immunofluorescence using zonula occludens-1 and paxillin antibody labeling.

Results : Knockdown of LOXL1-AS1 in SC cells led to significant expression changes in ECM target genes including neuronal adhesion protein, vascular adhesion protein, integrin alpha 2, and laminin gamma 1 (p<0.05). With LOXL1-AS1 knockdown, SC cells significantly increased the proportion of activated AKT (p=0.008), but no changes were observed with phosphorylation status of FAK or MAPK. Knockdown of LOXL1-AS1 resulted in a shortening of the major axis of SC cells (p=0.006). LOXL1-AS1 regulates genes that encode proteins responsible for ECM remodeling and mechanotransduction signaling in TM and SC cells, indicating a role in conventional outflow homeostasis. Also, knockdown of LOXL1-AS1 led to dysregulation of a small subset of ECM target genes including TUBGCP5, MMP2, MMP14, MMP28, MYC, DDX21, EIF5, PSMB9, IFIT1, OAS1 in one TM cell strain.

Conclusions : LOXL1-AS1 regulates genes that encode proteins responsible for ECM remodeling and mechanotransduction signaling in TM and SC cells, indicating a role in conventional outflow homeostasis.

This is a 2020 ARVO Annual Meeting abstract.

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