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Matrix Mechanotransduction via Yes-Associated Protein in Human Lamina Cribrosa Cells in Glaucoma.

Publication ,  Journal Article
Murphy, R; Irnaten, M; Hopkins, A; O'Callaghan, J; Stamer, WD; Clark, AF; Wallace, D; O'Brien, CJ
Published in: Invest Ophthalmol Vis Sci
January 3, 2022

PURPOSE: Extracellular matrix stiffening is characteristic of both aging and glaucoma, and acts as a promoter and perpetuator of pathological fibrotic remodeling. Here, we investigate the role of a mechanosensitive transcriptional coactivator, Yes-associated protein (YAP), a downstream effector of multiple signaling pathways, in lamina cribrosa (LC) cell activation to a profibrotic, glaucomatous state. METHODS: LC cells isolated from glaucomatous human donor eyes (GLC; n = 3) were compared to LC cells from age-matched nonglaucomatous controls (NLC; n = 3) to determine differential YAP expression, protein levels, and proliferation rates. NLC cells were then cultured on soft (4 kPa), and stiff (100 kPa), collagen-1 coated polyacrylamide hydrogel substrates. Quantitative real-time RT-PCR, immunoblotting, and immunofluorescence microscopy were used to measure the expression, activity, and subcellular location of YAP and its downstream targets, respectively. Proliferation rates were examined in NLC and GLC cells by methyl thiazolyl tetrazolium salt assays, across a range of incrementally increased substrate stiffness. Endpoints were examined in the presence or absence of a YAP inhibitor, verteporfin (2 µM). RESULTS: GLC cells show significantly (P < 0.05) increased YAP gene expression and total-YAP protein compared to NLC cells, with significantly increased proliferation. YAP regulation is mechanosensitive, because NLC cells cultured on pathomimetic, stiff substrates (100 kPa) show significantly upregulated YAP gene and protein expression, increased YAP phosphorylation at tyrosine 357, reduced YAP phosphorylation at serine 127, increased nuclear pooling, and increased transcriptional target, connective tissue growth factor. Accordingly, myofibroblastic markers, α-smooth muscle actin (α-SMA) and collagen type I, alpha 1 (Col1A1) are increased. Proliferation rates are elevated on 50 kPa substrates and tissue culture plastic. Verteporfin treatment significantly inhibits YAP-mediated cellular activation and proliferation despite a stiffened microenvironment. CONCLUSIONS: These data demonstrate how YAP plays a pivotal role in LC cells adopting a profibrotic and proliferative phenotype in response to the stiffened LC present in aging and glaucoma. YAP provides an attractive and novel therapeutic target, and its inhibition via verteporfin warrants further clinical investigation.

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Published In

Invest Ophthalmol Vis Sci

DOI

EISSN

1552-5783

Publication Date

January 3, 2022

Volume

63

Issue

1

Start / End Page

16

Location

United States

Related Subject Headings

  • YAP-Signaling Proteins
  • RNA
  • Proto-Oncogene Proteins c-yes
  • Optic Disk
  • Ophthalmology & Optometry
  • Mechanotransduction, Cellular
  • Humans
  • Glaucoma
  • Gene Expression Regulation
  • Gene Expression Profiling
 

Citation

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Murphy, R., Irnaten, M., Hopkins, A., O’Callaghan, J., Stamer, W. D., Clark, A. F., … O’Brien, C. J. (2022). Matrix Mechanotransduction via Yes-Associated Protein in Human Lamina Cribrosa Cells in Glaucoma. Invest Ophthalmol Vis Sci, 63(1), 16. https://doi.org/10.1167/iovs.63.1.16
Murphy, Rory, Mustapha Irnaten, Alan Hopkins, Jeffrey O’Callaghan, W Daniel Stamer, Abbot F. Clark, Deborah Wallace, and Colm J. O’Brien. “Matrix Mechanotransduction via Yes-Associated Protein in Human Lamina Cribrosa Cells in Glaucoma.Invest Ophthalmol Vis Sci 63, no. 1 (January 3, 2022): 16. https://doi.org/10.1167/iovs.63.1.16.
Murphy R, Irnaten M, Hopkins A, O’Callaghan J, Stamer WD, Clark AF, et al. Matrix Mechanotransduction via Yes-Associated Protein in Human Lamina Cribrosa Cells in Glaucoma. Invest Ophthalmol Vis Sci. 2022 Jan 3;63(1):16.
Murphy, Rory, et al. “Matrix Mechanotransduction via Yes-Associated Protein in Human Lamina Cribrosa Cells in Glaucoma.Invest Ophthalmol Vis Sci, vol. 63, no. 1, Jan. 2022, p. 16. Pubmed, doi:10.1167/iovs.63.1.16.
Murphy R, Irnaten M, Hopkins A, O’Callaghan J, Stamer WD, Clark AF, Wallace D, O’Brien CJ. Matrix Mechanotransduction via Yes-Associated Protein in Human Lamina Cribrosa Cells in Glaucoma. Invest Ophthalmol Vis Sci. 2022 Jan 3;63(1):16.

Published In

Invest Ophthalmol Vis Sci

DOI

EISSN

1552-5783

Publication Date

January 3, 2022

Volume

63

Issue

1

Start / End Page

16

Location

United States

Related Subject Headings

  • YAP-Signaling Proteins
  • RNA
  • Proto-Oncogene Proteins c-yes
  • Optic Disk
  • Ophthalmology & Optometry
  • Mechanotransduction, Cellular
  • Humans
  • Glaucoma
  • Gene Expression Regulation
  • Gene Expression Profiling