April 2010
Volume 51, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2010
Biophysical Cues Modulate TGM2 and MMP3 Expression in Cultured Human Trabecular Meshwork Cells
Author Affiliations & Notes
  • C. M. Reilly
    School of Veterinary Medicine: Pathology, Microbiology, and Immunology,
    University of California, Davis, California
  • H. Mochizuki
    School of Medicine: Ophthalmology,
    University of California, Davis, California
  • C. J. Murphy
    School of Medicine: Ophthalmology,
    University of California, Davis, California
  • P. Russell
    School of Veterinary Medicine: Surgical and Radiological Sciences,
    University of California, Davis, California
  • Footnotes
    Commercial Relationships  C.M. Reilly, None; H. Mochizuki, None; C.J. Murphy, None; P. Russell, None.
  • Footnotes
    Support  National Center For Medical Research Grant UL1 RR024146
Investigative Ophthalmology & Visual Science April 2010, Vol.51, 3213. doi:https://doi.org/
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      C. M. Reilly, H. Mochizuki, C. J. Murphy, P. Russell; Biophysical Cues Modulate TGM2 and MMP3 Expression in Cultured Human Trabecular Meshwork Cells. Invest. Ophthalmol. Vis. Sci. 2010;51(13):3213. doi: https://doi.org/.

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

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Abstract

Purpose: : To determine the effect of biophysical cues on the expression of transglutaminase-2 (TGM2) and matrix metalloproteinase-3 (MMP3) by cultured human trabecular meshwork (HTM) cells.

Methods: : HTM cells were harvested from donor corneas unsuitable for human transplant and cells were used at ≤ 7th passage. Cells were grown for 2 and 4 days on polyacrylamide substrates of varying biomimetic compliance (3 kPa, 30 kPa, 100 kPa) and cell culture plastic as a control. HTM cells were also grown for 3 days topographically patterned polyurethane surfaces having parallel ridges and grooves of 400 nm, 1400 nm, and 4000 nm pitch, using a planar surface as a control. RNA was extracted and real-time quantitative PCR was performed to determine TGM2 and MMP3 mRNA expression levels. Ribosomal 18S RNA was used as a loading control.

Results: : On Day 2, TGM2 expression was reduced to 66, 76, and 82% of control levels on 3 kPa, 30 kPa, and 100 kPa surfaces, respectively. On Day 4, TGM2 levels returned to control levels on all surfaces. MMP3 expression on Day 2 was increased by 1.3, 4.2, and 2.1 fold on 3 kPa, 30 kPa, and 100 kPa surfaces, respectively. On Day 4, MMP3 levels were increased roughly two-fold over control levels on all substrates. TGM2 levels on patterned surfaces were increased 1.5, 2.1, and 2.5 fold over control levels on 400 nm, 1400 nm, and 4000 nm pitch surfaces, respectively.

Conclusions: : TGM2 expression increased as substrate compliance decreased (stiffer), but the effect was transient. MMP3 levels were greatest on the 30 kPa substrate at Day 2, but remained elevated on all compliant surfaces on Day 4. These results suggest that changes in the biophysical properties of the ECM modulate expression of TGM2 and MMP3, which in turn affect the composition and turnover of the ECM. TGM2 decreases matrix turnover by crosslinking fibronectin and this enzyme has been shown to be increased in glaucoma. Recent work in our laboratory indicates that HTM compliance decreases in glaucoma, which may be related to ECM deposition and turnover. There was a pitch dependent increase in TGM2 levels on patterned surfaces, demostrating that distinct biophysical cues influence HTM cell behaviors. Additionally, these results suggest that HTM cellular responses in cultures grown on standard cultureware may not be reflective of in vivo responses.

Keywords: trabecular meshwork • extracellular matrix 
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