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Christopher M. Reilly, Joshua A. Wood, Christopher J. Murphy, Paul Russell; Nanoscale Topographic Cues And Antioxidant Status In Human Trabecular Meshwork Cells. Invest. Ophthalmol. Vis. Sci. 2011;52(14):6676.
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We have recently demonstrated that the biophysical properties of trabecular meshwork (HTM) are altered in glaucoma. Oxidative stress is also implicated in glaucoma progression. We hypothesized that HTM cells cultured on topographically patterned surfaces would have higher total antioxidant capacity than those cultured on tissue culture polystyrene, and that topographic substrates would modulate HTM cell response to oxidative challenge.
Primary HTM cell cultures were obtained from donor corneas unsuitable for transplant. Cells were used prior to the 7th passage. Cells from 4 donors were cultured in triplicate to confluence on polyurethane coated culture dishes with planar, 400nm, 1400nm, or 4000nm pitch (pitch = ridge width + groove width). Cells were treated with 250 micromolar hydrogen peroxide in serum free media or untreated serum free media (control) for 2 hours. Cells were harvested with trypsinization, counted, and digested in 0.1% Triton X-100. Total antioxidant status (TAS) was determined using a commercial colorimetric ABTS assay for Trolox equivalents, read at 645nm, and compared to a standard curve. Results were expressed as Trolox equivalents/105 cells, and normalized to the planar untreated control value.
TAS in untreated HTM cells was increased 1.8 and 1.5 fold on 1400nm and 4000nm surfaces, respectively, compared to planar controls. TAS was unchanged on 400nm surfaces. TAS was reduced by 0.6 to 0.7 fold in all peroxide treated groups.
These results suggest that topographic cues in the biomimetic range differentially modulate TAS in HTM cells. This may have implications in the progression of glaucoma, as HTM biophysical properties change. Hydrogen peroxide treatment lowered TAS to nearly identical levels in all groups, indicating topographic cues did not help maintain TAS in response to this acute oxidative stress in HTM cells.
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