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Rocio Vargas-Pinto, Kristin M. Perkumas, W Daniel Stamer, Mark Johnson; Modulus of Elasticity of Healthy and Glaucomatous Schlemm’s Canal Cells. Invest. Ophthalmol. Vis. Sci. 2011;52(14):4814.
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© ARVO (1962-2015); The Authors (2016-present)
Changes in stiffness of the inner wall endothelium of Schlemm’s Canal (SC) may contribute to the increased outflow resistance characteristic of glaucoma. We used Atomic Force Microscopy (AFM) with various tip sizes to assess the differences in elasticity between cultured SC cells from normal and glaucomatous donors.
SC cells were isolated from healthy donors (4 normal cell strains) and donors previously diagnosed with glaucoma (4 glaucoma cell strains). Cultured cells were stained with Calcein AM and Hoechst fluorescent dyes to determine cell viability and the location of the nucleus respectively. AFM force measurements were made using silicon nitride sharp pyramidal tips (nominal angle of 35°) and tips with polystyrene spherical attachments (diameters of 4.5 and 10 µm). Young’s Modulus (E) was calculated using the modified Hertz model. The AFM indentation was modeled using finite element software to determine the effect cortex stiffness would have on the measured modulus based on the tip geometry.
Young’s Modulus for SC cells measured with pyramidal tips was similar for normal cells (9.4± 12.4 kPa, n=76) and glaucoma cells (9.1± 10.6 kPa, n=98). When using spherical tips, lower values were obtained for normal cells (4.5 µm tip: 2.0± 2.4 kPa, n=69; 10 µm tip: 2.2± 3.5 kPa, n=108) compared to glaucoma cells (4.5 µm tip: 3.1± 3.9 kPa, n=75; 10 µm tip: 3.9± 9.9 kPa, n=82. Also, the mean variance in E measured with spherical tips was seven times greater for glaucomatous cells. No significant differences in stiffness were found between the cytoplasm and nuclear regions for either normal or glaucoma cells. The finite element model showed that measurements using the pyramidal tip are strongly affected by the stiffness of the cell cortex, while values measured with the spherical tip are only weakly influenced by the cortex stiffness.
The significantly higher modulus and greatly increased variance measured using spherical tips suggest that there are differences in stiffness between normal and glaucomatous SC cells. The lack of such a finding using pyramidal tips indicates that the altered stiffness is likely in the cell body and not in the cell cortex.
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