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P. Russell, J. Z. Gasiorowski, P. F. Nealey, C. J. Murphy; Nanoscale Topographic Cues Modulate Cell Behavior and Differentially Shape Gene and Protein Expression for Human Trabecular Meshwork Cells. Invest. Ophthalmol. Vis. Sci. 2008;49(13):3290.
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© ARVO (1962-2015); The Authors (2016-present)
To determine how nanoscale topographic cues influence human trabecular meshwork (HTM) cells.
HTM cells from different donors were plated on flat or anisotropically ordered nanopatterned surfaces comprised of repeating patterns of grooves and ridges. Analysis of cellular elongation and alignment was done using fluorescence microscopy. Cells were grown with or without 100 nM dexamethasone to determine if patterned surfaces influenced myocilin, versican, and alpha B-crystallin. Quantitative PCR was performed to examine mRNA levels of myocilin and versican isoforms. Western blots for myocilin and alpha B-crystallin were also done.
Anisotropically ordered nanopatterned surfaces caused HTM cells to align with the grooved patterns and actin filaments similarly oriented parallel to the patterned surface. Cells were markedly elongated on the nanogrooved substrates. Growth of cells on the 400 nm pitch surfaces increased mRNA expression for myocilin between 2 to 15 fold compared to cells on flat surfaces. Treatment with dexamethasone caused increases in myocilin mRNA on flat surfaces as expected, an effect that was augmented by the provision of the biomimetic grooved surfaces, increasing myocilin mRNA expression between 6 and 23 fold compared to cells grown on flat surfaces. Western blot verified increased myocilin protein levels for cells grown on nanopatterned surfaces from approximately 2 to 19 fold. Alpha B-crystallin expression was unaffected by the provision of topographic cues. The mRNA for versican isoforms, particularly the V0 and V1 isoforms, was increased in HTM cells by the presence of nanotopography although increases were generally 5 fold or less.
Nanoscale topographically patterned surfaces profoundly influence HTM cells. Cues from these surfaces affect cell orientation and elongation. Myocilin mRNA and certain of the versican isoform mRNAs were increased substantially when the cells were grown on biomimetic topographically patterned surfaces. The amount of myocilin protein was also increased, but western blots of alpha B-crystallin showed that not all proteins were influenced by these cues. The data suggest HTM cells grown on flat tissue culture plastic are not sensing some of the cues that they receive in vivo from the basement membranes upon which they dwell. These results point out the importance of topographic cueing for trabecular meshwork cells.
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