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R. Rogers, S. Culp-Stewart, J. G. Flanagan; The Mechanobiologic Response of Optic Nerve Head Cells to Mechanical Strain. Invest. Ophthalmol. Vis. Sci. 2009;50(13):888.
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Mechanical strain of optic nerve head (ONH) tissues likely plays a role in glaucomatous optic neuropathy. In this study we use proteomics to investigate the response of ONH cells to mechanical strain.
Human lamina cribrosa (LC) cells and astrocytes were isolated and grown from donor tissue (Eye Bank of Canada) in DMEM (4 mM L-glutamine;1g/L glucose; 1.5 g/L sodium bicarbonate; 10 % FBS; penicillin/ streptomycin) and DMEM/F12 (10 % FBS; penicillin/ streptomycin) at 37°C in a 5 % CO2 humidified incubator, respectively. Cells were seeded onto collagen I coated (LC cells) and collagen I and IV coated (astrocytes cells) BioFlex culture plates and grown to confluence. Cells were rinsed with DPBS and grown for 24 hours in serum-free media. The cells were then subjected to 3 or 12 % cyclic (1 Hz, sinusoid) equi-axial stretch for 2 or 24 hours using the Flexercell FX-4000 Tension Plus System. Control cells were serum-deprived and incubated without stretch for the duration of the experiment. Nano LC- MS/MS analysis using iTRAQ labelled samples and 2-D gel electrophoresis were both performed on the resulting cell lysates to determine up or down regulation of proteins.
There was a consistent and repeatable differential expression of proteins involved in the activation of astrocytes as well as the remodelling of the LC extracellular matrix for all test conditions relative to controls. This included upregulation of MMPs, GFAP and TGFβ.
The cells within the LC react in vitro to mechanical strain by upregulating proteins, such as TGFβ, that are believed to cause astrocytes to change into their active phenotype. Proteins responsible for cellular remodelling, such as MMPs, are also upregulated and implicated in degrading components of the LC ECM such as collagen, elastin and laminin.
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