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C. R. Ethier, Y. Lei, S. Rajabi, A. T. Read, D. R. Overby; Small Elevations of Pressure Do Not Affect Cultured Astrocytes But Gas Tension in the Media Does. Invest. Ophthalmol. Vis. Sci. 2010;51(13):2096.
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The response of optic nerve head (ONH) cells to mechanical stress may be important in glaucoma. Studies have reported biological effects of pressure on cultured ONH cells. Here we designed an apparatus to independently evaluate effects of pressure and of culture media gas tension (including oxygen tension) on cells.
DI TNC1 rat cortical astrocytes (ATCC) were seeded onto coverslips. A cell-free area (dia 4 mm) was created and cell migration rate and alpha-tubulin architecture were evaluated at 24, 48 and 74 hrs (Salvador-Silva et al, Glia, 2004). During the assay period, cells were exposed to one of four experimental conditions to independently test the effects of pressure and gas tension. 1) Control pressure, control gas tension: the coverslip was placed under a 15 mm tall column of media in an incubator under standard conditions. 2) High pressure, reduced gas tension: as in 1, but the media column was 100 mm tall (pressure = 7.4 mmHg). 3) Control pressure, reduced gas tension: the coverslip was placed in a horizontal column of media sized so that the pressure was as in 1, but the gas tension at the cells was as in 2. 4) High pressure, control gas tension: a closed culture chamber was connected to a gas exchange reservoir by a circulating pump (Obazawa et al, IOVS, 2004), designed so that cultured cells received fresh media with the same pressure as in 2 and gas tensions as in 1.
Calculations showed that cells in conditions 2 and 3 were hypoxic after ~48 hrs in culture. No effects of pressure were observed on cell migration rate or alpha-tubulin architecture. However, cells cultured under low gas tension (conditions 2 and 3) showed increased mobility at 48 and 72 hrs (p < 0.05), coincident with the predicted onset of hypoxia (Figure).
7.4 mmHg pressure alone has no effect on DI TNC1 astrocytes cultured on rigid coverslips, while hypoxia associated with a fluid column creating this pressure does. These results differ from a previous report (Salvador-Silva et al), the results of which may be explained by altered gas tensions in the culture media.
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