Increased hydrostatic pressure does not cause loss of retinal ganglion cell viability in human organotypic retinal cultures

Abstract

AbstractPurpose Raised intraocular pressure is the major risk factor for glaucoma. It is therefore vitally important to understand how increased pressure impacts on retinal ganglion cells (RGC) viability. These experiments investigate the effect of “pure” hydrostatic pressure on RGC survival in human organotypic retinal cultures (HORCs).Methods In order to achieve this, we have designed and built a highly sophisticated controlled environment pressure chamber to expose cells to increased hydrostatic pressure while maintaining constancy of other parameters.Results Exposure of HORCs to constant (60mmHg) or fluctuating (0‐100 mmHg; 1 cycle/min) pressure for 24 or 48h resulted in no loss of structural integrity compared to controls (n=4). No significant changes in levels of RGC markers (THY‐1 mRNA or NeuN) or LDH release were observed (n=4; p>0.05). RGC‐5 cells, used for comparison, showed no change in cell viability (MTS; n=4), LDH release (n=3) or apoptosis (TUNEL; n=3) with pressure at 24 or 72h (p>0.05). Oxygen/glucose deprivation in HORCs reduced RGC number (~40%) at 24h (n=9; p<0.05) and increased LDH release (~80%; n=10; p<0.05). p38 and JNK activation remained unchanged in HORCs exposed to pressure (n=3; p>0.05) whereas OGD increased activation of both (n=3; p<0.05).Conclusion These data clearly show that increased hydrostatic pressure on its own is insufficient to cause loss of cell viability, suggesting that direct hydrostatic pressure is not responsible for degeneration of RGCs occurring in glaucoma.