Basal nitric oxide production is enhanced by hydraulic pressure in cultured human trabecular cells

Abstract

Background/aims - Nitric oxide donors reduce intraocular pressure. Human trabecular cells in culture were examined for their nitric oxide production in response to hydraulic pressure. Methods - Human trabecular cells were cultured from trabeculum tissue fragments excised during trabeculectomy and exposed to hydraulic pressure change in a culture flask connected to a glass syringe. The pressure was exerted by automatic infusion of the piston of the syringe and monitored by a pressure gauge. The intracellular nitric oxide level was measured in real time with a nitric oxide binding fluorescent dye, diaminofluorescein-2. Results - Intracellular nitric oxide levels in cultured trabecular cells showed spontaneous fluctuation during 400 seconds of observation. Peak levels of intracellular nitric oxide were significantly higher at hydraulic pressure of 30, 40, and 50 mmHg, compared with 0 and 25 mmHg (p < 0.0001, one way ANOVA, and p < 0.05, Tukey-Kramer test). The fluctuation was completely abolished by the presence of N-methyl-L-arginine (L-NMMA), a nitric oxide synthase inhibitor. The cultured trabecular cells were shown by immunohistochemistry to express brain nitric oxide synthase (bNOS). Conclusion - Higher levels of hydraulic pressure enhanced basal production of nitric oxide in human trabecular cells. Nitric oxide would be a physiological mediator in the regulation of intraocular pressure.