Shear stress-triggered nitric oxide release from Schlemm's canal cells.
OBJECTIVE
Endothelial nitric oxide (NO) synthase is regulated by shear stress. At elevated intraocular pressures when the Schlemm's canal (SC) begins to collapse, shear stress is comparable with that in large arteries. We investigated the relationship between NO production and shear stress in cultured human SC cells.


METHODS
Schlemm's canal endothelial cells isolated from three normal and two glaucomatous human donors were seeded into Ibidi flow chambers at confluence, cultured for 7 days, and subjected to steady shear stress (0.1 or 10 dynes/cm(2)) for 6, 24, or 168 hours. Cell alignment with flow direction was monitored, and NO production was measured using 4-amino-5-methylamino-2',7'-difluorofluorescein (DAF-FM) and Griess reagents. Human trabecular meshwork (TM) and umbilical vein endothelial cells (HUVECs) were used as controls.


RESULTS
Normal SC strains aligned with the direction of flow by 7 days. Comparing 0.1 vs. 10 dynes/cm(2), NO levels increased by 82% at 24 hours and 8-fold after 7 days by DAF-FM, and similar results were obtained with Griess reagent. Shear responses by SC cells at 24 hours were comparable with HUVECs, and greater than TM cells, which appeared shear-insensitive. Nitric oxide production by SC cells was detectable as early as 6 hours and was inhibited by 100 μM nitro-L-arginine methyl ester. Two glaucomatous SC cell strains were either unresponsive or lifted from the plate in the face of shear.


CONCLUSIONS
Shear stress triggers NO production in human SC cells, similar to other vascular endothelia. Increased shear stress and NO production during SC collapse at elevated intraocular pressures may in part mediate IOP homeostasis.
Biomedical Engineering, Duke University, Durham, North Carolina, United States.
Nicole E
NE
Ashpole
Department of Bioengineering, Imperial College London, London, United Kingdom.
Darryl R
DR
Overby
Department of Bioengineering, Imperial College London, London, United Kingdom Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, Georgia, United States.
C Ross
CR
Ethier
Ophthalmology Department, Duke University, Durham, North Carolina, United States.
W Daniel
WD
Stamer
D000368:Aged; D000369:Aged, 80 and over; D000704:Analysis of Variance; D001082:Aqueous Humor / Q000378:metabolism*; D016022:Case-Control Studies; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000502:physiology*; D005260:Female; D005901:Glaucoma / Q000473:pathology / Q000503:physiopathology; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D007429:Intraocular Pressure / Q000502:physiology; D008297:Male; D008875:Middle Aged; D009569:Nitric Oxide / Q000096:biosynthesis*; D019001:Nitric Oxide Synthase / Q000378:metabolism; D033081:Shear Strength; D013314:Stress, Mechanical; D014129:Trabecular Meshwork / Q000166:cytology* / Q000502:physiology
D009569:Nitric Oxide; D019001:Nitric Oxide Synthase
aqueous humor; conventional outflow; eNOS; glaucoma; intraocular pressure