Endothelial Nitric Oxide Synthase-Related Mechanotransduction Changes in Aged Porcine Angular Aqueous Plexus Cells

Year: 2014

Type: article

Abstract: PURPOSE: To investigate effects of aging on endothelial nitric oxide synthase (eNOS) expression and signaling in angular aqueous plexus (AAP) (functional equivalent to human Schlemm's canal) cells subjected to shear stress. METHODS: The AAP cells were isolated differentially from porcine outflow tissues using puromycin selection. Cell aging was induced by culturing cells in hyperoxia condition (40% oxygen and 5% carbon dioxide) for 14 days. The AAP cells grown in chamber slides were exposed to a shear stress of 8 dynes/cm(2) for 24 hours. Expression of eNOS, eNOS-phospho Thr495, eNOS-phospho Ser1177, and Akt-phospho was tested by Western blot analysis and immunofluorescence staining. Nitric oxide (NO) levels were measured using the Griess assay. RESULTS: Compared with control, eNOS levels in aged cells were significantly reduced by 60% (P < 0.05; n = 6). Phosphorylation of eNOS at Ser1177 and Akt at Ser473 was 63% and 80% lower in aged cells, respectively, whereas phosphorylation of the eNOS inhibition site (Thr495) increased by 6.1-fold (P < 0.05; n = 6). Shear stress (8 dynes/cm(2) for 24 hours) increased eNOS abundance (total protein and at cell borders) and phosphorylation at Ser1177 by 1.7-fold and 1.8-fold, respectively (P < 0.05; n = 6), whereas aged cells were unresponsive. In control cells exposed to shear stress, the NO concentration was 1.8-fold higher than in the static group (P < 0.05; n = 4); however, aged cells were unresponsive to shear stress (mean ± SD, 4.3 ± 1.3 vs. 4.1 ± 1.4 μM). CONCLUSIONS: Aged AAP cells appear compromised in their mechanotransduction machinery involving eNOS, the protein product of the gene, NOS3, polymorphisms of which impart a risk for the development of glaucoma.

Source: Investigative Ophthalmology & Visual Science

DOI: 10.1167/iovs.14-14992

Authors: Y. Lei, W. D. Stamer, J. Wu, X. Sun

Institutions: Shanghai Medical College of Fudan University, Fudan University, Eye & ENT Hospital of Fudan University, Duke University, State Key Laboratory of Medical Neurobiology

Language: English

Topic: Glaucoma and retinal disorders

Subfield: Ophthalmology

Field: Medicine

Domain: Health Sciences

SDG: Good health and well-being

Open Access status: bronze

Funders: National Science Foundation, Research to Prevent Blindness, National Natural Science Foundation of China, Fudan University, National Health and Family Planning Commission of the People's Republic of China, Shanghai Municipal Health Bureau, Allergan, National Eye Institute

OpenAlex ID: W2082516236