Cytoskeletal dependence of adenosine triphosphate release by human trabecular meshwork cells.

Journal Article (Journal Article)

PURPOSE: To test whether adenosine triphosphate (ATP) release links cytoskeletal remodeling with release of matrix metalloproteinases (MMPs), regulators of outflow facility and intraocular pressure. METHODS: ATP release was measured by luciferin-luciferase. Ecto-ATPases from transformed human trabecular meshwork (TM) cells (TM5) and explant-derived TM cells were identified by RT-PCR. Actin was visualized by phalloidin staining. Cell viability was assayed by lactate dehydrogenase and thiazolyl blue tetrazolium bromide methods and propidium iodide exclusion, gene expression by real-time PCR, and MMP release by zymography. Cell volume was monitored by electronic cell sorting. RESULTS: Hypotonicity (50%) and mechanical stretch increased ATP release with similar pharmacologic profiles. TM cells expressed ecto-ATPases E-NPP1-3, E-NTPD2, E-NTPD8, and CD73. Prolonged dexamethasone (DEX) exposure (≥ 2 weeks), but not brief exposure (3 days), increased cross-linked actin networks and reduced swelling-triggered ATP release. Cytochalasin D (CCD) exerted opposite effects. Neither DEX nor CCD altered the cell viability, gene expression, or pharmacologic profile of ATP-release pathways. DEX accelerated, and CCD slowed, the regulatory volume decrease after hypotonic exposure. Activating A(1) adenosine receptors (A(1)ARs) increased total MMP-2 and MMP-9 release. DEX reduced total A(1)AR-triggered MMP release, and CCD increased the active form of MMP-2 release. The A(1)AR agonist CHA and the A(1)AR antagonist DPCPX partially reversed the effects of DEX and CCD, respectively. CONCLUSIONS: Cytoskeletal restructuring modulated swelling-activated ATP release, in part by changing the duration of cell swelling after hypotonic challenge. Modifying ATP release is expected to modulate MMP secretion by altering ecto-enzymatic delivery of adenosine to A(1)ARs, linking cytoskeletal remodeling and MMP-mediated modulation of outflow facility.

Full Text

Duke Authors

Cited Authors

  • Li, A; Leung, CT; Peterson-Yantorno, K; Stamer, WD; Civan, MM

Published Date

  • October 10, 2011

Published In

Volume / Issue

  • 52 / 11

Start / End Page

  • 7996 - 8005

PubMed ID

  • 21896846

Pubmed Central ID

  • PMC3220413

Electronic International Standard Serial Number (EISSN)

  • 1552-5783

Digital Object Identifier (DOI)

  • 10.1167/iovs.11-8170


  • eng

Conference Location

  • United States