Abstract
Abstract: :
Purpose: Studies were conducted to determine the roles of Fak and Src in the transcriptional pathway leading to increased production of matrix metalloproteinase 2 (MMP–2), myocilin, and fibronectin in trabecular meshwork cells subjected to mechanical stretch. Methods: Porcine trabecular meshwork cells were subjected to mechanical stretch. Changes in the overall levels of MMP–2, fibronectin, Fak, and Src were analyzed using zymograms or western immunoblots. Phosphorylation of Fak and Src was then determined. Inhibitors, such as PP–2, known to affect Fak and Src, were added prior to and during stretch and their effects were ascertained. Confocal microscopy was used to evaluate Fak localization following stretch. Results: Mechanical stretch causes an increase in the production of MMP–2, myocilin and fibronectin but does not change the overall protein levels of Fak or Src. Stretch increases phosphorylation of Fak on Y396, Y861, and Y925 as well as on Y418 of Src. Confocal microscopy shows a rapid increase of Fak in the nucleus following stretch. The Src inhibitor, PP–2, changes phosphorylation patterns. The fibronectin increase following stretch is blocked by PP–2, but the myocilin and MMP–2 increases are unaffected. Conclusions: Trabecular meshwork cells respond to mechanical stretch by increasing fibronectin, myocilin and MMP–2 levels, in addition to increasing Fak phosphorylation. The data suggest that both Src–dependent and Src–independent signal transduction pathways are involved.
Keywords: trabecular meshwork • signal transduction • phosphorylation