Expression of the Optineurin Gene in Human Trabecular Meshwork Cells Following Oxidative Stress and Phagocytosis

Abstract

Abstract: : Purpose: To examine the influence of stress, such as oxidative stress and phagocytic events on the expression of optineurin gene in cultured human trabecular meshwork (TM) cells. Methods: Early passaged human TM cells were treated for 30 minutes with 1.0 mM hydrogen peroxide (H₂O₂) or were incubated for 40 hr with latex beads. Proteins in the cell lysates and total RNA were extracted 1, 24 and 48 hrs after H₂O₂ treatment, as well as at the 0, 24 and 48 hr post–phagocytosis time points. Western blotting was performed to examine the optineurin protein level. The transcript level was determined by and real time PCR. Results: After oxidative stress, optineurin protein was increased 4.7–fold at the time point of 1 hr. The protein level then dropped off to normal level at later time points. A transient 2.8–fold increase of optineurin was also observed immediately following phagocytosis (0 hr time point). The protein expression again returned to normal at 24 and 48 hr time points. By real time PCR, the change of optineurin transcript paralleled that of protein in both experiments. Conclusions: The expression of the optineurin gene was rapidly and temporarily stimulated in human TM cells by oxidative stress and phagocytic challenge. Optineurin may be a stress response protein.

Keywords: trabecular meshwork • gene/expression • stress response