RPE Express MT1-MMP and its Levels Are Regulated by Estrogen and Oxidant Injury

Abstract

Abstract: : Purpose: Age-related macular degeneration (ARMD) is the most important cause of lost central vision in the elderly. Oxidant injury to the RPE has been implicated as a mechanism for the development of subRPE deposits and drusen in ARMD. Our previous data indicated that RPE express high levels of proMMP-2 and active MMP-2, and that the levels are regulated by estrogen. Following RPE oxidant injury, active MMP-2 declines but proMMP-2 increases. Since MT1-MMP is one of the key cell surface enzymes involved in activation of proMMP-2, we investigated whether MT1-MMP was expressed in RPE cells, if it was regulated by estrogen and if the levels of expression changed after oxidant injury as a mechanism for changes in MMP-2 activation. Methods: Confluent cultures of the human RPE-19 cell line were incubated in phenol red free medium containing 0.1% fetal bovine serum with and without 17ß-estradiol treatment. ARPE-19 cells oxidant injured after transient exposure to H2O2 and MPO were collected for protein. 100ug of protein extract was immunoprecipitated and western analysis performed for MMP-2 and MT1-MMP. Zymography for MMP-2 was also performed. Results: Western blotting indicated that abundant MT1-MMP protein was expressed in RPE-19 cells and cell membranes. After oxidant injury, active MMP-2 expression decreased. In contrast, injury increased MT1-MMP. Interestingly, treatment with 17ß-estradiol prevented the injury induced increase in MT1-MMP expression. Conclusions: RPE express the cell surface protease MT1-MMP, and its levels are regulated by estrogen and oxidant injury. Changes in MT1-MMP protein expression may be associated with dysregulation activation of proMMP-2 and other matrix molecules following oxidant injury, and treatment with 17ß-estradiol may prevent this effect.

Keywords: age-related macular degeneration • oxidation/oxidative or free radical damage • retinal pigment epithelium