The role of N-acetylcysteine amide in defending primary human retinal pigment epithelial cells against tert-butyl hydroperoxide- induced oxidative stress

Abstract

INTRODUCTIONAge-related macular degeneration (AMD) accounts for 8.7% of blindness worldwide, and it is a leading cause of blindness in the United States among adults age 60 and older. [1][2][3] Globally, about 1 in 32 visual impairments and 1 in 15 cases of blindness are due to macular disease. 4 Dry AMD is a degenerative condition that begins in Bruch's membrane and progresses to the retinal pigment epithelium (RPE) and the overlying photoreceptors with loss of central vision. The RPE is a single layer of pigment cells lying between the photoreceptors and choriocapillaris. The functions of the RPE include light absorption, epithelial transport, phagocytosis, secretion, and immune modulation. All of these functions are important to maintaining and supporting the photoreceptors. High blood flow rate and high O 2 saturation in the choriocapillaris render the RPE more exposed to oxidative stress. 5Oxidative stress seems to play a major role in the pathogenesis of dry AMD and retinal degeneration, although the pathophysiology of dry AMD is complex. 5,6 The macula is subject to increased levels of photooxidative stress because of its elevated metabolic rate and high proportion of polyunsaturated fatty acids, which are highly susceptible to lipid peroxidation. As the RPE cells of the macula age, oxidation of lipids and other cellular components results in an accumulation of indigestible lipofuscin in the lysosomes. The accumulation of lipofuscin granules closely parallels drusen formation in time course and distribution in the retina. Oxidative damage to the mitochondrial DNA in RPE cells has also been implicated in the development of AMD. Levels of antioxidants in retinal tissue, in particular glutathione (GSH), decrease with aging, making the macula even more vulnerable to progressive oxidative damage. 7,8 Endogenous antioxidants, such as GSH, cannot be replaced directly. Instead, compounds that can easily enter cells and increase intracellular antioxidant levels are preferred. Dietary antioxidants have been shown to slow the progression of retinal degeneration and halt progression of AMD in human clinical trials.9,10 GSH and its amino acid precursors protect RPE cells from oxidative injury and oxidant-induced apoptosis in vitro. N-acetylcysteine amide (NACA), a GSH prodrug, is a promising candidate for use as an antioxidant-based treatment for AMD. It has demonstrated efficacy in numerous in vitro and animal models of other oxidative stress-related conditions, including drug-induced RPE damage and cataracts. [11][12][13][14][15] Our lab has demonstrated that NACA prevented retinal degeneration in vitro and in animal models.14 The immortalized ARPE-19 cell line was used as an in vitro model because primary human RPE cells were not available at the time. ROS generation and lipid peroxidation induced by tert-butyl hydroperoxide (TBHP) were reversed by pretreatment with NACA. NACA also protected against TBHP-induced cell damage by increasing cellular levels of GSH, and maintaining cellular integrity, as measured by tra...