Effects of omega-3 fatty acid supplementation and retinal pigment epithelium on protein expression in cultured chick retinal neurons

It is well-established that the retinal pigment epithelium (RPE) plays an integral role in the function, nourishment, and survival of photoreceptors and other retinal neurons through multiple roles, including isomerization of retinal in the visual cycle and secretion of various growth factors and cell survival signals. Polyunsaturated fatty acids (PUFAs), such as docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), are selectively taken up from circulation, transported to retinal tissue through the RPE, and eventually incorporated into retinal cell membranes. PUFAs have been shown to protect the retina against oxidative damage resulting from pro-inflammatory and pro-apoptotic cell signals that are hallmarks of retinal degenerative diseases. While the positive effects of PUFA supplementation, especially DHA, on the antioxidant defense of retinal cells have been previously established, the objective of this study is to determine whether these neuroprotective effects are exerted on a local level using a DHA precursor. RPE and retinae were isolated and grown in vitro in the presence or absence of PUFAs. Embryonic day 18 chick embryos were sacrificed, and eyes were dissected to separate the RPE and retina into tissue explants to be grown as dissociated retinal cell cultures and dissociated retina and RPE co-cultures for 6 days. These cultures were supplemented with DHA or EPA and subsequently treated with hydrogen peroxide to simulate the oxidative stress conditions akin to those of retinal degenerative diseases. Immunocytochemical techniques in conjunction with confocal microscopy were utilized to visualize, quantify, and assess the expression of opsin and neuron-specific tubulin to investigate effects on rod photoreceptors as well as the neural cell population of the retina