Studio di modelli in vitro e in vivo di patologia oculare in risposta a stimolo neurotrofico: identificazione di nuovi biomarcatori e bersagli terapeutici

The retinal pigment epithelium (RPE), a key cell layer in the eye, plays an important role in retinal health. Dysfunction of the RPE is linked not only to age-related macular degeneration (AMD) but also to glaucoma, a major cause of blindness. RPE dysfunction contributes to the degeneration of retinal ganglion cells (RGCs), which is central to glaucoma-related vision loss. Oxidative stress and inflammation in RPE and Müller cells exacerbate glaucoma, and dysfunction of RPE can also disrupt the blood-retinal barrier, leading to the accumulation of toxic substances. Currently, neither biomarkers nor effective therapy are available for the disease. Nerve growth factor (NGF) has shown potential as a neuroprotective agent, but mechanisms of action remain unclear. In this study, after generating a differentiated RPE cell model from ARPE-19, which confirmed the expression of specific RPE markers (e.g. RPE65, CRALBP, MITF, RDH10), an in vitro model of glaucoma was set by using an original device able to induce pressure increase (HP) on cultured cells. The above-mentioned human RPE and rat Müller cells were used for experimental procedures. Several of the most important proteins involved in oxidative stress (e.g. DJ-1, NRF2, HO-1, CAT, SOD-1) and inflammatory (IL-6, IL-1beta) responses were analyzed in HP condition, in the presence or absence of NGF. At the same way, expression levels of three microRNAs (miR-34a-5p, miR-29c-3p, miR-146a-5p) were evaluated, and a direct target of miR-34a-5p (SIRT-1) was considered as well. Interestingly, results highlighted the suitability of the model for the study of glaucoma, evidencing, on one hand, expected and coherent levels of expression of the considered markers, and, on the other, the putative role of NGF as neuroprotective factor able to revert conditions caused by HP. Such effects were evidenced in terms of both oxidative stress/inflammation-related proteins’ and microRNAs’ expression levels in response to specific experimental conditions. In conclusion, the study provided interesting results and opens further analyses on preclinical models to assess the potential use of NGF for glaucoma treatment and validate candidate miRNAs here described as novel biomarkers/therapeutic targets. The industrial partner of the project, included in the PhD PON R&I program, was Dompé S.p.A.