CONICET | Buscador de Institutos y Recursos Humanos

Diabetic retinopathy (DR) is associated with persistent inflammation and with damage to the vascular bed. Previous results obtained by our group show that Alpha-1 Antitrypsin (A1AT) acts like an anti-inflammatory agent that could play a role on DR treatment. However, it is important to know the effect of A1AT on proteins that are relevant to retinal function and the molecular mechanisms involved. The retinal pigment epithelium (RPE) forms the outer component of the blood-retinal barrier. Connexin43 is a major gap junction protein expressed in RPE cells. Cx43 upregulation have been implicated in edema and loss of vascular integrity, leading to neuronal death, as in exudative age-related macular degeneration. AS160 is involved in insulin signaling affecting GLUT proteins. We evaluated Cx43, NFkB and AS160 expression and proteins implicated in different signaling pathways in an in vitro diabetic retinopathy cell model. ARPE-19 cells (ATCC® CRL-2302TM, USA) were maintained in DMEM/F12 (Invitrogen, USA) containing 2μM L-glutamine, 100U/ml penicillin, 100μg/ml streptomycin, and 10% fetal bovine serum. ARPE-19 cells (passages 9-12) were incubated 16hs with DMEM 5.5mM glucose (Control), DMEM 5.5mM glucose + 4.5mg/ml A1AT (Control + A1AT), DMEM 30mM glucose (Diabetic), DMEM 30mM glucose + 4.5mg/ml A1AT (Diabetic + A1AT). Cells were harvested with RIPA for Western blot or fixed for immunohistochemistry. A1AT diminished levels of Cx43 and AS160, A1AT also reduces AKT and pAKT1/2/3 expression levels, indicating PI3K/AKT pathway participation, and a possible crosstalk with Wnt and Insulin signaling. Besides, we could also observe a lower expression of NFkB p65 and iNOS, both proteins involved in the inflammatory response.Results support the hypothesis that A1AT regulates Cx43 and AS160 expression through PI3K/AKT, Wnt and Insulin signaling pathway. Taking together, these results indicate that A1AT is a promising molecule to be tested in retinal diseases as DR and age