Modulation of RAS and PI3-AKT pathway by Stavudine (d4T) in PBMC of Alzheimer’s Disease Patients

Background: Aβ42-deposition plays a pivotal role in AD-pathogenesis by inducing the activation of microglial cells and neuroinflammation. This process is antagonized by microglia-mediated clearance of Aβ plaques. Activation of the NLRP3 inflammasome is involved in neuroinflammation and in the impairments of Aβ-plaques clearance. Stavudine (d4T) on the other hand down-regulates the NLRP3 inflammasome and stimulates autophagy-mediated Aβ-clearing in a TPH-1 cell line model.We explored the effect of d4T on Aβ-autophagy using PBMC of AD patients that were primed with LPS and stimulated with Aβ in the absence/presence of d4T. We analyzed the NLRP3 activity by measuring NLRP3-ASC complexes formation by AMNIS Flow-sight and pro-inflammatory cytokines (IL-1β, IL-18 and Caspase-1) production by ELISA. Western blot analyses were used to measure phosphorylation and protein expression of p38, CREB, ERK and AKT, p70, LAMP 2A, Beclin-1 and Bax.Results: Data showed that d4T: 1) down regulates NLRP3 inflammasome activation and the production of down-stream proinflammatory cytokines even in PBMC; 2) stimulates the phosphorylation of AKT, ERK, p70 as well as LAMP2A production, but does modulate beclin-1, suggesting a selective effect of this compound on chaperone-mediated autophagy (CMA); 3) up regulates p-CREB and BAX, possibly diminishing Aβ-mediated cytotoxicity; and 4) reduces the phosphorylation of p-38, a protein involved in the production of pro-inflammatory cytokines.Conclusions: d4T reduces the activation of the NLRP3 inflammasome and stimulates CMA autophagy as well as molecular mechanisms that modulate cytotoxicity and reduce inflammation in cells of AD patients. It might be interesting to verify the possibly beneficial effects of d4T in the clinical scenario.