Modulation of MAPK- and PI3/AKT-Dependent Autophagy Signaling by Stavudine (D4T) in PBMC of Alzheimer's Disease Patients

Background: A beta(42) 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 A beta plaques. Activation of the NLRP3 inflammasome is involved in neuroinflammation and in the impairments of A beta-plaque clearance. On the other hand, stavudine (D4T) downregulates the NLRP3 inflammasome and stimulates autophagy-mediated A beta-clearing in a THP-1-derived macrophages. Methods: We explored the effect of D4T on A beta autophagy in PBMC from AD patients that were primed with LPS and stimulated with A beta oligomers in the absence/presence of D4T. We analyzed the NLRP3 activity by measuring NLRP3-ASC complex formation by AMNIS FlowSight and pro-inflammatory cytokine (IL-1 beta, IL-18 and Caspase-1) production by ELISA. The phosphorylation status of p38, ERK, AKT, p70, and the protein expression of CREB, LAMP2A, beclin-1, Caspase-3 and Bcl2 were analyzed by Western blot. Results: Data showed that D4T: (1) downregulates NLRP3 inflammasome activation and the production of down-stream pro-inflammatory cytokines in PBMC; (2) stimulates the phosphorylation of AKT, ERK and p70 as well as LAMP2A, beclin-1 and Bcl2 expression and reduces Caspase-3 expression, suggesting an effect of this compound on autophagy; (3) increases phospho-CREB, which is a downstream target of p-ERK and p-AKT, inducing anti-inflammatory cytokine production and resulting in a possible decrease of A beta-mediated cytotoxicity; and (4) reduces the phosphorylation of p38, a protein involved in the production of pro-inflammatory cytokines and tau hyperphosphorylation. Conclusions: D4T reduces the activation of the NLRP3 inflammasome, and it might stimulate autophagy as well as the molecular mechanism that modulates A beta cytotoxicity, and D4T might reduce inflammation in the cells of AD patients. It could be very interesting to check the possible beneficial effects of D4T in the clinical scenario.