Anti-oxidative stress and cognitive improvement of a semi-synthetic isoorientin-based GSK-3β inhibitor in rat pheochromocytoma cell PC12 and scopolamine-induced AD model mice via AKT/GSK-3β/Nrf2 pathway

Background: Alzheimer's disease (AD) is a neurodegenerative disease characterized by progressive cognitive deficits. Although the pathogenesis of AD is unclear, oxidative stress has been implicated to play a dominant role in its development. The flavonoid isoorientin (ISO) and its synthetic derivatives TFGF-18 selectively inhibit glycogen synthase kinase-3(3 (GSK-3(3), a potential target of AD treatment. Purpose: To investigate the neuroprotective effect of TFGF-18 against oxidative stress via the GSK-3(3 pathway in hydrogen peroxide (H2O2)-induced rat pheochromocytoma PC12 cells in vitro and scopolamine (SCOP)-induced AD mice in vivo. Method: The oxidative stress of PC12 cells was induced by H2O2 (600 mu M) and the effects of TFGF-18 (2 and 8 mu M) or ISO (12.5 and 50 mu M) were observed. The AD mouse model was induced by SCOP (3 mg/kg), and the effects of TFGF-18 (2 and 8 mg/kg), ISO (50 mg/kg), and donepezil (DNP) (3 mg/kg) were observed. DNP, a currently accepted drug for AD was used as a positive control. The neuronal cell damages were analyzed by flow cytometry, LDH assay, JC-1 assay and Nissl staining. The oxidative stress was evaluated by the detection of MDA, SOD, GPx and ROS. The level of ACh, and the activity of AChE, ChAT were detected by the assay kit. The expressions of Bax, Bcl-2, caspase3, cleaved-caspase3, p-AKT (Thr308), AKT, p-GSK-3(3 (Ser9), GSK-3(3, Nrf2, and HO-1, as well as p-CREB (Ser133), CREB, and BDNF were analyzed by western blotting. Morris water maze test was performed to analyze learning and memory ability. Results: TFGF-18 inhibited neuronal damage and the expressions of Bax, caspase3 and cleaved-caspase3, and increased the expression of Bcl-2 in vitro and in vivo. The level of MDA and ROS were decreased while the activities of SOD and GPx were increased by TFGF-18. Moreover, TFGF-18 increased the p-AKT, p-GSK-3(3 (Ser9), Nrf2, HO-1, p-CREB, and BDNF expression reduced by H2O2 and SCOP. Meanwhile, MK2206, an AKT inhibitor, reversed the effect of TFGF-18 on the AKT/GSK-3(3 pathway. In addition, the cholinergic system (ACh, ChAT, and AChE) disorders were retrained and the learning and memory impairments were prevented by TFGF-18 in SCOPinduced AD mice. Conclusions: TFGF-18 protects against neuronal cell damage and cognitive impairment by inhibiting oxidative stress via AKT/GSK-3(3/Nrf2 pathway.