In this study, radical scavenging effect and protective activity against oxidative stress of the extract and fractions from Acer okamotoanum (A. okamotoanum) were investigated. In addition, under Aβ25-35-induced Alzheimer''s disease (AD) model, the protective role from AD of A. okamotoanum was studies. A. okamotoanum was extracted with methanol (MeOH) and then fractionated in to 4 fractions including n-hexane, methylene chloride, ethyl acetate (EtOAc) and n-BuOH fractions.
The MeOH extract and fractions from A. okamotoanum showed strong radical scavenging effects against superoxide, hydroxyl radical, 2,2-diphenyl-1-picrylhydrazyl and nitric oxide. Among the MeOH extract and fractions, EtOAc fraction showed the strongest radical scavenging activities. In addition, EtOAc fraction showed the highest total flavonoid and phenol contents.
I investigated the protective role of MeOH extract and fractions against oxidative stress-induced neuronal cell damage using C6 glial cells. The treatment of hydrogen peroxide (H2O2) to C6 glial cells showed a loss of cell viability and an elevation of reactive oxygen species (ROS) level. However, the treatment of MeOH extract and fractions increased cell viability and decreased the ROS level. In addition, the protective mechanism of the active fraction, EtOAc fraction, against inflammation and apoptosis was studied. Control group treated with H2O2 significantly increased the up-regulation of inflammation factors, such as inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), interleukin-1 beta (IL-1β). However, treatment of EtOAc fraction effectively down-regulated iNOS, COX-2 and IL-1β. Furthermore, the increases of caspase 9, caspase 3, and PARP were significantly inhibited in dose-dependently manner by the treatment of EtOAc fraction.
The C6 glial cells showed a loss of cell viability and an elevation of ROS formation by treatment with Aβ25-35. However, treatment of MeOH extract and fractions increased cell viability and inhibited ROS formation. In addition, I investigated both inflammation and apoptosis related proteins in Aβ25-35-treated in C6 glial cells. The control group exhibited up-regulation of iNOS, COX-2, IL-1β, caspase 9, caspase 3 and PARP in C6 glial cells. However, EtOAc fraction treatment reduced the inflammation and apoptosis protein expression. Also, Aβ25-35 up-regulated the AD related proteins expression, amyloid precursor protein (APP), CTF-β, BACE, p-Tau. EtOAc fraction treatment inhibited expression of APP, CTF-β, BACE, p-Tau proteins in C6 glial cells. It indicated that EtOAc fraction exerted the protective effect from neuronal oxidative stress through the regulation of inflammation-and apoptosis-related protein expression.
I also studied the effect of EtOAc fraction on cognitive improvement and protective abilities in Aβ25-35-injected AD mouse model. EtOAc fraction was administered at an oral dose of 100 or 200 mg/kg/day for 2 weeks. The protective effect of EtOAc fraction from AD was observed using the behavioral tests including T-maze, object recognition, and Morris water maze tests. The control group injected with Aβ25-35 demonstrated significant impairments in recognition and memory function. However, the administration of EtOAc fraction showed improvement of cognition and memory function. Furthermore, administration of EtOAc fraction protected from Aβ25-35-induced lipid peroxidation and inhibited nitric oxide production in tissues. Acetylcholinesterase (AChE) was elevated in the brain by Aβ25-35, whereas administration of EtOAc fraction significantly decreased AChE level. In addition, I also investigated the protein expression of inflammation, apoptosis and AD related proteins using brain tissue of AD model mouse. Administration of EtOAc fraction groups significantly inhibited the expression of inflammation, apoptosis and AD related proteins.
In conclusion, the present study suggests that MeOH extract and fractions from A. okamotoanum would have the protective effect from oxidative stress induced by H2O2 and Aβ25-35 under C6 glial cells. In addition, the protective role of the EtOAc fraction against impairments of memory and cognition ability induced by Aβ25-35 under AD model of ICR mouse was also confirmed and it was related to the regulation of inflammation-and apoptosis-relation protein expression.