Neuroprotective Effects Of(Val~8)GLP-1-Glu-PLA Against Rotenone-induced Cytotoxicity On Hippocampal And Cortical Neurons

Background:Recent studies have shown that glucagon-like peptide-1(GLP-1) possesses beneficial effects on brain function and have neuroprotective effects in vivo and in vitro However, it is ineffective as a therapeutic agent due to its rapid degradation by dipeptidylpeptidase-IV (DPP-IV) and elimination from the body.(Val8)GLP-1-Glu-PAL, a novel form of GLP-1, binds to the GLP-1receptor, enables longer duration of action and has greater bioavailability.Objective:The aim of this study is to investigate the protective effects of (Val8)GLP-1-Glu-PAL on primary cultured hippocampal and cortical neurons.Methods:Primary hippocampal and cortical neurons were exposed to rotenone for48h in different concentrations (0,0.5,1,3, and5nM respectively) after a70-80%confluency for7-day in vitro. Neuronal cell viability was evaluated by Cell Counting Kit-8(CCK-8) assay. To decide whether (Val8)GLP-1-Glu-PAL had side effects, neuron toxicity were assessed by estimating lactate dehydrogenase (LDH) release. After that, neurons were pre-treated with different concentrations of (Val8)GLP-1-Glu-PAL(10,50,100,200and500nM) and then added with rotenone (0.5nM)2h later. CCK-8and LDH assay were used to evaluate the protective effects of (Val)GLP-1-Glu-PAL after7d of culture. The expression of three apoptotic factors; procaspase-3, cleaved caspase-3and Bcl-2were assessed using the Western blot technique.Results:Rotenone for increasing concentrations (0.5,1,3, and5nM) decreased cell viability after48h treatment in a dose dependent fashion (p<0.05). Different concentrations of (Val8)GLP-1-Glu-PAL(10,50,100,200and500nM) alone didn’t significantly change cell viability of neurons compared with control group. This suggested that (Val)GLP-1-Glu-PAL had no side effects on neurons. CCK-8assay demonstrated that the number of viable neurons was increased by treating the cells with (Val8)GLP-1-Glu-PAL in the presence of rotenone(0.5nM).(Val)GLP-1-Glu-PAL decreased the release of LDH from the dying neurons,(p<0.05) suggesting the neuroprotective effects of (Val8)GLP-1-Glu-PAL. These results suggested that (Val)GLP-1-Glu-PAL had protective effects against rotenone-induced cytotoxicity on hippocampal and cortical neurons. Western blot demonstrated that procaspase-3was decreased and cleaved caspase-3was increased significantly48h after rotenone, and the effects were reversed by pre-treatment with (Val8)GLP-1-Glu-PAL(100nM). Anti-apoptotic factor Bcl-2was down-regulated following rotenone treatment while (Val8)GLP-1-Glu-PAL reversed this down regulation.Conclusion:Our study demonstrated a neuroprotective effect of (Val8)GLP-1-Glu-PAL against rotenone-induced cytotoxicity.(Val8)GLP-1-Glu-PAL could remarkably prevent from the decrease of cell viability and mediate anti-apoptotic actions in neurons. Since (Val)GLP-1-Glu-PAL is a potent, long-acting GLP-1peptide and readily enters the brain through the blood brain barrier (BBB), it may be a potential novel agent to treat neurodegenerative disorders such as Alzheimer’s Disease and Parkinson Disease.