Alzheimer Hastalığı Tedavisinde Kullanılmak Üzere Donepezil Yüklü Plga-B-Peg Nanopartiküllerinin Hazırlanması ve Beyne Hedeflendirilmesi

Abstract

Alzheimer's disease (AD) is an irreversible and progressive neurodegenerative disease, which is caused by irreversible loss of neurons in the hippocampus and cortex regions of the brain. Although the molecular mechanism of the disease is still unclear, the deposition of the amyloid beta (Aβ) proteins (senile plaques) in the extracellular synaptic spaces of the neocortex is suggested to play a major role in progress of AD. Cholinesterase inhibitors are compounds that are used for symptomatic treatment of neuron transmission improvement in AD. Donepezil is a reversible and non-competitive cholinesterase inhibitor that preferably inhibits acetylcholinesterase compared to butyrylcholinesterase and is clinically used for treatment of AD. In the frame of this thesis, we developed donepezil loaded poli (laktic-co-glycolic acid)-block-poli (ethylen glycol) [PLGA-b-PEG] nanoparticles to increase efficiency of treatment in AD and to reduced the side effects of donepezil nanoparticles were directly targeted to the Aβ fibriles. By targeting donepezil to the brain in sufficient concentration we destabilized Aβ fibrile formation and minimized its tissue distrubition. Furthermore by setting up a blood brain barrier model in vitro, using microvascular endothelial cells and astrocytes we determined that this prepared nanoparticles passed across the blood brain barrier. In this set up we also evaluated the protective effect of donepezil on astrocytes in neuroinflammation by assessing cytokine expressions both in protein and gene levels. According to these results for inflammation cytokines IL-1β, IL-6, GM-CSF, TGF-β, MCP-1 and TNF-α, increase in both expression and protein levels was detected on astrocytes which were incubated with amyloid fibrils. After free donepezil and donepezil loaded nanoparticle administration to these stimulated astrocytes a significant decreased in both gene expression and protein levels were detected for IL-1β, IL-6, GM-CSF and TNF-α, whereas there were no significant changes observed for TGF-β and MCP-1. In vivo Alzheimer model was created using intracerebroventricular injection of Aβ(25-35) peptide. After the treatment period brain tissues of rats were removed and homogenized. Following the homogenization of the brain tissues; acetylcholinesterase activities were determined in each sample. According to the results acetylcholinesterase activity of Alzheimer model rats achieved with icv Aβ(25-35) injection were found to be significantly increased. Effect of donepezil loaded nanoparticles in this animal model were found to be more effective than free donepezil and significantly inhibited the acetylcholinesterase activity. Based on these results, ability of donepezil loaded nanoparticles to inhibit acetylcholinesterase activity is stronger than high dose free donepezil. The reason for this can be attributed to the controlled release of donepezil, longer duration of action and successful targetting of nanoparticles.