Mutant APP and amyloid beta-induced defective autophagy, mitophagy, mitochondrial structural and functional changes and synaptic damage in hippocampal neurons from Alzheimer's disease.

The purpose of our study was to determine the toxic effects of hippocampal mutant APP (mAPP) and amyloid beta (Aβ) in human mAPP complementary DNA (cDNA) transfected with primary mouse hippocampal neurons (HT22). Hippocampal tissues are the best source of studying learning and memory functions in patients with Alzheimer's disease (AD) and healthy controls. However, investigating immortalized hippocampal neurons that express AD proteins provide an excellent opportunity for drug testing. Using quantitative reverse transcriptase-polymerase chain reaction, immunoblotting & immunofluorescence and transmission electron microscopy, we assessed messenger RNA (mRNA) and protein levels of synaptic, autophagy, mitophagy, mitochondrial dynamics, biogenesis, dendritic protein MAP2 and assessed mitochondrial number and length in mAPP-HT22 cells that express Swedish/Indiana mutations. Mitochondrial function was assessed by measuring the levels of hydrogen peroxide, lipid peroxidation, cytochrome c oxidase activity and mitochondrial adenosine triphosphate. Increased levels of mRNA and protein levels of mitochondrial fission genes, Drp1 and Fis1 and decreased levels fusion (Mfn1, Mfn2 and Opa1) biogenesis (PGC1α, NRF1, NRF2 & TFAM), autophagy (ATG5 & LC3BI, LC3BII), mitophagy (PINK1 & TERT, BCL2 & BNIPBL), synaptic (synaptophysin & PSD95) and dendritic (MAP2) genes were found in mAPP-HT22 cells relative to WT-HT22 cells. Cell survival was significantly reduced mAPP-HT22 cells. GTPase-Drp1 enzymatic activity was increased in mAPP-HT22 cells. Transmission electron microscopy revealed significantly increased mitochondrial numbers and reduced mitochondrial length in mAPP-HT22 cells. These findings suggest that hippocampal accumulation of mAPP and Aβ is responsible for abnormal mitochondrial dynamics and defective biogenesis, reduced MAP2, autophagy, mitophagy and synaptic proteins & reduced dendritic spines and mitochondrial structural and functional changes in mAPP hippocampal cells. These observations strongly suggest that accumulation of mAPP and Aβ causes mitochondrial, synaptic and autophagy/mitophagy abnormalities in hippocampal neurons, leading to neuronal dysfunction.