Characterization of proteins involved in disease progression in Alzheimer disease

Alzheimer disease (AD) is a neurodegenerative disease and severe neuronal loss is already taking place at the time of diagnosis. AD affects nearly 50 million people worldwide and the incidence is expected to rise significantly in the coming decades, making the disease a high priority healthcare concern. There are two main hallmarks of AD pathology, the intraneuronal tau tangles and the mostly extracellular amyloid plaques. Current treatment strategies for AD have tried targeting both tau and amyloid plaques although with limited success. However, since AD is a multifactorial disease with many risk factors, a multifaceted approach is necessary to tailor treatment. Further understanding of the pathogenesis of AD is required to enable the development of novel drugs and treatment strategies. In this thesis, we use proteomics, bioinformatics, and microscopy in an attempt to identify and characterize proteins which might influence AD progression and pathogenesis. We utilize the AD mouse model AppNL-F/NL-F, which is a knock-in mouse model that overproduces amyloid β leading to Aβ plaque pathology without artifacts associated with APP overexpression applied in previous AD mouse models. We also study AD brain and neuronal cultures obtained from the AppNL-F/NL-F mouse.

In Paper I, we show that a combination of proteomics and bioinformatics can identify proteins involved in AD pathology in the AppNL-F/NL-F mice. Using immunofluorescence, we discovered that huntingtin, the pathogenic protein in Huntington disease, is abundant in the hippocampus of the AppNL-F/NL-F mouse at a presymptomatic stage. We furthermore localized the expression of huntingtin to pyramidal neuronal cells early in the mouse life span. In Paper II, we expand on our findings of huntingtin in the AppNL-F/NL-F mouse model by studying and characterizing the expression of huntingtin in the brain of AD patients and healthy controls. We found that huntingtin was increased in the frontal cortex and the hippocampus of AD patients. Huntingtin could be found in pyramidal neurons within both the frontal cortex and the hippocampus. The accumulation pattern of huntingtin in AD did not mimic the accumulation found in Huntington disease as there was no correlation between astrocytes and huntingtin in AD brain. Furthermore, using confocal microscopy we concluded that there was no association of tau protein and huntingtin in AD brain. In paper III, we investigated the localization of the protein DDX24, a protein identified via proteomics, in AD brain and neuronal cultures derived from the AppNL-F/NL-F mouse. DDX24 belongs to a family of proteins consisting of putative RNA helicases and is implicated in translation initiation, nuclear RNA splicing and ribosome assembly. We show that DDX24 accumulates in AD brain where it associates to areas important for memory formation. We also show that DDX24 is increased in the brain of AD patients compared to healthy controls. We found DDX24 to be increased in the brain and in the neuronal cells of the AppNL-F/NL-F mouse. Decreasing DDX24 levels increased APP levels in neuronal cells. DDX24 levels also appeared to be regulated by amyloid load or vice versa. In paper IV, we investigate neuritogenesis and neurogenesis in AD brain and in the AppNL-F/NL-F mouse model. We show that Ankyrin-3, a protein important for axonal development, is detectable in AD brain and in neurons derived from the AppNL-F/NL-F mouse. We also show that a known biomarker, doublecortin, is increased in embryonic and young mice brain derived from the AppNL-F/NL-F mouse when compared to controls.

Overall, our studies use proteomics, bioinformatics, and microscopy to describe the presence of huntingtin, DDX24, Ankyrin-3 and doublecortin in the brain of AD patients and AppNL-F/NL-F mouse. These results can prove helpful in our future understanding of AD pathogenesis.

List of scientific papers

I. Proteomics Time-Course Study of App Knock-In Mice Reveals Novel Presymptomatic Aβ42-Induced Pathways to Alzheimer’s Disease Pathology. Schedin-Weiss S, Nilsson P, Sandebring-Matton A, Axenhus M, Sekiguchi M, Saito T, Winblad B, Saido T, Tjernberg L. Journal of Alzheimer’s Disease. 2020;75(1):312-335.
https://doi.org/10.3233/JAD-200028

II. Huntingtin Levels are Elevated in Hippocampal Post-Morten Samples of Alzheimer’s Disease Brain. Axenhus M, Winblad B, Tjernberg L, Schedin-Weiss S. Current Alzheimer Research. 2020;17(9):858-867.
https://doi.org/10.2174/1567205017666201203125622

III. DEAD Box Helicase 24 is Increased in Alzheimer Disease Brain and Influences Presymptomatic Pathology. Axenhus M, Doeswijk T, Nilsson P, Winblad B, Tjernberg L, Schedin- Weiss S. [Manuscript]

IV. Neuritogenesis and Neurogenesis in Alzheimer Disease Brain and the AppNLF/NLF Mouse model. Axenhus M, Malik C S, Doeswijk T, Nilsson P, Winblad B, Tjernberg L, Schedin-Weiss S. [Manuscript]