A study on phenotypic and transcriptomic response to exercise in Alzheimer’s disease mice expressing mutant (N141I) presenilin-2

다국어 초록 (Multilingual Abstract)

Physical exercise is believed to produce beneficial effects on pathological features and behavioral symptoms of Alzheimer’s disease (AD). However, the molecular mechanisms by which exercise reduces pathophysiology of AD are not clearly understood. In the present study, it was examined whether the regular moderate exercise can improve cognitive function and produce transcriptomic response in the brain. To address this, four groups of mice were studied, non-transgenic control (Non-Tg), mice expressing the human presenilin-2 wild type (Tg-PS2w), mice expressing the human presenilin-2 with N141I mutation (Tg-PS2m), and mice expressing the human presenilin-2 with N141I mutation that were subjected to treadmill exercise (TE) treated at a speed of 10m/min for 50 min/day, 5 days/week, for 6 weeks (Tg-PS2m/Ex). Tg-PS2m/Ex mice exhibited increased preference in exploring a novel object compared to Tg-PS2m in the novel object recognition test, whereas effects of exercise on cognitive deficits of Tg-PS2m in the water maze test and passive avoidance test were subtle or undetected. Western blot and histological analysis using amyloid oligomer (A11) and Aβ (6E10) antibody indicated that amyloid oligomer-reactive bands and plaque deposition in the hippocampus tended to be reduced after TE treatment, although a statistical significance in both analyses was not obtained. Transcriptomic (RNA-sequencing) analysis of gene expression profiles and following protein analysis revealed that the cell cycle regulatory gene, Cdc28 protein kinase regulatory subunit 2 (Cks2), was decreased, and the cell cycle- and apoptotic cell death-related factors, including cyclin D1, PCNA, and cleaved caspase-3 were increased in the hippocampus of Tg-PS2m, whereas TE treatment reversed their altered expression. These results suggest that the regular moderate exercise for 6 weeks is not sufficient to clearly remove Aβ accumulation in the brain, but it improves cognitive deficits in the novel object recognition task and produces significant effects to reverse the transcriptomic responses driving cell-death in the hippocampus of Tg-PS2m mice. The results of the present study support the hypothesis that regular excise produces beneficial effects on pathological features and behavioral symptoms of AD through activation of transcriptomic responses in the hippocampus.

국문 초록 (Abstract)

이 연구는 운동이 Presenilin-2 돌연변이 유전자를 발현하는 알츠하이머 형질전환 마우스의 주요 병리지표와 전사체 발현에 미치는 영향에 대해 알아보고자 하였다. 연구목적을 달성하기 위해 모든 마우스는 Non-Tg (Non-transgenic control), Tg-PS2w (Transgenic mice expressing presenilin-2 wild-type), Tg-PS2m (Transgenic mice expressing mutated (N141I) presenilin-2), Tg-PS2m/Ex (Transgenic mice expressing mutated (N141I) presenilin-2 that were subjected to exercise) 집단으로 구분하였으며, 운동집단은 6주간 트레드밀 운동을 실시하였다. 그 결과, 수중미로검사(WMT)와 신물질탐색검사(NORT) 통해 Tg-PS2m 집단과 비교하여 Tg-PS2m/Ex 집단의 인지기능 수준이 개선된 것을 확인하였으며, Tg-PS2m 집단에서 해마(Hippocampus) 의존적으로 증가된 아밀로이드 베타(Amyloid beta; Aβ) 병변이 운동에 의해 감소하는 경향을 나타내는 것을 확인하였다. 또한, RNA-sequencing 기반 전사체 발현 분석과 단백질 정량분석을 통해 Tg-PS2m 집단에서 증가된 세포주기(Cell cycle) 및 세포사멸(Apoptosis) 관련 인자(Cyclin D1, PCNA, Bax, Cleaved caspase-3)의 발현이 운동에 의해 감소하는 것을 확인하였으며, 이를 매개하는 잠재적인 유전인자로서 Cdc28 protein kinaseregulatory subunit 2 (Cks2)의 변화를 관찰하였다. 결론적으로 6주간의 운동은 뇌에서 Aβ 병변을 제거하기에 충분하진 않지만, Tg-PS2m 알츠하이머 동물모델에서 해마 의존적으로 신경세포 사멸을 유도하는 유전적 반응과 인지기능 결함에 대한 개선효과를 나타냈다. 따라서, 운동은 알츠하이머 질환의 예방 및 치료를 위한 중재전략으로 활용 가능할 것으로 생각된다.

목차 (Table of Contents)

• Contents
• Ⅰ. Introduction 1
• 1. The background and objective of the present study 1
• 2. Objective of the thesis 5
• Ⅱ. Materials and Methods 6
• 1. Animals 6
• 2. Exercise protocol 6
• 3. Behavior assessment 7
• 1) Water maze test7
• 2) Novel object recognition test 7
• 3) Passive avoidance test 8
• 4. Tissue processing 8
• 5. RNA isolation 9
• 6. RNA sequencing 9
• 7. Bioinformatics analysis 9
• 8. Enzyme-linked immunosorbent assay 10
• 9. Immunoblotting 11
• 10. TUNEL assay 11
• 11. Immunofluorescence staining 12
• 12. Statistics 13
• Ⅲ. Result14
• Ⅳ. Discussion36
• Reference 41
• 국문초록 47