Increased DNA modification in the Alzheimer’s disease human brain

Abstract

Described for the first time in 1906, Alzheimer’s disease (AD) remains the most prevalent form of dementia throughout the world. AD is a complex neurodegenerative disease characterised by the progressive decline in cognitive functions, neuronal cell loss, and by the presence of amyloid beta (Aβ) plaques and neurofibrillary tangles that are composed of hyperphosphorylated tau, in the brain. Only a small percentage of the AD cases can be attributed to mutations in key genes that are involved in the pathophysiology of the disease. The vast majority of AD cases are sporadic, and the aetiology of these cases remains unknown. However, there is evidence that an alteration in epigenetics is involved in the pathophysiology of AD. Amongst the epigenetic markers, 5- methylcytosine (5mC) has been the most studied in AD. Previous human studies reported both increases and decreases in the level of 5mC at specific loci of genes that are involved in the pathophysiology of AD. However, only a few studies have investigated 5mC at global levels, and reported DNA hypomethylation in the AD brain. 5-hydroxymethylcytosine (5hmC), the oxidised form of 5mC, has been recently recognised as an epigenetic marker. The involvement of 5hmC in AD has not been investigated yet. In the present study, specific antibodies were used to study 5mC and 5hmC at global levels in two brain regions that are vulnerable to AD: the middle frontal gyrus and the middle temporal gyrus. Immunohistochemistry combined with high-throughput image analysis was used as the method to quantify levels of 5mC and 5hmC in post-mortem human brain tissue. The results demonstrated that DNA was globally hypermethylated and hyperhydroxymethylated in neurons from the AD brain. Increased levels of 5mC and 5hmC were significantly correlated with Aβ, tau, and ubiquitin loads; thus, demonstrating that these epigenetic changes were related to the progression of the disease. Dnmt1 and Dnmt3b, two of the enzymes involved in the regulation of 5mC were studied using the same approach as for 5mC and 5hmC. Results demonstrated that Dnmt1 was increased in the AD MTG, whereas Dnmt3b was decreased in the AD MTG. In vitro models of human brain cells were validated for the study of 5mC and 5hmC, and were used to test the effect of Aβ and 5-aza-2’-deoxycytidine (AzaC), a well-known demethylating agent, on levels of 5mC and 5hmC. Results demonstrated that Aβ can induce DNA hypermethylation and hyperhydroxymethylation in human precursor cells, but not in neurons and astrocytes differentiated from them. Finally, this study provided evidence that in addition to its effect on 5mC, AzaC is a strong inducer of 5hmC. Altogether, the results obtained in the present study clearly demonstrated the involvement of 5mC and 5hmC in AD, and provide the basis for the development of new diagnostic methods and therapeutic approaches.