Effects of an environmental enrichment protocol on epigenetic modifications in a rat transgenic model of Alzheimer’s disease-like amyloid pathology

CAMPUZANO, KAREN; DO CARMO, SONIA; DALMASSO, MARÍA CAROLINA; CASTAÑO, EDUARDO M; CUELLO, AUGUSTO CLAUDIO; MORELLI, LAURA; GALEANO, PABLO

Mendoza

Congreso; LVIII Reunión Anual de la Sociedad Argentina de Investigaciónes en Bioquímica y Biología Molecular; 2022

Sociedad Argentina de Investigaciónes en Bioquímica y Biología Molecular

Alzheimers disease is characterized by massive accumulation of amyloid beta in the cortex and hippocampus. Previous results in a transgenic Tg mice model of AD showed in hippocampus and cortex a decreased global DNA methylation levels together with a hypomethylation in the promotor of a critical enzyme for Abeta generation, BACE-1. Moreover, in human brains greater methylation in the BACE-1 promoter was associated with lower beta amyloid load in subjects with AD dementia, suggesting that epigenetic factors play a relevant role in AD amyloid pathology. Recently, it was shown that hydroxymethylation of cytosine residues, 5hmC, is much more abundant in the brain than in other tissues. However, this epigenetic mark has been little studied in AD brains. The aim of the present work was to determine the patterns of histone H3 methylation and acetylation, and DNA hydroxymethylation in the hippocampus of 9 month old Tg rats that recapitulate early stages of the AD like amyloid pathology, with intraneuronal Abeta accumulation in the absence of amyloid plaques, pre-plaque stage. Our results showed that there were not differences neither in the histone H3 methylation nor in the histone H3 acetylation levels between WT and Tg rats. Furthermore, given that epigenetic factors can be modified by environmental stimulation, we aimed to study whether an environmental enrichment protocol can reverse cognitive impairment and DNA hydroxymethylation patterns in Tg rats. To this end, WT and Tg rats were raised in standard cages and from 6 to 9 months of age subgroups of both genotypes were exposed to bigger cages (100 x 50 x 50 cm) that provided an environment of greater motor, cognitive, and social stimulation (EE). At approximately 8 months of age, spatial memory was assessed using the Morris Water Maze test. Then, rats were sacrificed and hippocampi were dissected and stored at -80 degree until further processing. Hippocampal DNA were isolated and sonicated, in an ultrasonic bath sonicator, in approximately 200-700 bp fragments. The size of the fragments was verified in a 2 per cent agarose gel and concentration and quality of DNA was measured in a NanoDrop spectrophotometer. Next, fragments were immunoprecipitated in order to obtain DNA fragments enriched in 5hmC and RT-qPCR were performed to measure the expression of Bdnf transcripts from promoters I and IV. Results showed that spatial memory was impaired in Tg rats compared to WT rats, and that the EE protocol was able to prevent this cognitive decline. Real Time qPCR for BDNF promoters I and IV were fine-tuned in control samples that were not immunoprecipitated and in the samples that were immunoprecipitated. The main objective of the present work is to perform next-generation sequencing (NGS) in order to identify differentially hydroxymethylated regions between the different treatments (genotypes and environmental protocols). This study was supported by PICT-2019-2019-00940 (ANPCyT) to PG and MCD.