Journal: Redox Biology

Article Title: Homocysteine induced N 6 -methyldeoxyadenosine modification perturbation elicits mitochondria dysfunction contributes to the impairment of learning and memory ability caused by early life stress in rats

doi: 10.1016/j.redox.2025.103668

Figure Lengend Snippet: 6 mA accumulated in the mitochondria of the hippocampus in MS rats. A. The immunofluorescence staining in the hippocampal neurons of rats with RNase treatment (scale bar: 100 μm), 6 mA signals (red) and their co-localization with NeuN (green). B. The 6 mA signals (green) and their co-localization with mitochondria marker (red) in primary neuron cells of the hippocampus in rats with RNase treatment; scale bar: 30 μm. C–E. The expression of methylase METTL4 and demethylase ALKBH1 in the mitochondria was detected by western blotting under MS stress. F–G. The overall mtDNA 6 mA modification level of the hippocampus in rats under MS stress. H. The distribution of 6 mA signals in mtDNA mapped using MeDIP-Seq. The map revealed the 6 mA modification sites distributed in the promoter region of D-loop and in mtDNA-encoding gene regions. I–J. Methylation immunoprecipitation (6 mA-IP) qPCR was performed to validate 6 mA sites in the promoter region and mtDNA encoded genes under MS stress. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, n = 3.

Article Snippet: The brain slices were blocked with goat serum for 1–2 h at 4 °C and incubated with the primary antibodies for 24 h at 4 °C, namely the NeuN Monoclonal Mouse Antibody (1:200, 66836, Proteintech) and the Rabbit Polyclonal 6 mA Antibody (1:200, # 202003, Synaptic Systems).

Techniques: Immunofluorescence, Staining, Marker, Expressing, Western Blot, Modification, Methylated DNA Immunoprecipitation, Methylation, Immunoprecipitation