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Fig. 3 | Molecular Neurodegeneration

Fig. 3

From: METTL3-dependent RNA m6A dysregulation contributes to neurodegeneration in Alzheimer’s disease through aberrant cell cycle events

Fig. 3

Decreased neuronal METTL3 and m6A in the hippocampus by AAV-shMettl3. (A) Highly conserved gene sequences of shRNA targeting region in mouse and rat Mettl3 mRNA. (B) Knockdown effect of shMettl3 was confirmed in mouse N2a cells 3 days after transient transfection (Data are means±SEM from 3 independent experiments). (C) AAV-mediated gene delivery in the hippocampus was confirmed by immunostaining for GFP. WT C57BL/6 mice receiving bilaterally stereotaxic injections of AAV encoding for eGFP plus anti-Mettl3 shRNA (shMettl3) or scrambled control shRNA (shCtrl) into the hippocampus at 2 months of age were analyzed 1 month later. (D-F) Representative images of the AAV-infected areas (GFP-positive) in hippocampus for different cell markers by immunocytochemistry after fixation: MAP 2 for neurons (D), GFAP for astrocytes (E), and Iba1 for microglia (F). (G, H) Representative image of immunoreactivity of METTL3 or m6A (G) in AAV-shRNA-injected mice and quantification analysis (H) confirmed that METTL3 protein and m6A modification level were reduced in mice injected with shMettl3 virus (n = 5–7 mice per group). GFP staining has been performed in adjacent brain slices at the same time and immunoreactivities of m6A and METTL3 were measured in GFP-positive regions. (Data are means±SEM, *p < 0.05, **p < 0.01, (B, H) unpaired student’s t-test)

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