Western blot

20 mg of mouse FC or HIP and human postmortem HIP were resuspended in Laemmli buffer (2% SDS, 10% glycerol, 60 mM Tris-Cl [pH 6.8], and 0.01% bromophenol blue), sonicated, and boiled for 5 min at 95°C. The sample concentration was determined by quantifying the absorbance at 260 nm with a NanoDrop One/OneC Microvolume UV spectrophotometer and using the equivalency between DNA and histone quantity (6 units A_260nm [DNA] = 1 μg/μL of protein). Proteins were transferred to a 0.2-μm nitrocellulose membrane (10600001, GE Healthcare, Chicago, IL, USA) and incubated overnight at 4°C with a primary antibody diluted in 5% skim milk in PBS containing 0.1% Tween 20 (663684B, Atlas Chemical, Houston, TX, USA). After three washes with PBS containing 0.1% Tween 20, membranes were incubated for 1 h at RT in a benchtop shaker with the secondary antibodies conjugated to horseradish peroxidase anti-rabbit IgG (1:10,000, A0545; Sigma-Aldrich, St. Louis, MO, USA) or anti-mouse IgG (1:5,000; NA9310, GE Healthcare, Chicago, IL, USA). Enhanced chemiluminescence (ECL) reagents (Luminata-HRT; Merck Millipore, Burlington, MA, USA) were used to visualize the proteins. Films were scanned, and the band analysis tool of the Quantity One software application was used for background subtraction and to determine the density of the bands. The proteins detected were MeCP2 (1:1,000; ab2829, Abcam, Cambridge, UK), SIRT2 (1:2,000; ab211033, Abcam, Cambridge, UK), acetyl-α-tubulin (1:2,000; T6793, Sigma-Aldrich, St. Louis, MO, USA), α-tubulin horseradish peroxidase (HRP) (1:5,000; ab40742, Abcam, Cambridge, UK), β-actin HRP (1:30,000; a3854, Sigma-Aldrich, St. Louis, MO, USA), TAU (1:1,000; ab64193, Abcam, Cambridge, UK), MAP6/STOP (1:1,000; 4265, Cell Signal, Danvers, MA, USA), STMN1 (1:500; 3352, Cell Signal, Danvers, MA, USA), S100b (1:1,000; Z0311, Dako, Carpinteria, CA, USA), GluR-1 (1:1,000; ab109450, Abcam, Cambridge, UK), GluR-2 (1:1,000, 13607, Cell Signal, Danvers, MA, USA), GluR-3 (1:2,000; 4676, Cell Signal, Danvers, MA, USA), acetyl-lysine (1:1,000; ab80178, Abcam, Cambridge, UK), ADAR2 (1:400; HPA018277, Atlas Antibodies, Bromma, Sweden), SOX2 (1:1,000; 3579, Cell Signal, Danvers, MA, USA), and Ki67 (1:1,000; ab16667, Abcam, Cambridge, UK).