Journal: Nature Communications

Article Title: HDAC1 modulates OGG1-initiated oxidative DNA damage repair in the aging brain and Alzheimer’s disease

doi: 10.1038/s41467-020-16361-y

Figure Lengend Snippet: a Representative images of control and Hdac1 cKO mice at the age of 3 months (3M) and 13 months (13M) stained for GFAP (red) and nuclei Hoechst (blue) in the hippocampal region. b Analysis of GFAP + cells. a , b Three mice were analyzed in each group (two sections/mouse). Cells analyzed for cell volume were: 144 (3M control), 168 (3M Hdac1 cKO), 181 (13M control), and 227 (13M Hdac1 cKO). c , d Representative images and quantification of the comet assay using hippocampal homogenates. c , d Nuclei analyzed (three mice/group) were: 1356 (3M control), 1268 (3M Hdac1 cKO), 1898 (13M control), and 1944 (13M Hdac1 cKO). e Analysis of freezing behavior during contextual fear conditioning test. Animals analyzed were: 13 (3M control), 10 (3M Hdac1 cKO), 14 (13M control), and 22 (13M Hdac1 cKO). f , g Control and Hdac1 cKO mice were subjected to the Morris water maze (MWM) test. Shown are the latencies to hidden platform during training and time spent in each quadrant during the probe trial. Images show the location of each quadrant and swim trace of mouse during the probe trial. T = target, O = opposite, R = right, L = left. Animals analyzed were: 9 (3M control), 7 (3M Hdac1 cKO), 7 (17M control), and 11 (17M Hdac1 cKO). h , i Hippocampal LTP induction in control and Hdac1 cKO mice. Animals/slices analyzed were: 3/6 (2M control), 3/7 (2M Hdac1 cKO), 4/8 (6M control), and 5/8 (6M Hdac1 cKO). Overlay of the representative field excitatory post-synaptic potential slope (fEPSPs) before (black trace) and after (red trace) LTP induction. Theta-burst stimulation (TBS). All values are shown as mean ± SEM. Statistical analysis: b , d one-way ANOVA with Tukey’s post hoc test; e , h , i two-tailed Student’s t test; f , g two-way repeated-measures ANOVA with Bonferroni’s post hoc test for acquisition phase; two-way ANOVA with Bonferroni’s post hoc test for probe trial. Source data are provided as a Source Data file.

Article Snippet: Released nuclei were then pelleted down at 1600 × g for 7 min and incubated with Alexa Flour APC-conjugated GFAP antibody (Cell Signaling Technology, 3657, 1:200) and Alexa Flour 488-conjugated NeuN (Millipore, MAB377X, 1:200) in cold PBS with 1% BSA and proteinase inhibitor cocktail on a tube rotator at 4 °C for 3 h. Pelleted nuclei were then resuspended in 0.5 mL PBS and were run through a 0.45 μm cell strainer prior to FACS sorting.

Techniques: Control, Staining, Single Cell Gel Electrophoresis, Two Tailed Test

Journal: Nature Communications

Article Title: HDAC1 modulates OGG1-initiated oxidative DNA damage repair in the aging brain and Alzheimer’s disease

doi: 10.1038/s41467-020-16361-y

Figure Lengend Snippet: a Enriched GO terms of DEGs from 13M animals were annotated using MSigDB , . b FPKM (Fragments Per Kilobase of transcript per Million mapped reads) values of DEGs identified in 13M Hdac1 cKO mice function in ion transport, aging process, PKC pathway, protein chaperone, and antioxidant response. Cell cycle-related genes were unaltered. c Known motifs identified by HOMER program within upstream and downstream (±) 500 base pairs (bp) from TSS (transcriptional start site) of DEGs. d ChIP-qPCR analysis indicates the enrichment of 8-oxoG at gene promoters (500 bp upstream of TSS). At least five animals were analyzed from two independent experiments. e , f Representative images and quantification plots of the alkaline and FPG-modified comet assay using hippocampal homogenates from 13M control and Hdac1 cKO mice. e , f Nuclei analyzed (three mice/group) were: alkaline, 1928 (13M control), 1412 (13M Hdac1 cKO); FPG + alkaline, 1872 (13M control) and 1500 (13M Hdac1 cKO). g Representative images of 13M control and Hdac1 cKO mice stained for 8-oxoG (green), NeuN (red), and GFAP (magenta). h Quantification plots showing the 8-oxoG intensity. g , h Four mice were analyzed (two sections/mouse). Cells analyzed were: NeuN + cells, 1585 (13M control), 2306 (13M Hdac1 cKO); GFAP + cells, 191 (13M control), 164 (13M Hdac1 cKO). All values are shown as mean ± SEM. Statistical analysis: a weighted Kolmogorov–Smirnov-like test followed by multiple hypothesis testing; c cumulative binomial distributions; d , f one-way ANOVA with Tukey’s post hoc test; h two-tailed Student’s t test. Source data are provided as a Source Data file.

Article Snippet: Released nuclei were then pelleted down at 1600 × g for 7 min and incubated with Alexa Flour APC-conjugated GFAP antibody (Cell Signaling Technology, 3657, 1:200) and Alexa Flour 488-conjugated NeuN (Millipore, MAB377X, 1:200) in cold PBS with 1% BSA and proteinase inhibitor cocktail on a tube rotator at 4 °C for 3 h. Pelleted nuclei were then resuspended in 0.5 mL PBS and were run through a 0.45 μm cell strainer prior to FACS sorting.

Techniques: ChIP-qPCR, Modification, Single Cell Gel Electrophoresis, Control, Staining, Two Tailed Test

Journal: Nature Communications

Article Title: HDAC1 modulates OGG1-initiated oxidative DNA damage repair in the aging brain and Alzheimer’s disease

doi: 10.1038/s41467-020-16361-y

Figure Lengend Snippet: a , b Representative images and quantifications of 8-oxoG intensity in NeuN + cells and GFAP + cells. 14M animals stained for 8-oxoG (green), NeuN (red), and GFAP (magenta). a , b Three mice were analyzed per group (two sections/mouse). Nuclei analyzed were: NeuN + cells, 409 (control), 683 ( Hdac1 cKO), 962 (5XFAD), 910 ( Hdac1 cKO; 5XFAD); GFAP + cells, 104 (control), 121 ( Hdac1 cKO), 204 (5XFAD), 155 ( Hdac1 cKO; 5XFAD). White arrows indicate GFAP + cells. c Known motifs identified by HOMER program within ±500 bp from TSS of DEGs c 13M control and 5XFAD mice. d 13M control and Hdac1 cKO; 5XFAD mice. e Bar graphs showing the amount of DEGs overlaps between groups ( Hdac1 cKO, 5XFAD, and Hdac1 cKO; 5XFAD mice at the age of 13 months) and enrichment significance. f Hippocampal tissues from 13M control and 5XFAD mice were assayed for HDAC1 activity. Five animals were analyzed in each group. g A Venn diagram showing the overlap in downregulated genes between Hdac1 cKO, 5XFAD, and Hdac1 cKO; 5XFAD mice at the age of 13 months. h Enriched GO terms of overlapping genes between Hdac1 cKO, 5XFAD, and Hdac1 cKO; 5XFAD mice at the age of 13 months, annotated using MSigDB , . All values are shown as mean ± SEM. Statistical analysis: b one-way ANOVA with Tukey’s post hoc test, n.s., not significant. P value of each comparison: Hdac1 cKO versus 5XFAD = 0.8823; Hdac1 cKO versus Hdac1 cKO; 5XFAD = 0.5983; 5XFAD versus Hdac1 cKO; 5XFAD = 0.1215; c , d cumulative binomial distributions; e two-tailed Fisher’s exact test; f two-tailed Student’s t test; h weighted Kolmogorov–Smirnov-like test followed by multiple hypothesis testing. Source data are provided as a Source Data file.

Article Snippet: Released nuclei were then pelleted down at 1600 × g for 7 min and incubated with Alexa Flour APC-conjugated GFAP antibody (Cell Signaling Technology, 3657, 1:200) and Alexa Flour 488-conjugated NeuN (Millipore, MAB377X, 1:200) in cold PBS with 1% BSA and proteinase inhibitor cocktail on a tube rotator at 4 °C for 3 h. Pelleted nuclei were then resuspended in 0.5 mL PBS and were run through a 0.45 μm cell strainer prior to FACS sorting.

Techniques: Staining, Control, Activity Assay, Comparison, Two Tailed Test