Journal: bioRxiv

Article Title: Early mitochondrial dysfunction proceeds neuroinflammation, synaptic alteration, and autophagy impairment in hippocampus of App knock-in Alzheimer mouse models

doi: 10.1101/2023.03.07.531542

Figure Lengend Snippet: (A) Hippocampi were dissected from two-, six-, 12-month-old App NL-G-F mice and six-, 12-, 18-month-old App NL-F mice and age matched WT controls. RNA was extracted from dissected hippocampi and cDNA libraries were synthesized. Finally, RNA sequencing was performed by a Hiseq 3000 sequencer (n = 3). Validation studies were performed by RT-qPCR using the same RNA samples for RNA sequencing. Western blotting with hippocampal brain homogenate and Olink proteomics were conducted for validations at protein level. Mitochondria and crude synaptosomal fraction were isolated for mechanistic studies of mitochondrial and autophagic functions. Mitochondrial dysfunction and autophagic alterations were revealed in App NL-G-F mice by electron microscopy. (B) t- SNE plot representing difference of App NL-F , App NL-G-F and WT transcriptomes. Each symbol represents one mouse individual. Each color represents one mouse genotype, red: App NL-F , blue: App NL-G-F , gray: WT, and each shape of symbol represents mouse age, circle: two-month-old, triangle: six-month-old, inverted triangle: 12-month-old, square: 18-month-old. (C) Venn diagram of significantly DEGs in App NL-F and App NL-G-F vs WT mice (FDR < 0.1). (D) Volcano plots of the gene expression profiles in different time points of App NL-F or App NL-G-F vs WT mice. Red and blue dots indicate significantly up- and downregulated genes (FDR < 0.1), respectively, in App knock-in mice. Grey dots indicate non-significantly altered genes. Genes validated by RT-qPCR are highlighted. (E) Heatmap of selected pathways related to AD, glucose metabolism, neuroinflammation and autophagy in App NL-F vs WT mice (columns 1 to 3), App NL-G-F vs WT mice (columns 4 to 6) and App NL-F vs App NL-G-F mice (columns 7 and 8). The p -value of each enriched pathway was converted to Z score. Significantly up- and downregulated pathways have the absolute value of Z scores ≥ 1.96. (F) Relative mRNA expression of Ide gene was normalized to Tubb3 (n = 3, left). Statistical significance was analyzed using Kruskal-Wallis tests followed by Dunn’s multiple comparison test. * p < 0.05. * App NL-F mice vs age matched WT controls, # vs two-month-old WT mice. IDE protein levels in hippocampal homogenates were quantified by Western blotting and normalized to β3-tubulin (n = 3, middle and right). Statistical significance was analyzed using one-way ANOVA followed by Tukey’s multiple comparisons test. *** p < 0.001, **** p < 0.0001. * vs age matched WT controls.

Article Snippet: Gene expression was normalized to b3-tubulin mRNA (Mm00727586_s1).

Techniques: Synthesized, RNA Sequencing, Biomarker Discovery, Quantitative RT-PCR, Western Blot, Isolation, Electron Microscopy, Gene Expression, Knock-In, Expressing, Comparison

Journal: bioRxiv

Article Title: Early mitochondrial dysfunction proceeds neuroinflammation, synaptic alteration, and autophagy impairment in hippocampus of App knock-in Alzheimer mouse models

doi: 10.1101/2023.03.07.531542

Figure Lengend Snippet: (A) Chord plot of significantly altered genes (FDR < 0.1) related to inflammatory response. Color of the circle edge boxes indicate up-(red) or down-(blue) regulation. (B) Relative mRNA expression levels of selected inflammatory genes were normalized to Tubb3 (n = 3). Statistical significance was analyzed using Kruskal-Wallis test followed by Dunn’s multiple comparison test. * vs age-matched WT mice, + vs age-matched App NL-F mice, # vs genotype-matched two-month-old mice, * p < 0.05, ** p < 0.01, *** p < 0.001. (C) CSF was withdrawn from cisterna magna through the dura mater of 18-month-old mice and analyzed by proximity extension assay technology using mouse exploratory panel to detect the changes in proteins related to inflammation. Statistical significance was analyzed using Kruskal-Wallis tests followed by Dunn’s multiple comparison test. * p < 0.05.

Article Snippet: Gene expression was normalized to b3-tubulin mRNA (Mm00727586_s1).

Techniques: Expressing, Comparison

Journal: bioRxiv

Article Title: Early mitochondrial dysfunction proceeds neuroinflammation, synaptic alteration, and autophagy impairment in hippocampus of App knock-in Alzheimer mouse models

doi: 10.1101/2023.03.07.531542

Figure Lengend Snippet: (A) Chord plot of significantly (FDR < 0.1) DEGs related to autophagy. Color of the circle edge boxes indicate up-(red) or down-(blue) regulation. (B) The relative mRNA expression of selected genes was normalized to Tubb3 (n = 3). Statistical significance was analyzed using Kruskal-Wallis tests followed by Dunn’s multiple comparison test. * p < 0.05. * vs age matched WT mice, # vs 2-month-old genotype-matched mice. (C) Co-immunofluorescence staining of Aβ, LC3 and synaptophysin with 12 months-old WT and App NL-G-F mice (n = 4). Red: Aβ, Green: LC3, Yellow: Synaptophysin, Blue: nucleus. Scale bar: 20 µm. (D, E) Signal intensity of Aβ (Red), LC3 (Green), Synaptophysin (Yellow) and nucleus (Blue) under the white line of WT and App NL-G-F brain staining gated in C. (F-I) Phospho-p62 (S403), total p62, LC3-I and LC3-II protein levels in hippocampal crude synaptosomal fraction (P2) or soluble fraction (S2) were visualized by Western blotting (n = 4). Protein levels were normalized to β3-tubulin. Statistical significance was analyzed using unpaired t test. * p < 0.05, ** p < 0.01.

Article Snippet: Gene expression was normalized to b3-tubulin mRNA (Mm00727586_s1).

Techniques: Expressing, Comparison, Immunofluorescence, Staining, Western Blot

Journal: Aging (Albany NY)

Article Title: TRDMT1 participates in the DNA damage repair of granulosa cells in premature ovarian failure

doi: 10.18632/aging.203080

Figure Lengend Snippet: Reduced TRDMT1 is associated with decreased ovarian function. ( A ) Histopathological examination of the ovaries of the WT, POF-PBS and POF-BMSCs groups. Scale bar: 100 μm. ( B ) Primordial follicles, primary follicles, secondary follicles, and atretic follicles were observed. ( A ) Transmission electron microscopy analysis of the ovarian structures in each group of mice. Scale bar: 2μm. ( A ) Apoptosis of GCs in ovaries was measured by TUNEL staining. Representative images of terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining show apoptotic GCs in each group. The TUNEL-positive apoptotic cells are indicated by green fluorescence. The nuclei (blue) were stained with DAPI. Scale bar: 100μm. ( A ) Representative immunohistochemical images of TRDMT1 in each group. Brown staining represents a positive TRDMT1 signal. Scale bar: 100μm. ( C ) Representative immunofluorescent staining of TRDMT1 in GCs from women with normal ovarian function and from patients with POF. Nuclei were stained with DAPI. Scale bar: 50μm. ( D ) TRDMT1 expression in the serum of healthy people and patients with POF was detected by Western blotting. An anti-Tubulin antibody was used as a loading control. *p<0.05, **P<0.01, and ***P<0.001. Statistical significance was determined using two-tailed t-tests for two groups and ANOVA for multiple comparisons. All values are means ± SD.

Article Snippet: After blocking, the membrane was incubated with primary antibodies against TRDMT1 and Tubulin (1:1000, mouse monoclonal, Boster, CHINA) combined with a horseradish peroxidase secondary antibody (1:5000, mouse monoclonal, Biosharp, CHINA).

Techniques: Transmission Assay, Electron Microscopy, TUNEL Assay, Staining, Fluorescence, Immunohistochemical staining, Expressing, Western Blot, Control, Two Tailed Test

Journal: Aging (Albany NY)

Article Title: TRDMT1 participates in the DNA damage repair of granulosa cells in premature ovarian failure

doi: 10.18632/aging.203080

Figure Lengend Snippet: TRDMT1 is involved in the regulation of GCs DNA damage and apoptosis. ( A ) Cells with FSHR expression are shown in red. Cell nuclei are stained blue. Scale bar: 100μm. ( B ) DNA damage of KGN, CTX-KGN-PBS and CTX-KGN-BMSCs was detected by comet assay. Scale bar: 100μm. ( C ) Annexin V-APC/AAD staining and flow cytometry analysis of the apoptotic rates of cells in the WT, CTX-KGN-PBS and CTX-KGN-BMSCs groups. Quantification of the apoptosis rates is shown. ( D ) Immunofluorescence staining and Western blotting of the expression levels of TRDMT1 in each group. Scale bar: 10μm. ( E ) DNA damage of CTX-KGN transfected with GFP-NC, GFP-TRDMT1, Sh-RNA and Sh-TRDMT1 was measured by comet assay. Comparison of the tail DNA of cells in the four groups. Scale bar: 100μm. ( F ) Annexin V-APC/AAD staining and flow cytometry analysis of the apoptotic rates of CTX-KGN transfected with GFP-NC, GFP-TRDMT1, Sh-RNA and Sh-TRDMT1. Comparison of the percentage of apoptotic cells in the four groups. *p<0.05, **P<0.01, and ***P<0.001. Statistical significance was determined using two-tailed t-tests for two groups and ANOVA for multiple comparisons. All values are means ± SD.

Article Snippet: After blocking, the membrane was incubated with primary antibodies against TRDMT1 and Tubulin (1:1000, mouse monoclonal, Boster, CHINA) combined with a horseradish peroxidase secondary antibody (1:5000, mouse monoclonal, Biosharp, CHINA).

Techniques: Expressing, Staining, Single Cell Gel Electrophoresis, Flow Cytometry, Immunofluorescence, Western Blot, Transfection, Comparison, Two Tailed Test

Journal: Aging (Albany NY)

Article Title: TRDMT1 participates in the DNA damage repair of granulosa cells in premature ovarian failure

doi: 10.18632/aging.203080

Figure Lengend Snippet: Oxidative stress induces DNA damage and apoptosis in GCs. ( G ) H 2 O 2 +KGN were harvested at the indicated time points for γ-H 2 AX staining. Positive cells in each group and the frequency of cells with foci were counted. Scale bar: 10μm. ( H ) KGN cells transfected with GFP-TRDMT1 (WT, G155V, and E63K mutant) were stimulated with H 2 O 2 , and their cell localization was observed. Scale bar: 50μm.Comparison of the nucleus/cytoplasm ratio between the two groups. *P<0.05, **p<0.01, and ***p<0.001. Statistical significance was determined using two-tailed t-tests for two groups and ANOVA for multiple comparisons. All values are means ± SD.

Article Snippet: After blocking, the membrane was incubated with primary antibodies against TRDMT1 and Tubulin (1:1000, mouse monoclonal, Boster, CHINA) combined with a horseradish peroxidase secondary antibody (1:5000, mouse monoclonal, Biosharp, CHINA).

Techniques: Staining, Transfection, Mutagenesis, Comparison, Two Tailed Test

Journal: Aging (Albany NY)

Article Title: TRDMT1 participates in the DNA damage repair of granulosa cells in premature ovarian failure

doi: 10.18632/aging.203080

Figure Lengend Snippet: TRDMT1 mediates GCs oxidative DNA damage repair. ( A ) The DNA damage of H 2 O 2 +KGN+PBS and H 2 O 2 +KGN+NAC transfected with GFP-NC, GFP-TRDMT1, Sh-RNA and Sh-TRDMT1 was measured via comet assay. Scale bar: 100μm. Comparison of the tail of DNA between the groups. ( B ) H 2 O 2 +KGN+PBS transfected with GFP-NC, GFP-TRDMT1, Sh-RNA and Sh-TRDMT1 were harvested at the indicated time points for γ-H 2 AX staining. Positive cells in each cell and the frequency of cells with foci were counted. Scale bar: 10μm. ( C ) H 2 O 2 +KGN+NAC transfected with GFP-NC, GFP-TRDMT1, Sh-RNA and Sh-TRDMT1 were harvested at the indicated time points for γ-H 2 AX staining. Positive cells in each group and the frequency of cells with foci were counted. Scale bar: 10μm.*P<0.05, **p<0.01, and ***p<0.001. Statistical significance was determined using two-tailed t-tests for two groups and ANOVA for multiple comparisons. All values are the means ± SD.

Article Snippet: After blocking, the membrane was incubated with primary antibodies against TRDMT1 and Tubulin (1:1000, mouse monoclonal, Boster, CHINA) combined with a horseradish peroxidase secondary antibody (1:5000, mouse monoclonal, Biosharp, CHINA).

Techniques: Transfection, Single Cell Gel Electrophoresis, Comparison, Staining, Two Tailed Test

Journal: Aging (Albany NY)

Article Title: TRDMT1 participates in the DNA damage repair of granulosa cells in premature ovarian failure

doi: 10.18632/aging.203080

Figure Lengend Snippet: The oxidative DNA damage repair promoted by TRDMT1 is related to RNA m5C methylation. ( A ) ROS in CTX-KGN were detected by DHE probe after transfection with GFP-NC or GFP-TRDMT1 (WT, G155V, and E63K mutant). Scale bar: 100 μm. ( B ) The DNA damage of CTX-KGN transfected with GFP-NC or GFP-TRDMT1 (WT, G155V, and E63K mutant) was measured via comet assay. Scale bar: 100μm ( C ) CTX-KGN transfected with GFP-NC and GFP-TRDMT1 (WT, G155V, and E63K mutant) were stained for γ-H 2 AX at the indicated time points. Scale bar: 10μm. Positive cells in each group and the frequency of cells with foci were counted. ( D ) Apoptosis of CTX-KGN transfected with GFP-NC or GFP-TRDMT1 (WT, G155V, E63K mutant) was assessed using Annexin V staining. Comparison of apoptotic rates between the groups. ( E ) The percentage of m5C in total RNA from CTX-KGN after transfection with GFP-NC or GFP-TRDMT1 (WT, G155V, and E63K mutant) was detected via ELISA-based assays. ( F ) The expression of RAD51 and RAD52 were detected after we stably expressed TRDMT WT and the mutants (TRDMT1 G155V/E63K ) in TRDMT1 KD CTX-KGN. *P<0.05, **p<0.01, and ***p<0.001. Statistical significance was determined using two-tailed t-tests for two groups and ANOVA for multiple comparisons. All values are means ± SD.

Article Snippet: After blocking, the membrane was incubated with primary antibodies against TRDMT1 and Tubulin (1:1000, mouse monoclonal, Boster, CHINA) combined with a horseradish peroxidase secondary antibody (1:5000, mouse monoclonal, Biosharp, CHINA).

Techniques: Methylation, Transfection, Mutagenesis, Single Cell Gel Electrophoresis, Staining, Comparison, Enzyme-linked Immunosorbent Assay, Expressing, Stable Transfection, Two Tailed Test

Journal: Aging (Albany NY)

Article Title: TRDMT1 participates in the DNA damage repair of granulosa cells in premature ovarian failure

doi: 10.18632/aging.203080

Figure Lengend Snippet: A schematic diagram showing the possible mechanism of TRDMT1 on GCs. CTX induced oxidative stress by generating ROS and thereby GCs apoptosis via DNA damage. The increased level of ROS leads to DNA damage. The increase in DNA damage promotes the apoptosis of GCs, thus promoting POF. TRDMT1 is important for DNA damage repair efficiency and survival. TRDMT1 participated in the DNA damage repair of GCs through methylation. m5C mRNA methylation is enriched at transcriptionally active sites with DNA damage. In short, oxidative damage to DNA induced the apoptosis of GCs and promoted the development of POF. TRDMT1 promoted DNA damage repair by regulating the methylation level, which reduced the apoptosis of GCs and inhibited the occurrence of POF. The regulation of oxidative DNA damage repair mediated by TRDMT1 was significantly correlated with its methylation activity.

Article Snippet: After blocking, the membrane was incubated with primary antibodies against TRDMT1 and Tubulin (1:1000, mouse monoclonal, Boster, CHINA) combined with a horseradish peroxidase secondary antibody (1:5000, mouse monoclonal, Biosharp, CHINA).

Techniques: Methylation, Activity Assay

Journal: The Journal of Neuroscience

Article Title: Genetic Contributions to Long-Range Temporal Correlations in Ongoing Oscillations

doi: 10.1523/JNEUROSCI.3083-07.2007

Figure Lengend Snippet: From broadband EEG to the DFA of narrow-band ongoing oscillations. A 10 s segment of EEG from occipital electrode O2 showing alpha oscillation bursts before (A) and after (B) bandpass filtering in the alpha-frequency range (8–13 Hz). We analyze the temporal structure of the amplitude (Ampl.) envelope of the oscillation, which is indicated with a thick line in B. Ongoing oscillations in the human EEG generally exhibit long-range temporal (auto-)correlations (LRTC), which are identified qualitatively as large variations in the duration and magnitude of the amplitude envelope over time as seen in the dizygotic twin siblings 1 and 2 (C, D). The temporal structure and correlations of the signal in C may be removed by randomly shuffling the signal in windows of 100 ms (E). The DFA exponent, α, provides a quantitative measure of LRTC, and the stronger correlations in dizygotic twin 1 (F, circles) compared with dizygotic twin 2 (F, squares) is reflected in a value of α closer to 1 (0.94 vs 0.68). The lack of temporal structure and correlations in E is reflected in the DFA exponent having the value of ∼0.5, which is characteristic of an uncorrelated random process (F, dots).

Article Snippet: Brain electric activity was recorded using Electro-cap electrodes placed according to the international 10–20 system ( n = 14 electrodes).

Techniques:

Journal: The Journal of Neuroscience

Article Title: Genetic Contributions to Long-Range Temporal Correlations in Ongoing Oscillations

doi: 10.1523/JNEUROSCI.3083-07.2007

Figure Lengend Snippet: Power-law decaying correlations in the amplitude fluctuations of alpha- and beta-frequency band oscillations. Three complementary autocorrelation analyses were performed: the autocorrelation function (Autocorr., A), the power spectral density (Power, B), and the DFA (C). Each analysis was performed on the amplitude envelope of alpha and beta oscillations and averaged across the 368 subjects and 14 electrodes (open circles; see Materials and Methods). The data have been fitted with a power-law function on time scales from 1 to 20 s (black lines). The autocorrelation function is plotted semilogarithmically, because the autocorrelation function can attain negative values; power spectral density and DFA are plotted in log-log coordinates. Each analysis points to a slow decay of correlation or LRTC. The dots indicate the analysis of computer-generated white noise that is filtered identically to the EEG data. The autocorrelation function is zero at all time lags, the power spectrum is flat, and the DFA exponent is close to 0.5 for both frequency bands, showing that the bandpass filters did not introduce autocorrelations in the amplitude time series on the long time scales investigated here.

Article Snippet: Brain electric activity was recorded using Electro-cap electrodes placed according to the international 10–20 system ( n = 14 electrodes).

Techniques: Generated, Introduce

Journal: The Journal of Neuroscience

Article Title: Genetic Contributions to Long-Range Temporal Correlations in Ongoing Oscillations

doi: 10.1523/JNEUROSCI.3083-07.2007

Figure Lengend Snippet: Genetic factors shape the power and the temporal correlation structure of ongoing oscillations. The scatter plots show the logarithmically transformed power (A) and the DFA exponents (B) of monozygotic (circles, n = 80 pairs) and dizygotic (pluses, n = 104 pairs) twin pairs for alpha and beta at occipital electrode O2. That both power and DFA of alpha and beta oscillations are heritable traits is indicated by the more than two times higher correlation between monozygotic twins than between dizygotic twins. Twin correlations, r, are indicated in the plots.

Article Snippet: Brain electric activity was recorded using Electro-cap electrodes placed according to the international 10–20 system ( n = 14 electrodes).

Techniques: Transformation Assay

Journal: The Journal of Neuroscience

Article Title: Genetic Contributions to Long-Range Temporal Correlations in Ongoing Oscillations

doi: 10.1523/JNEUROSCI.3083-07.2007

Figure Lengend Snippet: Genetic variances of DFA and power in ongoing oscillations are independent. There are no correlations between DFA exponents and power of ongoing alpha and beta oscillations at occipital electrode O2 (A), either before (upper plots) or after (lower plots) removing the subjects with a low SNR. B, At the central electrode C3, one may get the impression that DFA and oscillation power are correlated (upper plots). However, after removing 94 and 117 subjects with a low SNR in the alpha and beta band, respectively, we recover the nonsignificant zero correlation, which was observed also in occipital and parietal leads with a high SNR. The number of subjects included in each scatter plot is indicated with “n.” Significance levels of Pearson's coefficients of correlation: ns, p >0.05; **p < 0.001.

Article Snippet: Brain electric activity was recorded using Electro-cap electrodes placed according to the international 10–20 system ( n = 14 electrodes).

Techniques: