EMBO/soda-nel huggingface dataset:
Novel tau biomarkers phosphorylated at T181, T217 or T231 rise with subtle changes in Aβ pathology (2020)
Figure 3

Figure 3. Effect of Aβ pathology (Aβ PET) on p-tau biomarkers

(A, B, C, D, E) Dot and box-plot comparing each of the p-tau biomarker between the Aβ negative (A-; blue; n = 287) and the Aβ positive (A+; red; n = 42) groups. Aβ positivity was defined with Aβ PET visual read. The box-plots depict the median (horizontal bar), interquartile range (IQR, hinges) and 1.5 x IQR (whiskers). P-values were assessed by a one-way ANCOVA adjusted by age and sex.

(F, G, H, I, J) Scatter plots depicting the changes between each p-tau biomarker as a function of Aβ PET Centiloids (CL). The standardized regression coefficients (β) and the P-values were computed using a linear model adjusting for age and sex. The solid lines indicate the regression line and the 95% confidence intervals. The dashed green lines indicate the CL12 and CL30 cutoffs. Participants were also colour-coded based on the CSF Aβ42/40 ratio (A-, black; A+, turquoise).

(K, L, M, N, O) Dot and box-plots depicting comparison between each of the p-tau biomarker between Centiloid scale groups: (i) ≤12CL (blue; n = 278), (ii) 12 - 30CL (subthreshold Aβ pathology group; grey; n = 28), (iii) >30CL (red; n = 25). The box-plots depict the median (horizontal bar), interquartile range (IQR, hinges) and 1.5 x IQR (whiskers). P-values were assessed by a one-way ANCOVA adjusted by age and sex, followed by a Bonferroni-corrected post hoc pairwise comparison.

Data information: Abbreviations: CSF, cerebrospinal fluid; Mid, mid-region; N, N-terminal; p-tau, phosphorylated tau.
Novel tau biomarkers phosphorylated at T181, T217 or T231 rise with subtle changes in Aβ pathology (2020)
Figure 4

Figure 4. Discrimination of cognitively unimpaired Aβ-positive from Aβ negative individuals by p-tau biomarkers.

A-C ROC analysis was performed to test the accuracy to discriminate between Aβ-positive (A+) from Aβ-negative (A-) individuals. Aβ positivity was defined as CSF Aβ42/40 < 0.071 (A), Aβ PET positive visual read (B) or Aβ PET Centiloid (CL) > 12 (C). Abbreviations: CSF, cerebrospinal fluid; Mid, mid-region; NfL, neurofilament light; N, N-terminal; p-tau, phosphorylated tau; t-tau, total tau.
Novel tau biomarkers phosphorylated at T181, T217 or T231 rise with subtle changes in Aβ pathology (2020)
Figure 5

Figure 5. Comparison of the p-tau biomarkers between AT groups.

A-D Dot and box-plot showing the levels of each p-tau biomarker in each of the AT groups. Aβ-positive (A+) was defined by a CSF Aβ42/40 < 0.071 and Tau-positive (T+) by an Elecsys CSF Mid-p-tau181 > 24 pg/ml. The box-plots depict the median (horizontal bar), interquartile range (IQR, hinges) and 1.5 x IQR (whiskers). The horizontal dashed line indicates the median of the p-tau biomarker in the A-T- group. P-values were assessed by a one-way ANCOVA adjusted by age and sex followed by Bonferroni-corrected post hoc pairwise comparisons. Abbreviations: CSF, cerebrospinal fluid; Mid, mid-region; N, N-terminal; p-tau, phosphorylated tau.
Loss of T-bet confers survival advantage to influenza-bacterial superinfection (2018)
Figure 1

Figure 1. T-bet deficient lungs exhibit increased cellular infiltration and reduced leakiness following influenza virus infection.

Mice were infected with sub-lethal dose (25 pfu) of influenza virus and assessed as follows.

(A) Percentage body weight loss. See also Figs EV1A and B.

(B) Relative amount of PR8 viral NP mRNA, by qPCR.

(C) Histology of lungs by H&E staining with pathology scores. Scale bars = 200 μm. See also Fig EV1C.

(D) Absolute number of live CD45+ cells in whole lungs and BAL, by flow cytometry.

(E) Level of LDH in BAL, by LDH cytotoxicity assay.

(F) Expression of PDPN in lungs, by western blot. See also Figs EV1D and E.

(G) Total protein in BAL, by Bradford assay.

(H) Concentration of albumin in BAL, by ELISA.

Data information: Data are representative of three independent experiments with 3-5 mice per group and represented as mean ± S.E.M. Statistical significance in (D), (G) and (H) was determined with unpaired student's T-test.
Loss of T-bet confers survival advantage to influenza-bacterial superinfection (2018)
Figure 4

Figure 4. Increased IL-17 and IL-22 production in T-bet-/- lungs infected with influenza virus or superinfected with bacteria.

(A) Concentrations of cytokines or chemokines in the BAL of days 5 and 9 post sub-lethal influenza virus infection as determined by protein bead array. See also Figs EV4A and B.

(B, C) Mice were superinfected as described in Fig 3A. (B) Concentrations of IL-17 and IL-22 in BAL of mice, as determined by ELISA. See also Fig EV3E. (C) Concentrations of IL-17 and IL-22 in BAL of mice 12 h after with 2x105 cfu of S. pneumoniae serotype 19F alone (-Flu), or following superinfection (+Flu), assessed by ELISA.

Data information: Data in (A) are obtained from one independent experiment with 4 mice per group, and results of selected analytes IFNγ, CXCL10, IL-6, IL-13 and IL-17 are reconfirmed in a separate independent experiment by a customized protein bead array. Data in (B) and (C) are representative of two independent experiments with 3-4 mice per group. All data are represented as mean ± S.E.M. Significance in (A-C) was determined by unpaired student's T-test. n.s. = not significant.
Loss of T-bet confers survival advantage to influenza-bacterial superinfection (2018)
Figure 5

Figure 5. Increased immunity against bacterial superinfection in T-bet-/- mice is abrogated by neutralization of IL-17 or neutrophil depletion.

Mice were superinfected as described in Fig 3A.

(A) Bacterial load as determined by enumeration of cfu in BAL following administration of αIL-17 or αIL-22 blocking antibody. Control wild-type and T-bet-/- groups were administered with IgG isotype control antibody.

(B) T-bet-/- mice were administered αIL-17 or IgG isotype control antibody every other day from day -2 post-secondary bacterial infection and monitored for survival.

(C) Absolute neutrophil counts in the lungs following administration of αIL-17 or IgG isotype control antibody, as determined by flow cytometry.

(D) Pneumococcal cfu in BAL following αLy6G antibody mediated neutrophil depletion. Control T-bet-/- mice were administered with IgG isotype control antibody. See also Fig EV5A.

(E) mRNA expression of antimicrobial peptides in lungs following administration of αIL-17, IL-22 or IgG control antibody, as determined by qPCR.

Data information: Data in (A), (B) and (D) are combined from two independent experiments (n = 8-12 mice per group). Data in (C) and (E) are representative of two independent experiments with 3-4 in (C) or 4-6 in (E) mice per group. All data are represented as mean ± S.E.M. Statistical significance was determined with Kruskal-Wallis test followed by Dunn's multiple comparisons test in (A), logrank test in (B), unpaired-students t-test in (C), Mann-Whitney U test in (D) and one-way ANOVA followed by Tukey's multiple comparisons test in (E). n.s. = not significant. See also Figs EV5B and C.
Loss of T-bet confers survival advantage to influenza-bacterial superinfection (2018)
Figure 6

Figure 6. CD8+, CD4+ and γδ T cells are major sources of IL-17 in influenza and secondary bacterial superinfected T-bet-/- lungs.

Mice were superinfected as described in Fig 3A.

(A) Frequency of IFNγ+ and IL-17+ immune cells following intracellular cytokine staining and flow cytometry.

(B) Representative flow cytometry plots of total and IL-17+ lymphocyte subsets on day 3 post-secondary bacterial infected lungs following intracellular cytokine staining. Numbers in plots indicate the frequency of each IL-17+ cell subset over total numbers of respective cell-types.

(C) Frequency over total IL-17+ cells and absolute numbers of each IL-17 producing lymphocyte subset on days 0, 1 and 3 post-secondary bacterial superinfected lungs as assessed by flow cytometry of intracellular cytokine stained cells.

Data information: Data are representative of at least two independent experiments with 3 mice per group. All data are represented as mean ± S.E.M. Statistical significance in (A) and (C) was determined by unpaired student's T-test.
The dynamic membrane recruitment of miRNA processing complexes in neurons controls dendritogenesis (2017)
Figure 1

Figure 1: The microRNA-generating complex localizes at the neuronal endoplasmic reticulum

Components of the RLC co-precipitate with ER markers in density gradient fractionation. The post-nuclear supernatant of cortical neurons was loaded underneath a continuous Optiprep density gradient. ER markers Calnexin, Ribophorin I and Climp63, co-sedimented with ribosomes (RPS6, ribosomal protein subunit 6), mitochondria (Tim23) and RLC proteins at fractions 3-5. The bottom graph shows the density of each fraction. B) TRBP, PACT and Ago2 partially co-localize with the ER marker mCherry-CLIMP63. Hippocampal neurons (10 DIV) expressing mCherry-CLIMP63 were immunostained with TRBP, PACT or Ago2 antibodies. Boxed insets are magnifications of primary dendrites as depicted by the adjacent letters (scale bars; 10μm). C) GFP-Dicer partially co-localizes with mCherry-CLIMP63 at the neuronal soma and primary dendrites of hippocampal neurons. Neurons were imaged at 10 DIV using total internal reflection fluorescence (TIRF) microscopy using a penetration depth of 150nm. Boxed insets are magnifications of primary dendrites as depicted by the adjacent letters (scale bars; 2μm or 10μm). D) The majority of Dicer, TRBP and PACT are associated with neuronal membranes. Sequential detergent extraction was used to separate cytoplasmic from membrane fractions in 7 DIV cortical neurons. The relative quantity of depicted proteins was analyzed in three independent experiments (p<0.005, t-test type 3, n=3, error bars s.d.). Actin and ribophorin I were used as loading controls for cytoplasmic and membrane fractions, respectively. E) TRBP is cytoplasmic in primary astrocytes. Sequential detergent extraction was performed in primary astrocyte cultures. Actin and calnexin were used as loading controls for cytoplasmic and membrane fractions, respectively. F) The majority of Dicer, TRBP and PACT proteins are present in cytoplasmic fractions in HEK293T cells, isolated using sequential detergent extraction.
The dynamic membrane recruitment of miRNA processing complexes in neurons controls dendritogenesis (2017)
Figure 2

Figure 2: Short BDNF stimulation leads to the re-distribution of TRBP from the ER to the cytoplasm.

A) BDNF stimulation leads to a decrease in TRBP levels from ER microsomes. Cortical neurons were stimulated BDNF with or vehicle control and ER microsomes were isolated using differential centrifugation. Protein intensity values were normalized to calnexin in each of three independent experiments (p=0.008, t-test type 3, n=3, error bars; s.d.). B) Endogenous TRBP partially co-localizes with calnexin at the neuronal cell soma and primary dendrites. Hippocampal neurons were transfected with GFP at 7DIV and fixed at 10DIV following BDNF or control stimulation. Boxed insets are magnifications of the primary dendrite (15μm in length). Arrows point to overlapping puncta (scale bars; 20μm (GFP) or 10μm). C) BDNF leads to a decrease in co-localization between TRBP and calnexin at the soma and primary dendrites of hippocampal neurons. Manders co-localization co-efficients between TRBP and Calnexin were measured in the cell soma or primary dendrites (15μm) using GFP as a morphological mask. Average co-localization values were normalized to controls in each experiment (p<0.008, t-test type 3, n=4, error bars; s.d.). D) BDNF decreases mCherry-CLIMP63 co-localization with TRBP, but not with PACT or Ago2. Hippocampal neurons were transfected with GFP and mCherry-CLIMP63 at 7DIV and immunostained three days later, following BDNF or control stimulation. Manders co-localization was measured at the cell soma using GFP as a mask as indicated (p=0.01, t-test type 3, n=4, error bars; s.d.). E) BDNF leads to an increase in the levels of TRBP (p=0.03) and PACT (p=0.004) in the cytoplasmic fraction. Cortical neurons (7 DIV) were control- or BDNF-stimulated and cytoplasmic fractions were isolated using digitonin. Protein intensity values were normalized to actin in each experiment. The position of the star corresponds to the TRBP band used for analysis (~43kDa) (t-test type 3, n=4, error bars; s.d.). F) The BDNF-induced increase in cytoplasmic TRBP is dependent on intracellular calcium. Cortical neurons (7 DIV) were treated with vehicle control, BDNF and BAPTA-AM as indicated, and cytoplasmic fractions were isolated using digitonin. Protein intensity values were normalized to actin in each independent experiment (p=0.03, n=3, error bars; s.d.).
The dynamic membrane recruitment of miRNA processing complexes in neurons controls dendritogenesis (2017)
Figure 3

Figure 3: BDNF leads to TRBP dissociation from Dicer and decreased processing of pre-miR16.

BDNF stimulation causes a decrease in the TRBP-Dicer interaction. Young cortical neurons were treated with BDNF or vehicle control and co-immunoprecipitations (co-IPs) were performed using 2μg control IgG or polyclonal anti-TRBP antibodies. Dicer binding in co-IP was normalized to inputs in each experiment (p=0.001, t-test type 2, n=3, error bars represent s.d.). B) Short BDNF stimulation changes the levels of only a few microRNAs, such as miR-16-5p and miR-551b-5p. Volcano plot of small RNA sequencing data from BDNF- or control-treated cortical neurons (6DIV) was based on differential expression analysis using the edgeR package. Individual points represent the average fold change obtained in three independent experiments for each miRNA, and plotted against obtained p-values. MiRNAs above the red dashed line are significant (p<0.05, t-test type 2, n=3). C) BDNF stimulation leads to a transient decrease in mature miR-16-5p levels. 6DIV cortical neurons were treated with BDNF for 10 or 20 minutes and mature miRNA levels were measured in rtPCR. Ct values for each miRNA were normalized to U6 and to 20 minutes of control stimulation (n=3, p=0.02, t-test type 3, error bars; s.d.). D) BDNF stimulation leads to a transient increase in the levels of pre-miR16 after 10 minutes of BDNF stimulation. Ct values obtained in rtPCR were normalized to U6 and to 20 minutes control stimulation (n=4, t-test type 3, error bars; s.d.). E) BDNF causes a decrease in pre-miR16 binding to TRBP. Cortical neurons were treated with BDNF or vehicle control and RNA IPs were performed using a mouse monoclonal anti-TRBP antibody. Ct values measured in rtPCR were normalized to respective inputs (n=3, p=0.002, t-test type 3, error bars represent s.d.).
The dynamic membrane recruitment of miRNA processing complexes in neurons controls dendritogenesis (2017)
Figure 4

Figure 4: BDNF changes the relative isomiRs levels for members of the miR16 family.

A) BDNF changes the canonical isomiR proportion of a subset of miRNAs. IsomiR levels were compared to the total amount of the corresponding miRNA. Shown are the miRNAs that had significant changes in the canonical isomiR (annotated as '0', p<0.05, t-test type 2, n=3). Results are shown in order of decreasing significance. Shown in red are miRNAs of the miR16 family, miR-16-5p and miR-15b-5p. Black squares correspond non-existing isomiRs. Numbers on the top of the table represent the trimming (negative) or addition (positive) of nucleotides to the 3'- or 5'-end of the canonical 5p or 3p isomiRs, respectively. Only templated nucleotide additions and trimmings, which might reflect Dicer-mediated pre-miRNA cleavage, were considered for analysis. B) BDNF stimulation leads to a decrease in the canonical (22 nucleotides) isomiRs of the miR16 family, miR-16-5p (p=0.005) and miR-15b-5p (p=0.02). In both cases, this is accompanied by an increase in the levels of shorter isomiRs; the 19-nucleotide isoform of miR-16-5p (p=0.04) and the 21-nucleotide isoform of miR-15b-5p (p=0.007). The relative isomiRs proportion was calculated as described in A (t-test type 2, n=3, error bars; s.d.). C) Schematic representation of the pre-miRNA sequence and stem loop structure for pre-miR16 and pre-miR15b. The 5p and 3p canonical miRNA sequences are highlighted in pink. The red arrows depict the predicted Dicer cleavage sites that would lead to shorter isomiRs induced by BDNF, as shown in B.
The dynamic membrane recruitment of miRNA processing complexes in neurons controls dendritogenesis (2017)
Figure 6

Figure 6: The BDNF mRNA is a miR-16-5p target and is regulated during BDNF stimulation

Diagram showing the region of the BDNF 3'UTR bound by miR16 family of miRNAs that is conserved across species. Nucleotides colored in red depict the binding site of miR-16-5p (sequences obtained from www.targetscan.org). B) MiR-16-5p targets the 3'UTR of the BDNF mRNA. Cortical neurons were transfected with 100ng pmiRGlo dual-luciferase reporter consisting of the 3'UTR of BDNF (3'UTR WT) or a miR-16-5p-binding site mutant (3'UTR Mut) and either miR-Ctrl or miR-16. The Firefly to Renilla luciferase ratio was measured and values were normalized to the respective reporter expression without miRNA mimic co-transfection (n=3, p=0.02, t-test type 2). C) BDNF stimulation leads to an increase in the translation of 3'UTR WT (p=0.02, t-test type 3) but not 3'UTR Mut luciferase reporter, which is rescued by miR-16-5p overexpression (p=0.002; t-test type 2). Cortical neurons were transfected as in (B), stimulated with BDNF or vehicle control for 20 minutes, and lysed two hours after stimulation (n=4). D) Pre-miR16 is present in the neuronal cell soma and processes of hippocampal neurons. Neurons were plated on compartmentalized chambers and RNA was isolated from the neuronal cell body and processes separately. Pre-miR137 and pre-miR341 were used to assay somatic and neurite enrichment, respectively. Raw Ct values were normalized to GAPDH mRNA in each experiment (n=3).
The dynamic membrane recruitment of miRNA processing complexes in neurons controls dendritogenesis (2017)
Figure 7

Figure 7: Membrane-associated TRBP expression blocks the BDNF-induced decrease of miR16 and BDNF-induced dendritogenesis.

More Dicer binds to membrane-targeted GFP-TRBP (mGFP-TRBP) compared to GFP-TRBP. HEK293T cells were transfected with GFP, GFP-TRBP or mGFP-TRBP and cell lysates were processed for co-IP using anti-GFP antibody. Dicer binding was normalized to respective input lanes in each condition (n=4, p=0.02; t-test type 2, error bars are s.d.). B) mGFP-TRBP but not GFP-TRBP blocks the BDNF-induced decrease in miR-16-5p activity in dual-fluorescence sensor assay. Hippocampal neurons were transfected with 200ng miR16- or control-sensor (see Fig 5A) plus 50ng GFP, 400ng GFP-TRBP or 400ng mGFP-TRBP. Neurons were treated with BDNF or vehicle control prior to fixation and immunostained with anti-GFP antibody. BDNF led to a decrease in miR-16-5p activity in GFP-transfected neurons (p=0.03, n=4, t-test type 3), which was significantly different from mGFP-TRBP-expressing neurons (p=0.04, n=4, t-test type 2, error bars are s.d.). C&D) BDNF-induced dendritogenesis is blocked by mGFP-TRBP overexpression. C) Hippocampal neurons (4DIV) were transfected for three days with 200ng dsRed and 50ng GFP, 400ng GFP-TRBP or 400ng mGFP-TRBP. Neurons were treated with BDNF or vehicle control 24 hours before fixation. Pictures were obtained on a confocal microscope and threshold was set for dsRed to distinguish individual dendrites (scale bars are 10μm). D) The total number of intersections was calculated in Sholl analysis, and values were normalized to control treatment in each of three independent experiments. BDNF led to a significant increase in dendritic complexity (p=0.04; t-test type 3), which was significantly different in cells transfected with mGFP-TRBP, but not GFP-TRBP (n=3, p=0.03; t-test type 2, error bars; s.d.).
Reduction of Mitochondrial Iron Alleviates Cardiac Damage During Injury (2016)
Figure 3

Figure 3. Cardiac-specific ABCB8 overexpression reduces mitochondrial iron but does not alter cardiac function, ROS or the antioxidant system.

A. Mitochondrial non-heme iron in ABCB8 NTG and TG mice. N=4 mice for NTG and 6 mice for TG. * P=0.021 with two-tailed unpaired T-test.

B. Cytosolic non-heme iron in ABCB8 NTG and TG mice. N=4 mice for NTG and 6 mice for TG. Two-tailed unpaired T-test was performed.

C. Baseline ejection fraction of littermate ABCB8 NTG and TG mice. N=6 mice in each group. Two-tailed unpaired T-test was performed.

D. Baseline fractional shortening of littermate ABCB8 NTG and TG mice. N=6 mice in each group. Two-tailed unpaired T-test was performed.

E. Lipid peroxidation products in hearts of TG and NTG mice. N=6 mice in each group. Two-tailed unpaired T-test was performed.

F. Relative expression of antioxidant genes in NTG and TG mice. N=6 mice in each group. Two-tailed unpaired T-test was performed. All data are expressed as mean ± SEM. N.S. = not signigicant.
Reduction of Mitochondrial Iron Alleviates Cardiac Damage During Injury (2016)
Figure 4

Figure 4. ABCB8 transgenic mice have less cell death and better cardiac function after I/R injury.

A. Cardiac function in ABCB8 NTG and TG mice undergoing sham or I/R procedure. ANOVA followed by post-hoc Tukey test was performed for each time point. * P<0.05 compared with TG-sham at the same time point. $ P<0.05 compared with NTG-I/R at the same time point. § P<0.05 compared with NTG-sham at the same time point. N=5 mice for NTG-sham and NTG-I/R, and N=6 mice for TG-sham and TG-I/R. Exact P-values are included in Appendix Table S3.

B. Representative images of TUNEL staining for mice with indicated genotype undergone indicated procedure. Scale bar = 100μm.

C. Quantification of apoptosis in ABCB8 TG and NTG mice with indicated procedure. ANOVA followed by post-hoc Tukey test was performed. * P=0.035. N=4 mice for TG-I/R, and N=6 mice for all other groups.

D. Nppa expression in mice subjected to sham or I/R procedure. ANOVA followed by post-hoc Tukey test was performed. * P=3.24E-4 NTG-sham vs. NTG-I/R. * P=0.002 TG-sham vs. TG-I/R. * P=8E-6 NTG-I/R vs. TG-I/R. N=5 mice for NTG-sham and NTG-I/R, and N=6 mice for TG-sham and TG-I/R.

E. Nppb expression in mice subjected to sham or I/R procedure. ANOVA followed by post-hoc Tukey test was performed. * P<1E-8 NTG-sham vs. NTG-I/R. * P=0.032 NTG-I/R vs. TG-I/R. N=5 mice for NTG-sham and NTG-I/R, and N=6 mice for TG-sham and TG-I/R.

F. Myh7 expression in mice subjected to sham or I/R procedure. ANOVA followed by post-hoc Tukey test was performed. * P=0.04 NTG-sham vs. NTG-I/R. * P=0.034 NTG-I/R vs. TG-I/R. N=5 mice for NTG-sham and NTG-I/R, and N=6 mice for TG-sham and TG-I/R. All data are expressed as mean ± SEM. N.S. = not signigicant.