Journal: bioRxiv

Article Title: Antidepressants promote developmental-like plasticity through remodeling of extracellular matrix

doi: 10.1101/2025.01.03.631260

Figure Lengend Snippet: ( A ) Volcano plot of differentially expressed genes between mature granule cells from Cort and Cort + Flx (3wks) groups. ( B ) Top gene ontology terms for upregulated genes in (A). ( C ) Juvenile granule cell score defined as the top 100 genes upregulated in P5 granule cells versus P132 granule cells. Dataset from ; also see . ( D ) Transcriptomic profiles of dentate granule cells colored by treatment condition (Cort = red, Cort + Flx (3wks) = blue) and cell type (Mature = green outline, Immature = magenta outline). ( E ) Feature plot showing average expression of juvenile granule cell score (left) and quantification per mouse (right; One-way ANOVA, Tukey’s multiple comparisons test, n=3 mice/group). ( F ) Expression of Sox11 and Bdnf in granule cells. ( G ) SOX11 protein expression in the DG of juvenile (P10) and adult (∼P120) mice. PROX1 marks granule cell neurons. ( H ) Quantification of SOX11+ granule cells per mouse (t-test, n= 5 (juvenile), 4 (adult)). ( I ) SOX11 protein expression in the DG of adult mice treated with Cort or Cort + Flx (3wks). PROX1 marks granule cells, DCX marks immature granule cells. ( J ) Quantification of SOX11+ in mature (PROX1+ DCX-) and immature (PROX1+ DCX+) granule cells per mouse (One-way ANOVA, Tukey’s multiple comparisons test, n=3 (Cort), 3 (Cort + Flx 5d), 5 (Cort + Flx 3wks)). ( K ) Bdnf mRNA expression in the DG of juvenile (P10) and adult (∼P120) mice. ( L ) Quantification of Bdnf mRNA in the DG per mouse (t-test, n= 4 (juvenile), 3 (adult)). ( M ) Bdnf mRNA expression in the DG of adult mice treated with Cort or Cort + Flx (3wks). ( N ) Quantification of Bdnf mRNA in the DG per mouse (t-test, n= 4 (Cort), 4 (Cort + Flx 3wks)). ( O ) Representative image of BDNF protein expression in the hippocampus. Inset shows higher magnifications of BDNF overlapped with ZNT+ mossy fiber terminals. ( P ) BDNF protein levels in the hilar mossy fiber pathway of adult mice treated with Cort or Cort + Flx (3wks). ( Q ) Quantification of BDNF protein levels in the hilar mossy fiber pathway per mouse (One-way ANOVA, Tukey’s multiple comparisons test, n=4 mice/group). ( R ) Schematic for genetic and pharmacological approach to sparsely label mossy fiber terminals (left) and representative images of mossy terminals (right). ( S ) Quantification of mossy fiber terminal numbers per mouse in adult mice treated with Cort or Cort + Flx (3wks) (t-test, n= 4 mice/group). ( T ) Quantification of mossy fiber terminal area as cumulative distribution in adult mice treated with Cort or Cort + Flx (3wks) (Kolmogorov-Smirnov test, n= 200 terminals (Cort), 245 (Cort + Flx 3wks)). Data = mean ± SD. * p < 0 . 05 ; ** p < 0 . 01 ; *** p < 0 . 001 ; **** p < 0 . 0001 .

Article Snippet: Antigen retrieval for SOX11 (Sigma, MABE1929) and PROX1 (R&D Systems, AF2727) immunostaining was performed with the same procedure, but with a 10 minute incubation instead.

Techniques: Expressing

Journal: bioRxiv

Article Title: Antidepressants promote developmental-like plasticity through remodeling of extracellular matrix

doi: 10.1101/2025.01.03.631260

Figure Lengend Snippet: ( A ) Analysis of single cell RNA-seq dataset of the dentate gyrus from P5 and P132 mice showing unsupervised clustering analysis and identification of granule cells. ( B ) Volcano plot of differentially expressed genes between P5 versus P132 granule cells from (A). ( C ) Immature granule cell and axon regeneration scores defined as upregulated genes in 4-versus 8-week old neurons generated from adult hippocampal neurogenesis and regenerating versus non-regenerating spinal cord neurons . ( D ) Feature plot showing average expression of immature granule cell score (left) and quantification per mouse (right; One-way ANOVA, Tukey’s post-hoc test, n=3 mice/group). ( E ) Feature plot showing average expression of regeneration associated gene score (left) and quantification per mouse (right; One-way ANOVA, Tukey’s post-hoc test, n=3 mice/group). ( F ) Schematic for pharmacological and electroconvulsive shock approach for snRNA-seq of hippocampal nuclei (Dataset 2; see Methods ). ( G ) Unsupervised clustering of dentate granule cells from dataset 2 showing expression of Dock10 and module scores from and . ( H ) Quantification of module scores per mouse from dataset 2 (right; One-way ANOVA, Tukey’s post-hoc test, n=3 mice/group). ( I ) Normalized expression of Sox11 and Bdnf in mature granule cells corresponding to . ( J ) Proportion of DCX+ and DCX-neurons in the SOX11+ PROX1+ granule cell population in adult mice. ( K ) Representative image of SV2C levels in the hippocampus. Inset shows higher magnifications of SV2C overlapped with ZNT+ mossy fiber terminals. ( L ) SV2C protein levels in the hilar mossy fiber pathway of adult mice treated with Cort or Cort + Flx (3wks) and quantification of SV2C density per mouse (One-way ANOVA, Tukey’s multiple comparisons test, n=4 mice/group). Data = mean ± SD. * p < 0 . 05 ; ** p < 0 . 01

Article Snippet: Antigen retrieval for SOX11 (Sigma, MABE1929) and PROX1 (R&D Systems, AF2727) immunostaining was performed with the same procedure, but with a 10 minute incubation instead.

Techniques: RNA Sequencing Assay, Generated, Expressing

Journal: bioRxiv

Article Title: Breast Cancer Remodels Lymphatics in Sentinel Lymph Nodes

doi: 10.1101/2024.12.30.630756

Figure Lengend Snippet: a, UMAP plot of LECs from distant LNs (left) or metastatic LNs (right), color coded by cluster. b, Proportion of each LEC subset in distant and metastatic LNs, calculated from nine patients LECs (two-tailed, paired Student’s t-test), *p <0.05. c, Differential abundance testing using miloR. Neighborhoods are colored by their log fold abundance change between distant (blue) and metastatic (red) LNs. Non-differential abundance neighborhoods (FDR > 10%) are colored white. d, Beeswarm plot of the cell subset distribution of log fold change between normal and distant LNs. e, The trajectories of LN LEC differentiation shown in a UMAP plot. Six distinct LEC trajectories (T1-T6) were identified using Monocle single-cell trajectory analysis. f, Immunostaining of cancer cells and LEC markers PROX1 and MARCO in non-metastatic (left) and metastatic LNs (right). Zoomed-in images displaying SCS containing cytokeratin + cancer cells (left) and medullary sinuses (MS) both with and without cancer cells (right). A cancer cell in the MARCO + sinus is indicated by an arrow. Blue, cytokeratin; red, PROX1; green, MARCO. Scale bars, 500 μm. Images are representatives of two individuals with similar results. g, Immunostaining of CD200 + lymphatics and its quantification in metastatic (upper, black background) and non-metastatic (lower, grey background) LNs. CD200 + lymphatics and individual cancer cells within lymphatics are indicated by arrows and arrowheads, respectively. Blue, cytokeratin; green, PROX1; red, CD200. Scale bars, 500 μm. Images are representatives of seven individuals with simar results. SCS: subcapsular sinus; MS: medullary sinus; B: B cell zone. Circles in the bar plots represent biological replicates (mean±SEM, two-tailed, unpaired Student’s t-test). **p <0.005.

Article Snippet: The following primary antibodies were used: goat anti-human PROX1 (R&D Systems AF2727), rabbit anti-human MARCO (Atlas Antibodies, HPA063793), AF488 mouse anti-pan-cytokeratin (Thermo Fisher Scientific, MA5-18156), mouse anti-human MGP (Novus Biologicals, NBP2-45844), and BV421 mouse anti-human CD200 (Biolegend, 329209).

Techniques: Two Tailed Test, Immunostaining

Journal: bioRxiv

Article Title: Breast Cancer Remodels Lymphatics in Sentinel Lymph Nodes

doi: 10.1101/2024.12.30.630756

Figure Lengend Snippet: a, Differential abundance testing using MiloR and a heatmap of differentially expressed genes between differential abundance neighborhoods in LN LECs. The UMAP plot was shown in and is inserted here for clarity. In the heatmap, columns and rows represent neighborhoods and differentially expressed genes, respectively. Expression values for each gene are scaled between 0 and 1. The upper panel of the heatmap shows the neighborhood log fold change. FDR, false discovery rate. b, Heatmap showing the expression of the top DEGs between metastatic and distant LNs for each cell. Bars above the heatmap indicate the tissue and cluster origin of each cell (LNs, clusters). c, Violin plots displaying the top DEG expression between metastatic and distant LNs by cluster, with log-normalized expression value labeled. d, Immunostaining of MGP and its quantification in metastatic and distant LNs. Zoomed-in images show SCS and medullary sinuses containing cancer cells and MGP expression on PROX1 + LECs (arrows). Blue, cytokeratin; red, MGP; green, PROX1. Scale bars, 125 μm. Images are representative of four individuals with similar results. Circles in the bar plots represent biological replicates (mean±SEM, two-tailed, unpaired Student’s t-test). *p <0.05. e, Gene Ontology (GO) enrichment analysis of the top DEGs between metastatic and distant LNs using EnrichR package.

Article Snippet: The following primary antibodies were used: goat anti-human PROX1 (R&D Systems AF2727), rabbit anti-human MARCO (Atlas Antibodies, HPA063793), AF488 mouse anti-pan-cytokeratin (Thermo Fisher Scientific, MA5-18156), mouse anti-human MGP (Novus Biologicals, NBP2-45844), and BV421 mouse anti-human CD200 (Biolegend, 329209).

Techniques: Expressing, Labeling, Immunostaining, Two Tailed Test

Journal: eLife

Article Title: Increasing adult-born neurons protects mice from epilepsy

doi: 10.7554/eLife.90893

Figure Lengend Snippet: ( A ) Representative examples of hilar Prox1-ir in Cre- (1) and Cre+ (2) mice are shown. The boxes in a are expanded in b. Arrows point to hilar Prox1-ir cells, corresponding to hilar ectopic GCs. Calibration, 100 μm (a); 50 μm (b). ( B ) Prox1-ir is shown, within a hilar region of interest (ROI). The area of the ROI above the threshold, relative to the area of the ROI, is red. This area is called the area fraction, and was used to quantify hilar Prox1-ir. Calibration, 100 μm. ( C ) (1) Cre+mice had more hilar Prox1-ir cells than Cre- mice (t-test, p<0.001). (2) When sexes were divided, Cre+ mice had more hilar Prox1-ir cells than Cre- mice in both female (two-way ANOVA followed by Tukey’s post-hoc test, p<0.001) and male mice (p=0.001). ( D ) Correlations between hilar Prox1-ir cells and measurements of chronic seizures. (1) All Cre- and Cre- mice were compared regardless of sex. For the Cre- mice there was a significant inverse correlation between the # of Prox1-ir cells and # of chronic seizures (R 2 =0.296). Thus, the more Prox1-ir cells there were, the fewer chronic seizures there were. However, that was not true for Cre+mice (R 2 =0.072). (2) There was an inverse correlation between the number of hilar Prox1-ir cells and the seizure-free interval for Cre+ mice (R 2 =0.467) but not Cre- mice (R 2 =0.008). Thus, the more hilar Prox1-ir cells there were, the shorter the seizure-free periods were. However, this was not true for Cre- mice. (3) When data were divided by genotype and sex there was no significant correlation between hilar Prox1-ir cells and # of seizures (Cre- F, R 2 =0.0035; Cre+ F, R 2 =0.043; Cre- M, R 2 =0.104; Cre+ M, R 2 =0.083). (4) When data were divided by genotype and sex, there was a significant inverse correlation for the # of hilar Prox1-ir cells and seizure-free interval, but only for male Cre+ mice (R 2 =0.704). Cre+ females showed a trend (R 2 =0.395) and Cre- mice did not (Cre- F, R 2 =0.007, Cre- M, R 2 =0.046).

Article Snippet: anti-Prox1 (goat polyclonal) , 1:2000 , R and D Systems Cat# AF2727, RRID: AB_2170716 , Donkey anti-goat, Alexa Fluor 546 , 1:500 , Thermo Fisher Scientific Cat# A-11056, RRID: AB_2534103.

Techniques:

Journal: eLife

Article Title: Increasing adult-born neurons protects mice from epilepsy

doi: 10.7554/eLife.90893

Figure Lengend Snippet: ( A ) GluR2/3 hilar cells lacked Prox1 expression. (1) Cre- female mouse. Left: several GluR2/3+ cells (green) are located in the hilus within the box (marked by dotted white lines). Calibration, 70 µm. Right: The area within the box in the left panel is expanded. The merged image of GluR2/3+ (green) and Prox1+ (red) cells shows no double labeling. Calibration, 40 µm. (2) Cre+ female mouse. Similar results are shown for the Cre- mouse. White arrows mark ectopic GCs. Calibrations are the same as for the Cre- mouse. ( B ) A comparison of dorsal and ventral measurements for Cre- and Cre+ male mice shows no significant genotype effects. (1) GluR2/3. A two-way ANOVA showed no effect of dorsal or ventral location (F(1,13)=3.38; p=0.089) or genotype (F(1,13)=1.158; p=0.302). (2) SOM. A two-way ANOVA showed no effect of dorsal or ventral location (F(1,10)=0.172; p=0.687) or genotype (F(1,10)=0.014; p=0.908). (3) Parvalbumin. A two-way ANOVA showed no effect of dorsal or ventral location (F(1,13)=0.358; p=0.560) or genotype (F(1,13)=1.068 p=0.320). ( C ) A comparison of ventral measurements for both Cre- and Cre+ female and male mice. (1) There were significantly more GluR2/3+hilar cells in Cre+ female mice compared to Cre- female mice, like the dorsal hippocampus . Thus, GluR2/3+hilar cells were spared in Cre+ females in dorsal and ventral hippocampus. A two-way ANOVA showed o effect of sex (F(1,18)=0.744; p=0.400) or genotype (F(1,18)=0.386; p=0.542) but there was a significant interaction F(1,18)=5.433; p=0.0316, and post-hoc tests showed that Cre+ females had significantly more GluR2/3+ cells than Cre- females (p=0.045). (2) There were no significant differences among groups for SOM+ cells. Thus, there was an effect in dorsal but not ventral hippocampus. Also, SOM cells were spared in Cre+ females dorsally but not ventrally. A two-way ANOVA showed no effect of sex (F(1,17)=0.718; p=0.408) or genotype (F (1, 17)=0.769; p=0.393). (3) There were no significant differences in numbers of parvalbumin+ cells, like dorsal hippocampus . Thus, parvalbumin cells were similar regardless of genotype in dorsal and ventral hippocampus. A two-way ANOVA showed no significant effect of sex (F(1,16)=0.401; p=0.536) or genotype (F(1,16)=0.221; p=0.645).

Article Snippet: anti-Prox1 (goat polyclonal) , 1:2000 , R and D Systems Cat# AF2727, RRID: AB_2170716 , Donkey anti-goat, Alexa Fluor 546 , 1:500 , Thermo Fisher Scientific Cat# A-11056, RRID: AB_2534103.

Techniques: Expressing, Labeling, Comparison

Journal: eLife

Article Title: Increasing adult-born neurons protects mice from epilepsy

doi: 10.7554/eLife.90893

Figure Lengend Snippet: Key resources.

Article Snippet: anti-Prox1 (goat polyclonal) , 1:2000 , R and D Systems Cat# AF2727, RRID: AB_2170716 , Donkey anti-goat, Alexa Fluor 546 , 1:500 , Thermo Fisher Scientific Cat# A-11056, RRID: AB_2534103.

Techniques: Plasmid Preparation

Journal: eLife

Article Title: Increasing adult-born neurons protects mice from epilepsy

doi: 10.7554/eLife.90893

Figure Lengend Snippet: Key resources.

Article Snippet: anti-Prox1 (goat polyclonal) , 1:2000 , R and D Systems Cat# AF2727, RRID: AB_2170716 , Donkey anti-goat, Alexa Fluor 546 , 1:500 , Thermo Fisher Scientific Cat# A-11056, RRID: AB_2534103.

Techniques: Plasmid Preparation

Journal: bioRxiv

Article Title: Differential contribution of P73 + Cajal-Retzius cells and Reelin to cortical morphogenesis

doi: 10.1101/2024.10.15.618167

Figure Lengend Snippet: (A) Drawing indicating the regions shown in B-L (top) and color key for the panel shown in B. (B) Immunostaining for Reln and Tomato in the neocortex or hippocampus of P1 control or Reln cKO animals. (C) Immunostaining for Reln in control and cKO showing the decreased Reln expression, in the dorsolateral cortex and hippocampus. (D) Quantification of the fraction of Reln + cells among Tomato + CRs in the dorsal cortex and hippocampus. Each point corresponds to one animal. (E) Quantification of the density of Tomato + CRs in the dorsal cortex MZ of control and cKO mice. Each point corresponds to one animal. (F) Density of Tomato + CRs along the medio-lateral axis of the cortical MZ (n=3 animals per genotype). (G) Quantification of the fluorescence intensity of Reln (arbitrary units) along the HF and cortical MZ. The dashed lines represent each animal considered (n=3 per genotype). (H) Immunostaining for Tbr1and Brn2, or for CTIP2 and Tomato in a cortical column of the presumptive somatosensory cortex of P1 control and Reln cKO . (I) In situ hybridization for Tbr1 and Rorb in a rostral section of the cortex from P1 control, Reln cKO and PGK Cre ;Reln lox/- mice. (J) Immunostaining for CTIP2 in a cortical column of the presumptive somatosensory cortex of P1 control and PGK Cre ;Reln lox/- mice. (K) In situ hybridization for Nr4a2 in the lateral cortex of P1 control, Reln cKO and PGK Cre ;Reln lox/- mice. CLA: claustrum. (L) DAPI staining of nuclei in the rostromedial cortex of P1 control, Reln cKO and PGK Cre ;Reln lox/- mice. Arrows point to cells abnormally positioned in the MZ of mutants. (M) Immunostaining for Foxg1 and Tomato along the medio-lateral axis of the caudal cortex of P1 control or Reln cKO . (N) Immunostaining for CTIP2 and Prox1 in the P1 hippocampus of control and Reln cKO . (O) Immunostaining for CTIP2 and Prox1 or Laminin in the P1 hippocampus of control and PGK Cre ;Reln lox/- mice. The arrow points to the defect in compaction of the CA1 pyramidal layer. HF: hippocampal fissure. Scale bars: 50µm in B, K. 100µm in F, H. 200µm in G, J, L, M. 500µm in I, K.

Article Snippet: The following primary antibodies were used: goat anti-Brn2 (POU3F2, Abcam ab101726 1:1000), rat anti-CTIP2 (BCL11B, Abcam ab18465 1:600), rabbit anti-FOXG1 (Abcam ab18259 1:2000), rabbit anti-Laminin (Sigma-Aldrich L9393 1:600), goat anti-Neuropilin-1 (R&D Systems AF566 1:800), rabbit anti-p73 (Cell signaling 14620 1:250), goat anti-Nurr1 (NR4A2, R&D Systems AF2156 1:200), goat anti-Prox1 (R&D Systems AF2727 1:1000), goat anti-Reelin (R&D Systems AF3820 1:2000), rabbit anti-TBR1 (Abcam ab31940 1:1000).

Techniques: Immunostaining, Control, Expressing, Fluorescence, In Situ Hybridization, Staining

Journal: bioRxiv

Article Title: Differential contribution of P73 + Cajal-Retzius cells and Reelin to cortical morphogenesis

doi: 10.1101/2024.10.15.618167

Figure Lengend Snippet: (A) Drawing indicating the region shown in B-E (top) and region specificity of the markers used (bottom). (B) Immunostaining for CTIP2 on coronal sections of the hippocampus from E16 to P1 in control and Gmnc -/- animals. DG: dentate gyrus anlage. (C) Immunostaining for EdU on sections of the hippocampus at P2 in control and Gmnc mutants following EdU incorporation at E15. SO: stratum oriens , SP: stratum pyramidale . The yellow dashed line separates the regions considered normal and folded in mutants. Quantification of EdU labelled cells in control and mutant (n=3 each), considering normal or folded regions of CA1. (D) Immunostaining for Laminin (green) on coronal sections of the hippocampus in E18. HF: hippocampal fissure, only distinguishable in controls. (E) In situ hybridization for Nr4a2 and Zbtb20 , and immunostaining for Prox1 on coronal sections of the hippocampus at P1 in control and Gmnc -/- animals. Scale bars: 200µm in B, C, E. 50µm in D.

Article Snippet: The following primary antibodies were used: goat anti-Brn2 (POU3F2, Abcam ab101726 1:1000), rat anti-CTIP2 (BCL11B, Abcam ab18465 1:600), rabbit anti-FOXG1 (Abcam ab18259 1:2000), rabbit anti-Laminin (Sigma-Aldrich L9393 1:600), goat anti-Neuropilin-1 (R&D Systems AF566 1:800), rabbit anti-p73 (Cell signaling 14620 1:250), goat anti-Nurr1 (NR4A2, R&D Systems AF2156 1:200), goat anti-Prox1 (R&D Systems AF2727 1:1000), goat anti-Reelin (R&D Systems AF3820 1:2000), rabbit anti-TBR1 (Abcam ab31940 1:1000).

Techniques: Immunostaining, Control, Mutagenesis, In Situ Hybridization