Journal: bioRxiv

Article Title: Supracellular Mechanics and Counter-Rotational Bilateral Flows Orchestrate Posterior Morphogenesis

doi: 10.1101/2025.11.18.689090

Figure Lengend Snippet: (A-B) 2D-projected confocal images of the near-blastopore region (orange arrowhead) at stages 18 and 24. Schematics indicates approximate imaging locations and plane (Blue shaded box). (A) En face views of F-actin, keratin 8, and fibronectin and fibronectin in parasagittal section. The sagittal section is rotated 90° to align the blastopore with en face images. The regions of tissue rotation are located approximately 100 to 150 µm from the center of the blastopore. (B) En face views of keratin 8 and fibronectin, with a zoomed inset (yellow box). Aligned fibronectin fibrils are most prominent within 80 to 120 µm of the blastopore. (C) Polar histograms (rose plots) of fiber orientations at stage 18 and 24 (n, number of embryos used; Square goodness-of-fit test, ∗∗∗∗p<0.0001) (D) Workflow for image processing and analysis of immunostained samples. (E) Fibronectin morphological features from stage 18 and stage 24. (C and E) measured within a 100 by 100 µm region ventral to the blastopore. Each symbol represents the mean value per embryo (Mann-Whitney U, ∗p=0.02). Bars; mean ± 95% CI. (A-B) scale bars, 20 µm. (A-B) Xenopus illustrations © Natalya Zahn (2022).

Article Snippet: Immunofluorescence staining was carried out with primary antibodies against fibrillin-2 (JB3, Developmental Studies Hybridoma Bank; 1:200), collagen-2 (II-II6B3, Developmental Studies Hybridoma Bank; 1:100), laminin-a1 (L9393, Sigma-Aldrich; 1:500), mouse anti-fibronectin monoclonal antibody (4H2, courtesy of Douglas DeSimone, University of Virginia, Charlottesville, VA, USA; 1:500), rabbit anti-fibronectin polyclonal antibody (F3648, Sigma; 1:200), acetylated tubulin (T6793; Sigma; 1:125), keratin (1h5; Developmental Studies Hybridoma Bank; 1:500), and GFP (N86/8; Developmental Studies Hybridoma Bank; 1:50) and incubated overnight on a nutator at 4 °C.

Techniques: Imaging, MANN-WHITNEY

Journal: bioRxiv

Article Title: Supracellular Mechanics and Counter-Rotational Bilateral Flows Orchestrate Posterior Morphogenesis

doi: 10.1101/2025.11.18.689090

Figure Lengend Snippet: (A) Dorso-anterior index (DAI) of the severity of dorsalized and ventralized phenotypes. (B) Representative phenotypes of each treatment at stage 20, showing characteristic morphological differences between dorsalized, ventralized, and control embryos. (C) Final frame of brightfield timelapse sequence, overlaid with a yellow deformation map (see also Video S6). (D) Vorticity overlaid randomized dot plots deformed by calculated displacements, and max-projected across all timepoints to visualize movement patterns. Representative images are shown for each treatment. (E) SWIRL predicted vortices highlight vortex structure and organization. (F-G) Quantitative comparison of predicted vortex characteristics in dorsalized (8 vortices from 11 embryos), control (32 vortices from 19 embryos), and ventralized (11 vortices from 11 embryos) embryos. Each symbol represents a single predicted vortex (Mann-Whitney U, ∗∗p=0.0012; ∗∗p=0.0061; ∗∗∗p=0.0002; ∗∗∗∗p<0.0001). (F) Vortex compactness (mean ± 95% CI). (G) Vortex swirling strength (mean ± 95% CI). (H) Fibronectin networks in posterior tissues from stage 24 embryos within a 100 by 100 µm region ventral to the blastopore. (I) Fibronectin orientation frequency in each treatment (n, number of embryos used; Square goodness-of-fit test, ∗∗∗∗p<0.0001) (J) Morphological features of fibronectin network in dorsalized, control, and ventralized embryos. Each symbol represents the mean value per embryo (Mann-Whitney U, ∗p=0.0426; ∗∗∗∗ p < 0.0001). Bars indicate mean ± 95% CI. (B-C) scale bars, 100 µm; 20 µm in (H). (A) DAI diagram with permission of the publisher. Xenopus illustrations © Natalya Zahn (2022).

Article Snippet: Immunofluorescence staining was carried out with primary antibodies against fibrillin-2 (JB3, Developmental Studies Hybridoma Bank; 1:200), collagen-2 (II-II6B3, Developmental Studies Hybridoma Bank; 1:100), laminin-a1 (L9393, Sigma-Aldrich; 1:500), mouse anti-fibronectin monoclonal antibody (4H2, courtesy of Douglas DeSimone, University of Virginia, Charlottesville, VA, USA; 1:500), rabbit anti-fibronectin polyclonal antibody (F3648, Sigma; 1:200), acetylated tubulin (T6793; Sigma; 1:125), keratin (1h5; Developmental Studies Hybridoma Bank; 1:500), and GFP (N86/8; Developmental Studies Hybridoma Bank; 1:50) and incubated overnight on a nutator at 4 °C.

Techniques: Control, Sequencing, Comparison, MANN-WHITNEY

Journal: bioRxiv

Article Title: Supracellular Mechanics and Counter-Rotational Bilateral Flows Orchestrate Posterior Morphogenesis

doi: 10.1101/2025.11.18.689090

Figure Lengend Snippet: (A) Methods used for targeted disruption of fibronectin organization and integrin-fibronectin interactions. (B) Representative phenotypes showing morphological changes following fibronectin disruption. (C) 2D max-projected confocal image of fibronectin within a 100 by 100 µm region ventral to the blastopore. (D) Morphological features of fibronectin matrix for control (mAb 4H2) and function-blocking (mAb P8D4) treatments. Each symbol represents the per-embryo mean (Mann-Whitney U, ∗p=0.0350; ∗∗p=0.0023). Bars indicate mean ± 95% CI. (E) Final frame of brightfield timelapse sequence overlaid with yellow deformation map (see also Video S7). (F) Time-projected displacement of random dot plot overlaid with vorticity. Disruptions to fibronectin result in less distinct or absent bi-directional vortices compared to controls. (G) SWIRL predicted vortex structure and spatial distribution across treatments. (H-J) Vortex characteristics across treatments. Each symbol in (H) and (I) represents a single predicted vortex: 4H2 (19 vortices from 10 embryos), P8D4 (8 from 11), COMO (21 from 13), and FNMO (18 from 13). In (J), each symbol represents an embryo with a predicted vortex pair. Vortex formation was significantly disrupted in P8D4- and FNMO-treated embryos, with a reduction in detected vortices and alterations in vortex compactness (H) and swirling strength (I). (J) Vortex asymmetry index (Mann-Whitney U, ∗p<0.0290; ∗∗p=0.0054; ∗∗p=0.0024; ∗∗∗p=0.0002). Bars indicate mean ± 95% CI, except in (H). Scale bars, 100 µm in (B,E); 20 µm in (C).

Article Snippet: Immunofluorescence staining was carried out with primary antibodies against fibrillin-2 (JB3, Developmental Studies Hybridoma Bank; 1:200), collagen-2 (II-II6B3, Developmental Studies Hybridoma Bank; 1:100), laminin-a1 (L9393, Sigma-Aldrich; 1:500), mouse anti-fibronectin monoclonal antibody (4H2, courtesy of Douglas DeSimone, University of Virginia, Charlottesville, VA, USA; 1:500), rabbit anti-fibronectin polyclonal antibody (F3648, Sigma; 1:200), acetylated tubulin (T6793; Sigma; 1:125), keratin (1h5; Developmental Studies Hybridoma Bank; 1:500), and GFP (N86/8; Developmental Studies Hybridoma Bank; 1:50) and incubated overnight on a nutator at 4 °C.

Techniques: Disruption, Control, Blocking Assay, MANN-WHITNEY, Sequencing

Journal: Nature Communications

Article Title: Lymph nodes molecular subtypes unravel lymph nodes heterogeneity and clinical implications in colorectal cancer

doi: 10.1038/s41467-025-63200-z

Figure Lengend Snippet: Dot plot of PCA based on fibroblasts-specific ( A ) and endothelial cell-specific ( B ) gene markers to estimate their ability in distinguish the LNs subtypes ( n = 68). C Box plots illustrating the differences in COL1A1, α-SMA, FN1, CD31, and LYVE1 gene expression between LNs subtypes ( n = 50, 35, 31, 32, 11, and 14). Each data point represents an individual LN (biological replicates). Box plots represent the median (center line), 25th and 75th percentiles (bounds of the box), and minimum and maximum values (whiskers). The mIF staining and quantification reveal differential expression of α-SMA, FN1, COL1A1 ( n = 20 and 16) ( D ) and LYVE-1, CD31 ( n = 24 and 18) ( E ) between NLN_C1 and NLN_C4. Each data point represents an individual LN (biological replicates). Data are presented as mean values ± SD. The p value was calculated using a two-sided Student’s t test. Scale bars indicate 100 μm, 500 μm, and 1000 μm respectively. Source data are provided as a Source Data file. F Sirius Red staining results of differences in fibrosis levels between NLN_C1 and NLN_C4. Scale bars indicate 1000 μm. PC principal component, PCA principal component analysis, mIF multiplex immunofluorescence, SD standard deviation.

Article Snippet: The primary antibodies included rabbit polyclonal antibody CXCL13/BCA1 (Proteintech, 10927-1-AP); rabbit polyclonal antibody LYVE1 (Proteintech, 28321-1-AP); mouse monoclonal antibody CD20 (Proteintech, 60271-1-Ig); mouse monoclonal antibody FN1 (Proteintech, 66042-1-Ig); mouse monoclonal antibody CD31 (Proteintech, 66065-2-Ig); mouse monoclonal antibody COL1A1 (Proteintech, 67288-1-Ig); mouse monoclonal antibody PDPN (Proteintech, 67432-1-Ig); mouse monoclonal antibody CD35 (Proteintech, 68033-1-Ig); rabbit recombinant antibody α-SMA (Proteintech, 80008-1-RR); All images were captured using an EVOS FL Auto 2 Imaging System (Thermo Fisher Scientific).

Techniques: Gene Expression, Staining, Quantitative Proteomics, Multiplex Assay, Immunofluorescence, Standard Deviation

Journal: Development (Cambridge, England)

Article Title: Application and measurement of tissue-scale tension in avian epithelia in vivo to study multiscale mechanics and inter-germ layer coupling

doi: 10.1242/dev.204561

Figure Lengend Snippet: Mechanical coupling across germ layers. (A) Schematic of the embryonic region shown in B,C. Ventral view. A, anterior; ML, mediolateral; P, posterior. (B) Overlay of brightfield and fluorescent images prior to (top; applied strain E app =0) and after application of stretch to the endoderm (bottom; E app =0.6); endoderm nuclei are labeled by electroporation with pCAG-H2B-GFP (green). Magenta arrowheads and dashed yellow lines denote the positions of two nuclei and lateral somite boundaries, respectively. Scale bar: 100 µm. (C) Kymograph of stretch progressively applied to endoderm, illustrating concomitant deformation of somites over time. Scale bar: 100 µm. (D) Left: Schematic (top) and brightfield images (bottom) of the embryo following dorsal cuts to isolate ventral endoderm–somite interactions. Right: Comparison of Lagrangian strains in endoderm and mesoderm for control and dorsal cut embryos. ns, not significant ( P =0.52, Welch's t -test on the slopes of the strain transfer per sample). Circles represent the mean, error bars represent s.d. (E) Transverse view of fibronectin (FN; red) staining of the endoderm–somite interface of control (top) and Dispase-treated (bottom) embryos. DAPI was used to stain nuclei (gray). en, endoderm; no, notochord; so, somite. Scale bar: 10 µm. (F) Quantification of applied force versus in-plane Lagrangian strain in the endoderm for control and Dispase-treated embryos. (G) Relative stiffness quantified from the force-strain curves shown in F. * P =0.012 (Welch's t -test). (H) Comparison of Lagrangian strains in endoderm and mesoderm for control and Dispase-treated embryos. * P =0.012 (Welch's t -test on the slopes of the strain transfer per sample). Circles represent the mean, error bars represent s.d.

Article Snippet: Embryos were washed (PBS with 0.1% Triton X-100 was used for all washes) and left in blocking solution (10% heat inactivated goat serum) for 2 h, then incubated overnight at 4°C with mouse anti-fibronectin primary antibody (DSHB, B3/D6) diluted in PBS+0.1% Triton X-100 at 1:200.

Techniques: Labeling, Electroporation, Comparison, Control, Staining