Journal: Journal of Biomedical Science

Article Title: Many but not all pathogen-associated molecular patterns aggravate neurogenic heterotopic ossification after spinal cord injury

doi: 10.1186/s12929-026-01237-y

Figure Lengend Snippet: Expression of PRRs by mouse and human skeletal muscle cells. Expression of PRR mRNA transcripts in mouse satellite cells (SC), FAPs (FP), endothelial cells (EC) and monocyte/macrophages (MΦ) sorted from skeletal muscles. Whole mouse bone marrow (BM) cells and splenocytes (SP) were used as positive controls for these transcripts. Human PDGFRα + FAPs were sorted from muscles surrounding NHO biopsies, and CD14 + monocytes were isolated from peripheral blood from healthy donors. A TLR1, B TLR2, C TLR3, D TLR4, E TLR5, F TLR6, G TLR7, H TLR8, I TLR9, J STING1, K RIGI, L MDA5 (IFIH1), M PRK, N NOD1, O NOD2, P Dectin-1 (CLEC7A), Q Dectin-2 (Clec4n/CLEC6A), R Mincle (CLEC4E). For each gene, left hand histogram represents expression of the mouse gene in mouse cells and the right-hand histogram represents expression of the human gene in human cells. For mouse cells, relative mRNA expression was quantified relative to house-keeping gene Hprt . For human cells, values were normalized using three references genes HPRT , RPLP0 and PPIA for FAPs and ACTB , GAPDH and PPIA for CD14 + cells. Each dot represents a mouse or a human donor, bars represent mean ± SD

Article Snippet: Human MPCs were trypsinized and incubated for 30 min with biotinylated anti-human PDGFRα (Cat# BAF322, R&D Systems) goat polyclonal antibody and CD56-PE (clone B159, BD Pharmigen) monoclonal antibody in PBS 2% FCS, 2 mM EDTA or with control isotypes IgG1 PE (Cat# A07796, Beckman Coulter) and biotinylated goat IgG (Cat# BAF108, R&D Systems).

Techniques: Expressing, Muscles, Isolation

Journal: Journal of Biomedical Science

Article Title: Many but not all pathogen-associated molecular patterns aggravate neurogenic heterotopic ossification after spinal cord injury

doi: 10.1186/s12929-026-01237-y

Figure Lengend Snippet: OSM and IL-1 neutralization in monocyte-conditioned media strongly inhibit hFAPs mineralization. A OSM, IL-1α and IL-1β concentrations quantified in conditioned media from human CD14 + macrophages stimulated with 200 ng/ml Pam2CSK4 (CM Pam2CSK4 ), 200 ng/ml Pam3CSK4 (CM Pam3CSK4 ) or non-stimulated (CM ∅ ). B OSM, IL-1α and IL-1β concentrations in CM Pam2CSK4 , CM Pam3CSK4 and CM ∅ were correlated with hFAP calcium mineralization measured by Alizarin Red staining after 2 weeks of culture in osteogenic conditions in the presence of the conditioned media. Each dot represents a different conditioned medium sample. C–F Mouse anti-human OSM antibody, isotype control antibody and IL-1RA were used to neutralize OSM and IL-1 in human CD14 + monocyte conditioned media as indicated below each chart. Calcium mineralization of hFAPs cultured for 2 weeks in osteogenic conditions with C CM Pam2CSK4 or D CM Pam3CSK4 was measured using Alizarin Red staining and quantified by spectrophotometry. E, F RUNX2 protein quantification by Western blot using Stain-Free normalization for hFAPs cultured for 2 weeks in osteogenic conditions with E CM Pam2CSK4 or F CM Pam3CSK4 . In C-F, each dot represents a hFAP individual donor. Bars represent mean ± SD, One-way ANOVA Dunnett’s multiple comparison test versus CM ∅ in ( A ) and ( B ), versus CM Pam2CSK4 in ( C ) and ( E ), or versus CM Pam3CSK4 in ( D ) and ( F ), * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: Human MPCs were trypsinized and incubated for 30 min with biotinylated anti-human PDGFRα (Cat# BAF322, R&D Systems) goat polyclonal antibody and CD56-PE (clone B159, BD Pharmigen) monoclonal antibody in PBS 2% FCS, 2 mM EDTA or with control isotypes IgG1 PE (Cat# A07796, Beckman Coulter) and biotinylated goat IgG (Cat# BAF108, R&D Systems).

Techniques: Neutralization, Staining, Control, Cell Culture, Spectrophotometry, Western Blot, Comparison

Journal: Developmental Cell

Article Title: Lumen Expansion Facilitates Epiblast-Primitive Endoderm Fate Specification during Mouse Blastocyst Formation

doi: 10.1016/j.devcel.2019.10.011

Figure Lengend Snippet: EPI and PrE Expression Levels Are Reduced in ATP1-Inhibited Embryos (A) Immunofluorescence images of TE (Cdx2), EPI (Sox2), and PrE (Gata4) fate in pre-treatment control (E3.5 WT), Atp1 inhibited (E4.0 500 μM and E4.0 250 μM), and end-stage control (E4.0 DMSO) embryos. Lumen boundaries outlined by dashed white line and mean lumen volume in white text. Scale bars, 10 μm. (B) Boxplot of lumen volume for E3.5 WT (N = 21), E4.0 DMSO (N = 24), E4.0 250 μM Atp1 inhibited (N = 14) and E4.0 500 μM Atp1 inhibited (N = 31) embryos indicating that the impact on lumen volume is concentration dependent. (C) Boxplot of fluorescence levels of Cdx2 (gray), Sox2 (green), and Gata4 (magenta) in E4.0 500 μM Atp1 inhibited embryos compared to E4.0 DMSO controls. (D) Schematic 2D representation of 3D analysis method for spatial segregation of ICM lineages. P 1,2,3,4 are 3D points. L ↔ is a 3D line ( P 1 P 2 ↔ equivalent) that defines the embryonic-abembryonic axis. d ‾ is the 3D line segment ( P 3 P 4 ‾ equivalent) that measures the perpendicular distance from the center of a cell to L ↔ . See Image Analysis for formal definitions of all geometric entities. (E) Boxplot of spatial overlap between EPI and PrE lineages within E4.0 control (DMSO, N = 15), E4.0 Atp1 inhibited (500 μM, N =13) and simulated data of maximal overlap in E4.0 WT embryos (Simulation, N = 27). ∗∗∗∗ p < 0.0001, ∗∗∗ p < 0.001, ∗∗ p < 0.01, ∗ p < 0.05. n.s., not significant. For boxplots: central mark indicates the median; lower edge, 25%; upper edge, 75%; lower whisker, Q1 − (1.5 × IQR), where IQR = Q3 − Q1; upper whisker, Q3 + (1.5 × IQR). See also and ; .

Article Snippet: The following primary antibodies were used in this study: rabbit anti-pERM (Cell Signaling, 3726), mouse anti-Cdx2 (BioGenex, MU392A-UC), goat anti-Sox2 (Santa Cruz Biotechnology, sc-17320), goat anti-Sox2 (R&D Systems, AF2018-SP), rabbit anti-Sox2 (Cell Signaling, 23064), rabbit anti Gata4 (Santa Cruz Biotechnology, sc-9053), goat anti-Gata4 (R&D Systems, AF2606-SP), rabbit anti-GFP (MBL, 598), mouse anti-Rab11 (BD Biosciences; 610656), rat anti-integrin-β1 (Merck Millipore, MAB1997), goat anti-biotinylated Gata4 (R&D Systems, BAF2606) mouse anti-Oct3/4 (Santa Cruz Biotechnology, sc-5279) and mouse anti-Hsp47 (Enzo Life Sciences, M16.10A1).

Techniques: Expressing, Immunofluorescence, Control, Concentration Assay, Fluorescence, Whisker Assay

Journal: Developmental Cell

Article Title: Lumen Expansion Facilitates Epiblast-Primitive Endoderm Fate Specification during Mouse Blastocyst Formation

doi: 10.1016/j.devcel.2019.10.011

Figure Lengend Snippet: PrE Specification and Spatial Segregation of ICM Lineages Is Impaired by Mechanical Inhibition of Lumen Expansion (A) Brightfield images of mechanical deflation. Magenta asterisk marks the needle tip. Dotted magenta line indicates lumen boundary. (B) Immunofluorescence images of EPI (Sox2) and PrE (Gata4) fate in pre-manipulation control (E3.5 WT), E4.0 post-manipulation control (E4.0 WT), and E4.0 mechanically inhibited (E4.0 Mechanical) embryos. Magenta arrowheads indicate the position of cells expressing high levels of Gata4 within the ICM. White dotted line indicates lumen boundaries. Average lumen volume in white text. Scale bars, 10 μm. (C) Boxplot of fluorescence levels of Sox2 (green) and Gata4 (magenta) in mechanically inhibited (Mech., N = 33) and post-manipulation control (WT, N = 28) E4.0 embryos. (D) Boxplot of spatial overlap between EPI and PrE lineages within post-manipulation control (WT, N = 27), mechanically inhibited (Mech., N = 33), E4.0 procedural control (Control, N = 11), and E4.0 simulation of complete overlap in WT conditions (Simulation, N = 27). ∗∗∗∗ p < 0.0001, ∗∗ p < 0.01. n.s., not significant. For boxplots: central mark indicates the median; lower edge, 25%; upper edge, 75%; lower whisker, Q1 − (1.5 × IQR), where IQR = Q3 − Q1; upper whisker, Q3 + (1.5 × IQR). See also Figure S6 and .

Article Snippet: The following primary antibodies were used in this study: rabbit anti-pERM (Cell Signaling, 3726), mouse anti-Cdx2 (BioGenex, MU392A-UC), goat anti-Sox2 (Santa Cruz Biotechnology, sc-17320), goat anti-Sox2 (R&D Systems, AF2018-SP), rabbit anti-Sox2 (Cell Signaling, 23064), rabbit anti Gata4 (Santa Cruz Biotechnology, sc-9053), goat anti-Gata4 (R&D Systems, AF2606-SP), rabbit anti-GFP (MBL, 598), mouse anti-Rab11 (BD Biosciences; 610656), rat anti-integrin-β1 (Merck Millipore, MAB1997), goat anti-biotinylated Gata4 (R&D Systems, BAF2606) mouse anti-Oct3/4 (Santa Cruz Biotechnology, sc-5279) and mouse anti-Hsp47 (Enzo Life Sciences, M16.10A1).

Techniques: Inhibition, Immunofluorescence, Control, Expressing, Fluorescence, Whisker Assay

Journal: Developmental Cell

Article Title: Lumen Expansion Facilitates Epiblast-Primitive Endoderm Fate Specification during Mouse Blastocyst Formation

doi: 10.1016/j.devcel.2019.10.011

Figure Lengend Snippet: Perturbation of FGF4 Signaling in the Lumen Impacts Molecular Specification of EPI and PrE Lineages (A) Immunofluorescence images of EPI (Sox2) and PrE (Gata4) fate in E3.5 post-FGF4 deposition (E3.5 FGF4 Inj.), E3.5 post-PD173074 deposition (E3.5 PD Inj.), and E3.5 post-PBS deposition (E3.5 PBS Inj.). White dotted line indicates lumen boundaries. Average lumen volume in white text. Scale bars, 10 μm. (B) Boxplot of fluorescence levels of Sox2 (green) and Gata4 (magenta) in E3.5 post-FGF4 deposition (FGF4 Inj., N = 24), E3.5 post-PD173074 deposition (PD Inj., N = 21), and E3.5 post-PBS deposition (PBS Inj., N = 13) embryos. (C) Boxplot of luminal volume in E3.5 post-FGF4 deposition (FGF4 Inj., N = 24), E3.5 post-PD173074 deposition (PD Inj., N = 21), and E3.5 post-PBS deposition (PBS Inj., N = 13). ∗∗∗∗ p < 0.0001, ∗∗ p < 0.01. n.s., not significant. For boxplots: central mark indicates the median; lower edge, 25%; upper edge, 75%; lower whisker, Q1 − (1.5 × IQR) where IQR = Q3 − Q1; upper whisker, Q3 + (1.5 × IQR). See also Figure S7 .

Article Snippet: The following primary antibodies were used in this study: rabbit anti-pERM (Cell Signaling, 3726), mouse anti-Cdx2 (BioGenex, MU392A-UC), goat anti-Sox2 (Santa Cruz Biotechnology, sc-17320), goat anti-Sox2 (R&D Systems, AF2018-SP), rabbit anti-Sox2 (Cell Signaling, 23064), rabbit anti Gata4 (Santa Cruz Biotechnology, sc-9053), goat anti-Gata4 (R&D Systems, AF2606-SP), rabbit anti-GFP (MBL, 598), mouse anti-Rab11 (BD Biosciences; 610656), rat anti-integrin-β1 (Merck Millipore, MAB1997), goat anti-biotinylated Gata4 (R&D Systems, BAF2606) mouse anti-Oct3/4 (Santa Cruz Biotechnology, sc-5279) and mouse anti-Hsp47 (Enzo Life Sciences, M16.10A1).

Techniques: Immunofluorescence, Fluorescence, Whisker Assay

Journal: Developmental Cell

Article Title: Lumen Expansion Facilitates Epiblast-Primitive Endoderm Fate Specification during Mouse Blastocyst Formation

doi: 10.1016/j.devcel.2019.10.011

Figure Lengend Snippet: Luminal Deposition of FGF4 Partially Rescues EPI-PrE Specification in ATP1-Inhibited Embryos (A) Immunofluorescence images of EPI (Sox2) and PrE (Gata4) fate in E3.5 post-FGF4 deposition and Atp1 inhibition (E3.5 FGF4 250 μM), E3.5 post-PBS deposition and Atp1 inhibition (E3.5 PBS 250 μM), and E3.5 control embryos (E3.5 DMSO). White dotted line indicates lumen boundaries. Average lumen volume in white text. Magenta arrowhead indicates cell with high Gata4 expression relative to neighboring cells. Scale bars, 10 μm. (B) Boxplot of fluorescence levels of Sox2 (green) and Gata4 (magenta) in E3.5 post-FGF4 deposition and Atp1 inhibition (250 μM FGF4 Inj., N = 42 embryos), E3.5 post-PBS deposition and Atp1 inhibition (250 μM PBS Inj., N = 12), and E3.5 control embryos (DMSO, N = 12). (C) Boxplot of luminal volume in E3.5 post-FGF4 deposition and Atp1 inhibition (250 μM FGF4 Inj., N = 49 embryos), E3.5 post-PBS deposition and Atp1 inhibition (250μM PBS Inj., N = 9), and E3.5 control embryos (DMSO, N = 38). ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗∗ p < 0.0001. n.s., not significant. For boxplots: central mark indicates the median; lower edge, 25%; upper edge, 75%; lower whisker, Q1 − (1.5 × IQR), where IQR = Q3 − Q1; upper whisker, Q3 + (1.5 × IQR).

Article Snippet: The following primary antibodies were used in this study: rabbit anti-pERM (Cell Signaling, 3726), mouse anti-Cdx2 (BioGenex, MU392A-UC), goat anti-Sox2 (Santa Cruz Biotechnology, sc-17320), goat anti-Sox2 (R&D Systems, AF2018-SP), rabbit anti-Sox2 (Cell Signaling, 23064), rabbit anti Gata4 (Santa Cruz Biotechnology, sc-9053), goat anti-Gata4 (R&D Systems, AF2606-SP), rabbit anti-GFP (MBL, 598), mouse anti-Rab11 (BD Biosciences; 610656), rat anti-integrin-β1 (Merck Millipore, MAB1997), goat anti-biotinylated Gata4 (R&D Systems, BAF2606) mouse anti-Oct3/4 (Santa Cruz Biotechnology, sc-5279) and mouse anti-Hsp47 (Enzo Life Sciences, M16.10A1).

Techniques: Immunofluorescence, Inhibition, Control, Expressing, Fluorescence, Whisker Assay

Journal: Developmental Cell

Article Title: Lumen Expansion Facilitates Epiblast-Primitive Endoderm Fate Specification during Mouse Blastocyst Formation

doi: 10.1016/j.devcel.2019.10.011

Figure Lengend Snippet:

Article Snippet: The following primary antibodies were used in this study: rabbit anti-pERM (Cell Signaling, 3726), mouse anti-Cdx2 (BioGenex, MU392A-UC), goat anti-Sox2 (Santa Cruz Biotechnology, sc-17320), goat anti-Sox2 (R&D Systems, AF2018-SP), rabbit anti-Sox2 (Cell Signaling, 23064), rabbit anti Gata4 (Santa Cruz Biotechnology, sc-9053), goat anti-Gata4 (R&D Systems, AF2606-SP), rabbit anti-GFP (MBL, 598), mouse anti-Rab11 (BD Biosciences; 610656), rat anti-integrin-β1 (Merck Millipore, MAB1997), goat anti-biotinylated Gata4 (R&D Systems, BAF2606) mouse anti-Oct3/4 (Santa Cruz Biotechnology, sc-5279) and mouse anti-Hsp47 (Enzo Life Sciences, M16.10A1).

Techniques: Recombinant, Software

Journal: PLoS Genetics

Article Title: Congenital lipodystrophy induces severe osteosclerosis

doi: 10.1371/journal.pgen.1008244

Figure Lengend Snippet: A) Transplanted fat depots 3 months after surgery. B) Fat depot weight 3 months after transplantation. C) Serum leptin and adiponectin of FF mice 3 months after fat depot transplantation. WT and non-transplanted FF mice serve as control. D) μCT analysis of trabecular bone volume and bone mineral density of distal femurs of FF mice 3 months after sham operation or transplantation of fat derived from WT or adipokine-deficient mice. Data are presented as mean ± SD. **p<0.01; *** p<0.001 as determined by ANOVA with Holm-Sidak's post hoc analysis for multiple comparisons test. D) comparison with FF Sham except where detailed.

Article Snippet: The primary antibody cocktail contained rat anti-mouse CD45-BUV395 (BD Horizon, clone 30-F11, final dilution factor 1:200), rat anti-mouse TER-119-APC (BioLegend, clone TER-119, 1:200), rat anti-mouse CD41-BV421 (BioLegend, clone MWReg30, 1:300), rat anti-mouse/human CD11b (BioLegend, clone M1/70, 1:400), and rat-anti mouse Leptin receptor (LepR)-biotin (R&D Systems, polyclonal, 1:50) in Brilliant Stain Buffer (BD Biosciences) containing 10 μg/mL FcBlock.

Techniques: Transplantation Assay, Control, Derivative Assay, Comparison