Journal: Frontiers in Immunology

Article Title: Host Genetics and Antiviral Immune Responses in Adult Patients With Multisystem Inflammatory Syndrome

doi: 10.3389/fimmu.2021.718744

Figure Lengend Snippet: Antiviral and inflammatory responses of patient PBMCs in response to SARS-CoV-2 infection. (A–H) Patient (P1-P5), healthy controls (HC) (n=5) and COVID-19 intensive care unit (ICU) patient (COVID-19) (n=9) peripheral blood mononuclear cells were left untreated (UT) or infected with SARS-CoV-2 at a multiplicity of infection of 0.5 for 24 h. Induction of IFNs and proinflammatory cytokines was measured in supernatants by U-plex mesoscale technology. Data are pooled from two independent experiments, but the experiment was performed only once for each patient due to limited patient material. All stimulations were done in triplicate in the individual experiments. (I) Autoantibodies against IFNα and IFNω measured in serum from patients, healthy controls (n=15) and COVID-19 ICU patients (n=13) by ELISA, y-axis depicts blank-corrected OD450-630. Values above 0.5 (dashed line) are considered positive. Statistical differences were calculated using Kruskal-Wallis with correction for multiple comparisons by Dunn’s test. *p < 0.05, **p < 0.01, ***p < 0.001. Each line, showing the result of a statistical test, compares one patient with the HC or the COVID-19 ICU group. Only comparisons between patient and controls, which are statistically different, are indicated on the graph.

Article Snippet: Briefly, ELISA plates were coated with 1 μg/mL IFNα (130-093-874, Miltenyi Biotec) or IFNω (BMS304, Invitrogen) overnight at 4°C followed by blocking in 5% skimmed milk.

Techniques: Infection, Enzyme-linked Immunosorbent Assay

Journal: International Journal of Oral Science

Article Title: Gli2 and Gli3 synergistically mediate HH-TGF-β crosstalk in mesenchymal progenitor cells to orchestrate tooth root morphogenesis

doi: 10.1038/s41368-026-00427-6

Figure Lengend Snippet: Spatiotemporal co-expression pattern of Gli2 and Gli3 in the developing root. a UMAP plot showing cell clusters of dental epithelium and mesenchyme in mouse molars at P3.5. b , c Feature plots showing the expression of Gli2 and Gli3 in the first molar at P3.5. d – g Colocalization of Gli2 and Gli3 by RNAscope in the first molar at P3.5. h – k Colocalization of Gli2 and Gli3 by RNAscope in the first molar at P7.5. l – o Colocalization of Gli2 and Gli3 by RNAscope in the first molar at P12.5. p , p’ Colocalization of Gli2 and tdTomato (tdT) at P7.5 after tamoxifen (TAM) induction at P3.5. q , q’ Colocalization of Gli3 and tdT at P7.5 after TAM induction at P3.5. White boxes represent the apical region. White dotted lines outline the Hertwig’s epithelial root sheath (HERS). White arrows denote positive signals of Gli2 or Gli3 . White arrowheads indicate colocalized signals. Scale bars, 100 μm

Article Snippet: The antibodies used for CUT&Tag were: Gli2 (1:50, NB-600-874, Novus Biologicals, USA), Gli3 (5 μg/10 6 cells, AF3690, R&D systems, USA), rabbit control IgG (1:50, AC005, Abclone, China), goat anti-rabbit IgG H&L (Ab207, Vazyme, China), and rabbit anti-goat IgG (H + L) (1:100, AS081, Abclone, China).

Techniques: Expressing, RNAscope

Journal: International Journal of Oral Science

Article Title: Gli2 and Gli3 synergistically mediate HH-TGF-β crosstalk in mesenchymal progenitor cells to orchestrate tooth root morphogenesis

doi: 10.1038/s41368-026-00427-6

Figure Lengend Snippet: Concurrent deletion of Gli2 and Gli3 in Gli1 + progenitor cells exacerbates root dysplasia. a – e Micro-CT sagittal sections, 3D reconstruction images, and HE staining of molars in control mice at P14.5 and P21.5. f – j Micro-CT sagittal sections, 3D reconstruction images, and HE staining of molars in Gli1-CreER;Gli2 fl/fl mice at P14.5 and P21.5. k – o Micro-CT sections, 3D reconstruction images, and HE staining of molars in Gli1-CreER;Gli3 fl/fl mice at P14.5 and P21.5. p – t Micro-CT sections, 3D reconstruction images, and HE staining of molars in Gli1-CreER;Gli2 fl/fl ;Gli3 fl/fl mice at P14.5 and P21.5. u – w Quantitative analysis of root length in control and mutant mice at P21.5 ( n = 4). White arrows denote the measurement of root length. Black arrows indicate normal alveolar bone. Red arrows indicate compromised alveolar bone. Data are presented as mean ± SD. **P < 0.01, ***P < 0.001 , ****P < 0.000 1, ns, no significance. Scale bars, ( b , e , g , j , l , o , q , t ), 200 μm; others, 500 μm

Article Snippet: The antibodies used for CUT&Tag were: Gli2 (1:50, NB-600-874, Novus Biologicals, USA), Gli3 (5 μg/10 6 cells, AF3690, R&D systems, USA), rabbit control IgG (1:50, AC005, Abclone, China), goat anti-rabbit IgG H&L (Ab207, Vazyme, China), and rabbit anti-goat IgG (H + L) (1:100, AS081, Abclone, China).

Techniques: Micro-CT, Staining, Control, Mutagenesis

Journal: International Journal of Oral Science

Article Title: Gli2 and Gli3 synergistically mediate HH-TGF-β crosstalk in mesenchymal progenitor cells to orchestrate tooth root morphogenesis

doi: 10.1038/s41368-026-00427-6

Figure Lengend Snippet: Deletion of Gli2 , Gli3 or both in dental epithelium has no effect on tooth root development. a – d Micro-CT images and HE staining of molars in control mice at P21.5. e – h Micro-CT images and HE staining of molars in K14-CreER;Gli2 fl/fl mice at P21.5. i-l Micro-CT images and HE staining of molars in K14-CreER;Gli3 fl/fl mice at P21.5. m – p Micro-CT images and HE staining of molars in K14-CreER;Gli2 fl/fl ;Gli3 fl/fl mice at P21.5. q – s Quantitative analysis of root length in control and mutant mice ( n = 4). White arrows point to root length measurement. Black boxes represent the furcation region of the first molar. Black dotted lines indicate the area of periodontal ligament. Data are presented as mean ± SD. ns, no significance. Scale bars, ( c , g , k , o ), 200 μm; ( d , h , l , p ), 100 μm; others, 500 μm

Article Snippet: The antibodies used for CUT&Tag were: Gli2 (1:50, NB-600-874, Novus Biologicals, USA), Gli3 (5 μg/10 6 cells, AF3690, R&D systems, USA), rabbit control IgG (1:50, AC005, Abclone, China), goat anti-rabbit IgG H&L (Ab207, Vazyme, China), and rabbit anti-goat IgG (H + L) (1:100, AS081, Abclone, China).

Techniques: Micro-CT, Staining, Control, Mutagenesis

Journal: International Journal of Oral Science

Article Title: Gli2 and Gli3 synergistically mediate HH-TGF-β crosstalk in mesenchymal progenitor cells to orchestrate tooth root morphogenesis

doi: 10.1038/s41368-026-00427-6

Figure Lengend Snippet: Concurrent loss of Gli2 and Gli3 profoundly disrupts cell differentiation and proliferation. a – c RNAscope staining of Dspp , and immunostaining of Periostin and Sp7 of molars in control mice at P21.5. d – f RNAscope staining of Dspp , and immunostaining of Periostin and Sp7 of molars in Gli1-CreER;Gli2 fl/fl mice at P21.5. g – i RNAscope staining of Dspp , and immunostaining of Periostin and Sp7 of molars in Gli1-CreER;Gli3 fl/fl mice at P21.5. j – l RNAscope staining of Dspp , immunostaining of Periostin and Sp7 of molars in Gli1-CreER;Gli2 fl/fl ;Gli3 fl/fl mice at P21.5. m – p Visualization of tdT + cells in molars from control, Gli1-CreER;Gli2 fl/fl , Gli1-CreER;Gli3 fl/fl and Gli1-CreER;Gli2 fl/fl ;Gli3 fl/fl mice at P14.5. q – t , q’ – t’ Immunostaining of Ki67 + in the first molar in control and mutant mice at P5.5. u – v Quantification of Ki67 + cells in epithelium and mesenchyme in control and mutant mice ( n = 4). White boxes represent the apical region. White dotted lines in ( c , f , i , l ) indicate the area of alveolar bone, and in ( q ’– t ’) outline HERS. White arrows indicate positive signals, while yellow arrows denote impaired signals. Data are presented as mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001, ns, no significance. Scale bars, ( a – t ), 100 μm; ( q ’– t ’), 50 μm

Article Snippet: The antibodies used for CUT&Tag were: Gli2 (1:50, NB-600-874, Novus Biologicals, USA), Gli3 (5 μg/10 6 cells, AF3690, R&D systems, USA), rabbit control IgG (1:50, AC005, Abclone, China), goat anti-rabbit IgG H&L (Ab207, Vazyme, China), and rabbit anti-goat IgG (H + L) (1:100, AS081, Abclone, China).

Techniques: Cell Differentiation, RNAscope, Staining, Immunostaining, Control, Mutagenesis

Journal: International Journal of Oral Science

Article Title: Gli2 and Gli3 synergistically mediate HH-TGF-β crosstalk in mesenchymal progenitor cells to orchestrate tooth root morphogenesis

doi: 10.1038/s41368-026-00427-6

Figure Lengend Snippet: Simultaneous deletion of Gli2 and Gli3 downregulates TGF-β signaling activity. a Clustered heatmap of bulk RNA-seq data from control and mutant groups at P5.5. b – d Volcano plots illustrating differentially expressed genes (DEGs) between control and mutant groups. e , f KEGG pathway analysis of downregulated genes showing top enriched signaling pathways in Gli3 mutant and double mutant groups. g Western blot of p-SMAD2, p-SMAD3, and total SMAD2/3 protein levels of tooth germ in control and mutant groups at P5.5. h – o Immunofluorescence staining of p-SMAD3 and p-SMAD2 in the first molar in four groups at P5.5. White boxes represent the apical region of the molar. White dotted lines outline HERS. White arrows indicate positive signals, while yellow arrows denote impaired signals. Scale bars, ( h – o ), 100 μm

Article Snippet: The antibodies used for CUT&Tag were: Gli2 (1:50, NB-600-874, Novus Biologicals, USA), Gli3 (5 μg/10 6 cells, AF3690, R&D systems, USA), rabbit control IgG (1:50, AC005, Abclone, China), goat anti-rabbit IgG H&L (Ab207, Vazyme, China), and rabbit anti-goat IgG (H + L) (1:100, AS081, Abclone, China).

Techniques: Activity Assay, RNA Sequencing, Control, Mutagenesis, Protein-Protein interactions, Western Blot, Immunofluorescence, Staining

Journal: International Journal of Oral Science

Article Title: Gli2 and Gli3 synergistically mediate HH-TGF-β crosstalk in mesenchymal progenitor cells to orchestrate tooth root morphogenesis

doi: 10.1038/s41368-026-00427-6

Figure Lengend Snippet: Gli2 and Gli3 bind to the promoter of Acvr2b to activate its expression. a Venn diagram showing commonly downregulated genes associated with TGF-β signaling among three comparison groups. b Feature plot of Acvr2b in the mouse molar at P3.5. c RT-qPCR showing the relative mRNA expression of Acvr2b in control and mutant molar mesenchyme ( n = 4). d – g RNAscope staining of Acvr2b expression in control and mutant mice. h Heatmap of CUT&Tag showing the binding profiles of Gli2 and Gli3 around ±3kb of the transcription start site (TSS). i Motif enrichment analysis for Gli2 and Gli3 in the molar mesenchyme. j Peak calling analysis showing enhanced binding of Gli2 and Gli3 to the promoter region of Acvr2b . The annotated peak is enlarged in the black box below. k CUT&Tag-qPCR showing enrichment at the Acvr2b promoter relative to the IgG control ( n = 3). White dotted lines outline HERS. *P < 0.05, **P < 0.01, ns, no significance. Scale bars: ( d – g ), 50 μm

Article Snippet: The antibodies used for CUT&Tag were: Gli2 (1:50, NB-600-874, Novus Biologicals, USA), Gli3 (5 μg/10 6 cells, AF3690, R&D systems, USA), rabbit control IgG (1:50, AC005, Abclone, China), goat anti-rabbit IgG H&L (Ab207, Vazyme, China), and rabbit anti-goat IgG (H + L) (1:100, AS081, Abclone, China).

Techniques: Expressing, Comparison, Quantitative RT-PCR, Control, Mutagenesis, RNAscope, Staining, Binding Assay

Journal: International Journal of Oral Science

Article Title: Gli2 and Gli3 synergistically mediate HH-TGF-β crosstalk in mesenchymal progenitor cells to orchestrate tooth root morphogenesis

doi: 10.1038/s41368-026-00427-6

Figure Lengend Snippet: Upregulation of TGF-β signaling activity partially rescues tooth root defects in double mutant mice. a – i Micro-CT images and HE staining of molars from control mice treated with vehicle, Gli1-CreER;Gli2 fl/fl ;Gli3 fl/fl mice treated with vehicle, and Gli1-CreER;Gli2 fl/fl ;Gli3 fl/fl mice treated with SRI-011381. Red arrows indicate the sites of tooth eruption. White arrows indicate the measurement of the tooth root length. j Quantitative analysis of root length among three treatment groups ( n = 4). k – n Immunofluorescence staining and quantitative analysis of Periostin in three treatment groups ( n = 3). o – r RNAscope staining and quantitative analysis of Dspp in three treatment groups ( n = 3). White arrows indicate positive signals, yellow arrows indicate impaired signals, and white arrowheads indicate restored signals. Data are presented as mean ± SD. *P < 0.05, **P < 0.01 , ***P < 0.001, ****P < 0.000 1. Scale bars, ( a , b , d , e , g , h ), 500 μm; ( c , f , i ), 200 μm; ( k – q ), 100 μm

Article Snippet: The antibodies used for CUT&Tag were: Gli2 (1:50, NB-600-874, Novus Biologicals, USA), Gli3 (5 μg/10 6 cells, AF3690, R&D systems, USA), rabbit control IgG (1:50, AC005, Abclone, China), goat anti-rabbit IgG H&L (Ab207, Vazyme, China), and rabbit anti-goat IgG (H + L) (1:100, AS081, Abclone, China).

Techniques: Activity Assay, Mutagenesis, Micro-CT, Staining, Control, Immunofluorescence, RNAscope