Journal: Nature Communications

Article Title: Gli1 identifies osteogenic progenitors for bone formation and fracture repair

doi: 10.1038/s41467-017-02171-2

Figure Lengend Snippet: Gli1 marks MMPs in postnatal growing mice. a – f Confocal images of immunofluorescence staining for Endomucin a , b , Aggrecan c , d , or Lepr e , f on frozen sections of the proximal tibia. Boxed areas shown in magnified images to the right. GP growth plate. Arrowhead in f denotes a typical Lepr + tdTomato + cell. Percentage (mean ± SD, n = 3) of double positive cells over total tdTomato + cells was calculated from cancellous bone region extending 300 µm from growth plate in two sections each of three mice. Same below. Scale bars: 100 µm ( a , e ) or 500 µm c . g , h Confocal images of direct fluorescence on frozen sections of proximal tibia. Boxed area shown at a higher magnification to the right. Arrowheads in h denote Pdgfra + tdTomato + cells. Scale bar: 100 µm g . i FACS analyses of Lepr (left) or Pdgfra (right) expression among CD31 − CD45 − Ter119 − tdTomato + cells from the cancellous bone region beneath the growth plate of either distal femur or proximal tibia. Red and blue lines represent control IgG and specific antibody, respectively. j – o Confocal images of Runx2 j , k or Osx l – o immunofluorescence staining on frozen sections of the proximal tibia. Boxed areas are shown in magnified images to the right k , m , o . DAPI signal was omitted in k , m , o for better visualization. Scale bar: 100 µm. p FACS analyses of Lepr (left) or Pdgfra (right) expression among CD31 − CD45 − Ter119 − tdTomato + from the bone marrow. Red and blue lines represent the control and specific antibody, respectively. Quantification is presented as mean ± SD, n = 3. q – t Confocal images of immunofluorescence staining for perilipin on frozen sections of distal femur. Boxed areas are shown at higher magnification to the right. Percentage (mean ± SD, n = 3) of perilipin + tdTomato + over perilipin + adipocytes was acquired from cancellous bone region extending 400 µm from the growth plate in two sections each of three mice. Scale bar: 500 µm. u – z Confocal images showing Gli1 + cells (red) and immunofluorescence signal of Aggrecan (green) in distal femur of Gli1-CreER T2 ;Ai9 mice treated with TM at different ages and harvested after 24 h. Scale bar: 500 µm. Boxed areas are shown at higher magnification to the right v , x , z . (AA) Differential expression of MSC marker genes in Gli1 + MMPs vs. Gli1 − mesenchymal cells. Heatmap generated from RNA-seq data of three paired biological replicates. Scale derived from voom-transformed data indicating relative increase (red) or decrease (blue) in MMPs

Article Snippet: The antibodies used in this study include anti-mouse CD140a (Pdgfra) APC (17-1401-81, eBioscience, 1:100), goat-anti-Lepr-biotin (AF497, R&D, 1:100), anti-Ter119-PE-cy7 (25-5921-81, eBioscience, 1:1000), anti-CD31-PE-cy7 (25-0311-81, eBioscience, 1:1000), and anti-CD31-PE-cy7 (25-0451-81, eBioscience, 1:1000).

Techniques: Immunofluorescence, Staining, Fluorescence, Expressing, Marker, Generated, RNA Sequencing Assay, Derivative Assay, Transformation Assay

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