Journal: Bone Research

Article Title: Spatially resolved osteoblast-traced transcriptomics uncovers TGF-β as a combination target with sclerostin in osteoporosis

doi: 10.1038/s41413-026-00521-9

Figure Lengend Snippet: Transcriptomic profiles reveal that TGF-β signaling is negatively correlated with osteoblast activity. a Uniform manifold approximation and projection (UMAP) plot of all isolated regions, colored by group. b Heatmap showing the average expression of the top 20 up- and downregulated genes compared to D35. c Boxplots of osteogenesis and Wnt signaling inhibitor gene expression. Box = IQR; line = median; whisker = 1.5 × IQR. The Wilcoxon rank-sum test is used for statistics. d Violin plots of osteoblast marker gene set scores based on gene set variation analysis (GSVA). Each point = single region; Wilcoxon rank-sum test was used for statistics. e Bar plots of normalized enrichment scores of significant pathways identified by gene set enrichment analysis (GSEA). Color represents gene ratio with each pathway, representing the proportion of genes detected in the dataset relative to the total number of genes in each canonical gene set. f Violin plots of TGF-β signaling activity scores derived from GSVA. Each data point represents an individual region. The Wilcoxon rank-sum test was used for statistics. g Scatter plot showing a negative correlation between TGF-β signaling activity and osteoblast marker scores. Pearson correlation coefficient ( r ) and P -value are calculated across regions. * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.000 1; ns not significant

Article Snippet: First, TGF-β blocking antibody (10 mg/kg, BE0057, BioXCell, USA) was intraperitoneally injected three times a week.

Techniques: Activity Assay, Isolation, Expressing, Gene Expression, Whisker Assay, Marker, Derivative Assay

Journal: Bone Research

Article Title: Spatially resolved osteoblast-traced transcriptomics uncovers TGF-β as a combination target with sclerostin in osteoporosis

doi: 10.1038/s41413-026-00521-9

Figure Lengend Snippet: scRNA-seq shows downregulated TGF-β signaling in active osteoblasts. a Experimental timeline for scRNA-seq using the Dmp1 -CreERt2:mTmG mouse model across activation states and Scl-Ab treatment. b Overview of scRNA-seq workflow. Hindlimb bones were dissociated, CD45- cells were sorted, and subjected to scRNA-seq ( n = 4). c UMAP of 3 968 cells, with clusters identified by differential gene sets. Two osteoblast-related clusters (clusters 4 and 10) and one osteocyte cluster (cluster 17) were annotated. Color indicates the respective cluster. d Feature plots of osteoblast markers: Col1a1 , Bglap , Alpl , Runx2 , Dmp1 , and Phex . e Bar plots of GFP+ cells proportions by cluster. Green bars indicate the highest proportions. f UMAP of osteoblast-lineage clusters, colored by experimental group. g Stacked bar plots showing cell distribution across clusters. Left: within total cells; right: within GFP+ cells. h GSEA enrichment plot of TGF-β signaling

Article Snippet: First, TGF-β blocking antibody (10 mg/kg, BE0057, BioXCell, USA) was intraperitoneally injected three times a week.

Techniques: Activation Assay

Journal: Bone Research

Article Title: Spatially resolved osteoblast-traced transcriptomics uncovers TGF-β as a combination target with sclerostin in osteoporosis

doi: 10.1038/s41413-026-00521-9

Figure Lengend Snippet: TGF-β drives the transition of active osteoblasts into BLC-like cells in DBP-based bone organoid. a Experimental timeline of osteoblast culture on DBP and treatment with VD3, PGE2, or TGF-β. b Representative immunofluorescence images, showing actin-phalloidin (green) and nucleus-DAPI (blue). Scale bar = 100 μm. c Box plots of actin angles ( n = 8). Wilcoxon rank-sum test was used for statistics. d Representative confocal images, showing actin-phalloidin (green) and nucleus-DAPI (blue). e Bar plot of cell height quantified from confocal images ( n = 8). A t -test was used for statistics. f Representative immunofluorescence images, showing Ki67 (green), and nucleus-DAPI (blue). g Bar plot of the Ki67-positive cell ratio of stained cells ( n = 8). Proportions of Ki67-positive cells were calculated as the number of Ki67-positive cells divided by the number of DAPI-positive cells. Each point represents individual well. A t -test was used for statistics. *** P < 0.001; **** P < 0.000 1

Article Snippet: First, TGF-β blocking antibody (10 mg/kg, BE0057, BioXCell, USA) was intraperitoneally injected three times a week.

Techniques: Immunofluorescence, Staining

Journal: Bone Research

Article Title: Spatially resolved osteoblast-traced transcriptomics uncovers TGF-β as a combination target with sclerostin in osteoporosis

doi: 10.1038/s41413-026-00521-9

Figure Lengend Snippet: Lineage tracing reveals that TGF-β regulates osteoblast-BLC conversion and reactivation. a Experimental timeline of tamoxifen pulse and TGF-β/TGFβ-Ab administration in Dmp1 -CreERt2:mTmG mice. b Representative confocal images of femoral periosteum showing GFP (green), tdTomato (red) signals, and DAPI (blue). Scale bar = 20 μm. c Violin plot of GFP+ cell thickness. Each point represents individual cells. The Wilcoxon rank-sum test was used for statistics. d Bar plot of GFP+ cells per unit length. Cell numbers were determined from three comparable sections per mouse, with eight fields (400× magnification) analyzed per section. The Wilcoxon rank-sum test was used for statistics. e Box plot of serum P1NP levels. A t -test was used for statistics. Group sizes: 8 weeks ( n = 6), 9 weeks ( n = 5), TGF-β ( n = 4), Inactive (12 weeks control + 13 weeks control; n = 6), Scl-Ab+TGF-β ( n = 3), Scl-Ab ( n = 3), TGFβ-Ab ( n = 4), and Scl-Ab+TGFβ-Ab ( n = 5). * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.000 1; ns not significant

Article Snippet: First, TGF-β blocking antibody (10 mg/kg, BE0057, BioXCell, USA) was intraperitoneally injected three times a week.

Techniques: Control

Journal: Bone Research

Article Title: Spatially resolved osteoblast-traced transcriptomics uncovers TGF-β as a combination target with sclerostin in osteoporosis

doi: 10.1038/s41413-026-00521-9

Figure Lengend Snippet: Dual inhibition of TGF-β and sclerostin increases bone mass in a hindlimb unloading model. a Experimental timeline of hindlimb unloading using C57B1/6 J mice and treatment with TGFβ-Ab, Scl-Ab, or both. Mice euthanized at week 12. b Representative μCT images of femoral diaphysis. c Representative confocal images of trabecular bone showing calcein (green), alizarin (red) signals, and DAPI (blue). Scale bar = 100 μm. d Representative images of TRAP-stained femur sections; violet = TRAP-positive, green = counterstain. Scale bar = 200 μm. e – h Bar plot of trabecular bone volume/total volume (Tra. BV/TV, e ), trabecular thickness (Tb. Th, f ), trabecular separation (Tb. Sp, g ), and trabecular number (Tb. N, h ) from μCT. Data = mean ± standard deviation. i , j Bar plot of mineral apposition rate (Tra. MAR, i ) and trabecular bone formation rate per bone surface (Tra. BFR/BS, j ) from labeling images. Data = mean ± standard deviation k Box plot of TRAP+ osteoclasts per bone surface (OC.N/BS), measured through ImageJ from TRAP-stained slides. l – n Box plot of serum P1NP ( l ) and TRAP ( m ) levels, and RANKL/OPG ratio ( n ). Group sizes: Control ( n = 5), Un_con ( n = 5), Scl-Ab ( n = 6), TGFβ-Ab ( n = 6), and Scl-Ab+TGFβ-Ab ( n = 7). Each point represents an individual mouse. ANOVA was used for statistics; * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.000 1; ns not significant

Article Snippet: First, TGF-β blocking antibody (10 mg/kg, BE0057, BioXCell, USA) was intraperitoneally injected three times a week.

Techniques: Inhibition, Staining, Standard Deviation, Labeling, Control

Journal: bioRxiv

Article Title: Downregulation of Satb1 is required to prevent autoimmunity by maintaining Tfh homeostasis

doi: 10.64898/2026.02.03.703482

Figure Lengend Snippet: a, Experimental workflow for in vitro Th cells subsets differentiation. b, Expression of CXCR5 + Bcl6 + in Th subsets: Th0, Th1, Th2, Th17, TfhA (anti-TGF-β), Tfh5, Tfr cells differentiated in vitro from naive CD4 + T cells (age= 35 weeks) (WT n=7, KI n=8). c, Contour plot of flow cytometry showing expression of CXCR5 vs Bcl6 in TfhA and Tfh cells. d, In vivo analysis of splenic Th1, Th2, Th17, and Treg cells from aged WT and KI mice (WT n=7-10, KI n=7). e, Flow cytometric analysis of the percentage and total numbers of splenic Tfh cells from aged WT and KI mice (WT n=10, KI n=10). f, Flow cytometric analysis of the percentage and total numbers of splenic Tfr cells from aged WT and KI mice (WT n=10, KI n=10). g, In vivo analysis of Th1, Th2, Th17, and Treg cells in mLN from aged WT and KI mice (WT n=6-7, KI n=6-7). h, Flow cytometric analysis of the frequency of Tfh and Tfr in Satb1 deficient and Satb1 sufficient mice (WT n=7, KI n=4). d-g, Data are pooled from at least two independent experiments and are analyzed by unpaired t -test; ns indicates not significant p > 0.05, p < 0.05 = *; p < 0.01 = **; p < 0.001 = *** p < 0.0001 = ****

Article Snippet: In TfhA cultures, TGF-β was blocked with anti- TGF-β antibodies (BioXCell, BE0057) at 10 μg/ml.

Techniques: In Vitro, Expressing, Flow Cytometry, In Vivo