Fig 1.
Alterations in Hedgehog signaling-related genes are key characteristics in the transcriptomic landscape of OP patients.
A: Volcano plots showing the DEGs in OP vs. controls (Log2Foldchange>1.0; FDR<0.05). Upregulated and downregulated genes are shown in black and red, respectively, while pink dots represent genes without significant differences. The genes labeled by the black arrow are hub genes involved in the Hedgehog signaling pathway. B: Dot plot showing the results of KEGG pathway enrichment analysis on differentially expressed genes in OP vs. control comparisons. The dot color represents the P value of each pathway. C: Protein-protein interaction network of SHH, IHH, GLI2, and RUNX2. The dot color and label size represent the importance of each gene in this topological network. D: Dot plots showing KEGG pathway enrichment analysis results of genes involved in the protein-protein network.
Fig 2.
Histological assessment of bone tissue in an osteoporosis rat model after BSHX treatment.
A: The morphology of femoral head in each experimental group under light microscope Scale bar, 500 μm. B: Immunofluorescence staining shows the expression of collagen I in BMSCs. The expression level of collagen I is represented by the degree of green fluorescence, while nuclei were counterstained by using DAPI (blue fluorescence). Scale bar, 100 μm.
Table 1.
Comparison of the bone mineral density in five experimental groups.
Fig 3.
Changes in the Hedgehog signaling pathway after BSHX treatment.
A: Immunohistochemical staining of SHH, IHH, GLI2, and RUNX2. Scale bar, 500 μm. B: RT-qPCR analysis of Shh, Ihh, Gli2, and Runx2 mRNA levels in rat bone tissue. *P<0.05 compared with Blank group; #P<0.05 compared with Calcium group. C: the concentrations of SHH, IHH, Gli2, and Runx2 in rat bone tissue were measured by ELISA. *, P<0.05.
Table 2.
Comparison of the optical density of SHH, IHH, GLI2, and RUNX2 between groups.
Fig 4.
Effect of BSHX on expression levels of ALP, BMP, and COL1A1 in BMSCs.
The concentration of ALP (A), BMP (B) and COL1A1 (C) in BMSCs after BSHX, osteogenic inducer, or estradiol treatment.
Fig 5.
mRNA and protein expression levels of SHH, IHH, GLI2, and RUNX2 in BMSCs after BFHX treatment.
A: qRT-PCR results displayed the mRNA expression levels of Shh, Gli2, and Runx2 in BMSCs after BFHX treatment. B: Western blot and quantification of protein levels of SHH, IHH, GLI2 and RUNX2 in BMSCs after BSHX treatment. *, P<0.05; **, P<0.01.
Fig 6.
Quercetin is the hub active compounds of BSHX in improving OP.
A: Enrichment analysis of metabolites in BSHX. B: Protein-protein interaction network of targets of metabolites in BSHX. C: Scatter plot showing the results of KEGG pathway enrichment analysis on targets of bioactive compounds in BSHX. The dot color represents the P value of each pathway. D: Pharmacology network of targets of bioactive compounds in BSHX. E: RT-qPCR assay detects the expression levels of Hedgehog-related genes in OP model rats after quercetin treatment.