Osteopontin Antibody - #AF0227

Figure 4. Biocompatibility and osteogenic differentiation ability of HN25. G) Immunofluorescence staining of ALP and OPN after culturing 14 d on different samples. Scale bar = 100 μm. H) Volcano plot showing differently expressed genes in MSCs from HN25 under US irradiation compared to the control.

Figure 5. In vitro biocompatibilities and osteogenic activities of the substrates. (A) CCK-8 assay of hBMSCs on the indicated surfaces after 1, 3 and 7 days of culture. * denotes p < 0.05 and ** denotes p < 0.01 compared to Ti-NTs, # denotes p < 0.05 compared to the corresponding control group without peptides. Error bars denote the standard deviations over quadruplicate measurements with separately implants. (B) Confocal fluorescence microscopy images of hBMSCs stained with vinculin, F-actin and DAPI after being cultured for 24 h. Scale bar, 50 μm. (C-F) qRT-PCR assay of osteogenic gene expression of (C) ALP, (D) RUNX-2, (E) COL1 and (F) OPN of hBMSCs after 7 and 14 days of culture. * denotes p < 0.05, ** denotes p < 0.01 and *** denotes p < 0.001 compared to Ti-NTs-P-A; # denotes p < 0.05, & denotes p < 0.01 and $ denotes p < 0.001 compared to the corresponding control group without peptides. All error bars denote the standard deviations over quadruplicate measurements with separately implants. (G) Immunofluorescence staining of hBMSCs cultured on Ti-NTs-P-A for 7 days (ALP and RUNX-2) and 14 days (OPN). The images were obtained by confocal fluorescence microscopy. Scale bar, 50 μm. (H) Western blotting of hBMSCs cultured on the substrates for 7 and 14 days. At each time point, left lane was Ti-NTs, middle lane was Ti-NTs-A and right lane was Ti-NTs-P-A. * denotes p < 0.05, ** denotes p < 0.01 and *** denotes p < 0.001 compared to Ti-NTs-P-A. Error bars denote the standard deviations over triplicate measurements with separately Western blotting results.

Fig. 6 VSIG4 promoted SPP1 expression through enhancing the histone H3 lysine 18 lactylation and promoting the transcription activity of STAT3. (A) Co-culture of mATC with BMDMs isolated from VSIG4-KO and WT mice to examine the lactate concentration. (B) Isolation of TAMs from mATC-derived tumors in VSIG4-KO (n = 3) and WT mice (n = 3) on day 32 to examine the lactate concentration. Data are presented as mean ± S.E.M. (C-D) Co-culture of mATC with BMDMs isolated from VSIG4-KO and WT mice to examine the expressions of histone H3 lysine 18 lactylation (H3K18la). (E) Genome browser track analysis using dataset GSE192358 showed the H3K18la lactylation levels in the binding region of Spp1 with or without the lactate treatment, where the left area of the dotted line represented the binding region of H3K18la on the Spp1 promotor. The mRNA (F) and protein (G-H) level of Spp1 in BMDM with or without the lactate treatment (n = 3). (I-J) The expression of Stat1, pStat1, Stat3, pStat3, and pStat6 in BMDMs after VSIG4 knockout (n = 3). (K) The JASPAR database predicted the presence of STAT3 binding sites in the SPP1 promoter region. The mRNA (L) and protein (M-N) level of Spp1 in BMDMs after STAT3 inhibitor Stattic treatment (n = 3). (O) The transcriptional activity of STAT3 on SPP1 was determined by dual luciferase reporter gene assay (n = 3). Data are presented as mean ± S.D. *P 

Fig. 5 In vitro evaluation of C-T@Ti3C2 nanosheets for promoting osteogenesis. a Viability of rat bone marrow mesenchymal stem cells after incubation with different concentrations of C-T@Ti3C2 nanosheets (n = 3 for each group). b CLSM of AM/PI-stained rBMMSCs after various concentrations of C-T@Ti3C2 nanosheets. c CLSM images of the expression of BMP2, OCN, RUNX2 and OPN (at Day 14) in rBMMSCs after different treatments. A representative image of three replicates from each group is shown. d Relative mRNA levels of osteogenic genes (COL-I, BMP2, OPN and RUNX2). e Representative Western blots of total COL-I, OPN, BMP2, RUNX2 and β-actin after induction for 14 days in rBMMSCs. Data are presented as the mean ± SD, and statistical significance was calculated using the two-tailed t test

Fig. 5 In vitro evaluation of C-T@Ti3C2 nanosheets for promoting osteogenesis. a Viability of rat bone marrow mesenchymal stem cells after incubation with different concentrations of C-T@Ti3C2 nanosheets (n = 3 for each group). b CLSM of AM/PI-stained rBMMSCs after various concentrations of C-T@Ti3C2 nanosheets. c CLSM images of the expression of BMP2, OCN, RUNX2 and OPN (at Day 14) in rBMMSCs after different treatments. A representative image of three replicates from each group is shown. d Relative mRNA levels of osteogenic genes (COL-I, BMP2, OPN and RUNX2). e Representative Western blots of total COL-I, OPN, BMP2, RUNX2 and β-actin after induction for 14 days in rBMMSCs. Data are presented as the mean ± SD, and statistical significance was calculated using the two-tailed t test

Fig. 7. In vitro osteogenic efficiency-related gene expression detection of porous scaffolds. A–D: rBMSCs were cocultured with four types of porous scaffolds. After 7 and 14 days of osteogenic induction, RT‒PCR was performed to detect the expression of osteogenic-related genes, including OPN (A), RunX2 (B), Col-1 (C), and OCN (D). E and F: Western blotting assay used to detect osteogenic-related proteins. All statistical data are represented as mean ± SD (n = 3; ∗∗∗∗P < 0.0001).