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SIS3 reprograms CAFs (a) Schematic representation depicting the in vitro activation of NIH3T3 cells into CAFs using an OS-conditioned medium. (b) Western blotting is used to assess the expression levels of CAFs activation <t>markers</t> <t>α-SMA</t> and FAP. (c) Immunofluorescence (IF) staining is used to visualize the expression of CAFs activation markers α-SMA and FAP (scale bar: 25 μm). (d) IF staining is used to detect the expression of α-SMA and FAP in NIH3T3 cells, CAFs, and CAFs co-cultured with SIS3 (scale bar: 25 μm). (e) Statistical analysis of relative fluorescence intensity (FI), presented as mean ± SD (n = 3). (f) Schematic illustration of the TGF-β/SMAD3 signaling pathway. (g) Western blotting is used to evaluate the activation status of the TGF-β/SMAD3 signaling pathway in NIH3T3 cells, CAFs, and CAFs co-cultured with SIS3 and the statistical analysis of relative <t>SMAD</t> and p-SMAD expression, presented as mean ± SD (n = 3). (h) Representative images from collagen gel contraction assays on NIH3T3 cells, CAFs, and CAFs co-cultured with SIS3 and the statistical analysis of relative contraction degree, presented as mean ± SD (n = 3). (i) Representative images from wound healing assays conducted on NIH3T3 cells, CAFs, and CAFs co-cultured with SIS3 (scale bar: 200 μm) and the statistical analysis of relative wound healing degree, presented as mean ± SD (n = 3). (j) Representative images from transwell migration assays performed on NIH3T3 cells, CAFs, and CAFs co-cultured with SIS3 (scale bar: 100 μm) and the statistical analysis of the relative number of migrated cells, presented as mean ± SD (n = 3).
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SIS3 reprograms CAFs (a) Schematic representation depicting the in vitro activation of NIH3T3 cells into CAFs using an OS-conditioned medium. (b) Western blotting is used to assess the expression levels of CAFs activation markers α-SMA and FAP. (c) Immunofluorescence (IF) staining is used to visualize the expression of CAFs activation markers α-SMA and FAP (scale bar: 25 μm). (d) IF staining is used to detect the expression of α-SMA and FAP in NIH3T3 cells, CAFs, and CAFs co-cultured with SIS3 (scale bar: 25 μm). (e) Statistical analysis of relative fluorescence intensity (FI), presented as mean ± SD (n = 3). (f) Schematic illustration of the TGF-β/SMAD3 signaling pathway. (g) Western blotting is used to evaluate the activation status of the TGF-β/SMAD3 signaling pathway in NIH3T3 cells, CAFs, and CAFs co-cultured with SIS3 and the statistical analysis of relative SMAD and p-SMAD expression, presented as mean ± SD (n = 3). (h) Representative images from collagen gel contraction assays on NIH3T3 cells, CAFs, and CAFs co-cultured with SIS3 and the statistical analysis of relative contraction degree, presented as mean ± SD (n = 3). (i) Representative images from wound healing assays conducted on NIH3T3 cells, CAFs, and CAFs co-cultured with SIS3 (scale bar: 200 μm) and the statistical analysis of relative wound healing degree, presented as mean ± SD (n = 3). (j) Representative images from transwell migration assays performed on NIH3T3 cells, CAFs, and CAFs co-cultured with SIS3 (scale bar: 100 μm) and the statistical analysis of the relative number of migrated cells, presented as mean ± SD (n = 3).

Journal: Bioactive Materials

Article Title: Hydrogel delivering antifibrotic agent and nano-sonosensitizer enhances efficacy of sonodynamic therapy in osteosarcoma treatment

doi: 10.1016/j.bioactmat.2025.10.001

Figure Lengend Snippet: SIS3 reprograms CAFs (a) Schematic representation depicting the in vitro activation of NIH3T3 cells into CAFs using an OS-conditioned medium. (b) Western blotting is used to assess the expression levels of CAFs activation markers α-SMA and FAP. (c) Immunofluorescence (IF) staining is used to visualize the expression of CAFs activation markers α-SMA and FAP (scale bar: 25 μm). (d) IF staining is used to detect the expression of α-SMA and FAP in NIH3T3 cells, CAFs, and CAFs co-cultured with SIS3 (scale bar: 25 μm). (e) Statistical analysis of relative fluorescence intensity (FI), presented as mean ± SD (n = 3). (f) Schematic illustration of the TGF-β/SMAD3 signaling pathway. (g) Western blotting is used to evaluate the activation status of the TGF-β/SMAD3 signaling pathway in NIH3T3 cells, CAFs, and CAFs co-cultured with SIS3 and the statistical analysis of relative SMAD and p-SMAD expression, presented as mean ± SD (n = 3). (h) Representative images from collagen gel contraction assays on NIH3T3 cells, CAFs, and CAFs co-cultured with SIS3 and the statistical analysis of relative contraction degree, presented as mean ± SD (n = 3). (i) Representative images from wound healing assays conducted on NIH3T3 cells, CAFs, and CAFs co-cultured with SIS3 (scale bar: 200 μm) and the statistical analysis of relative wound healing degree, presented as mean ± SD (n = 3). (j) Representative images from transwell migration assays performed on NIH3T3 cells, CAFs, and CAFs co-cultured with SIS3 (scale bar: 100 μm) and the statistical analysis of the relative number of migrated cells, presented as mean ± SD (n = 3).

Article Snippet: The primary antibodies used were: α-SMA (1:1000, ABclonal, A17910), FAP (1:1000, ABclonal, A23789 ), SMAD (1:2000, Proteintech, 66516-1-Ig), p-SMAD (S423+S425) (1:2000, Abcam, ab52903), STING (1:1000, Cell Signaling Technology, D2P2F), p-STING (Ser365) (1:1000, Cell Signaling Technology, D8F4W), p-TBK1 (Ser172) (1:1000, Cell Signaling Technology, D52C2), and p-IRF3 (Ser396) (1:1000, Cell Signaling Technology, D6O1M).

Techniques: In Vitro, Activation Assay, Western Blot, Expressing, Immunofluorescence, Staining, Cell Culture, Fluorescence, Migration