Journal: Science Advances

Article Title: Stem cell–homing biomimetic hydrogel promotes the repair of osteoporotic bone defects through osteogenic and angiogenic coupling

doi: 10.1126/sciadv.adq6700

Figure Lengend Snippet: ( A ) Micro-CT scan and 3D reconstruction results (scale bar, 1 mm); quantitative analysis of bone density, volume, and other parameters. BV/TV, bone tissue volume/total tissue; Tb.Th, trabecular thickness; Tb.N, trabecular number. ( B ) H&E staining images of different groups (scale bar, 1 mm and 125 μm). ( C ) Masson staining images of different groups (scale bar, 1 mm and 125 μm). ( D ) Flow cytometry showed BMSCs from osteoporotic rats were positive for CD44 (99.8%), CD29 (99.8%), CD73 (98.9%), and CD90 (77.3%) and negative for CD34 (2.57%) and CD45 (2.5%). ( E ) Alizarin Red S, Toluidine blue, and Oil Red O staining were used to detect osteogenic, chondrogenic, and adipogenic differentiation, respectively (scale bar, 100 μm). ( F ) SA-β-gal staining detected senescent cells (scale bar, 100 μm). ( G ) Western blotting analysis of Nanog, Sox2, and Oct4 protein expression in BMSCs from different groups. ( H ) Western blotting analysis of P53, P21, and P16 protein expression in BMSCs from different groups. ( I ) Representative immunofluorescence staining of γ-H2AX foci formation in BMSCs from different groups (scale bar, 25 μm). DAPI, 4′,6-diamidino-2-phenylindole. ( J ) Representative immunofluorescence staining of CD90 and Runx2 in BMSCs from the different groups (scale bar, 25 μm). ( K ) Representative immunofluorescence staining of CD90 and Opn in BMSCs from different groups (scale bar, 25 μm). ( L ) Western blotting analysis of Runx2 and Opn protein expression in BMSCs from different groups. ( M ) ALP staining of BMSCs from the different groups. ( N ) Alizarin Red S staining of BMSCs from different groups. n = 3 for each group. Error bars denote means ± SEM; ns, no significance; * P < 0.05, ** P < 0.01, and *** P < 0.001.

Article Snippet: Following two washes in PBS, BMSCs were incubated for 6 hours in ALP staining buffer (Beyotime Institute of Biotechnology, China).

Techniques: Micro-CT, Staining, Flow Cytometry, Western Blot, Expressing, Immunofluorescence

Journal: Science Advances

Article Title: Stem cell–homing biomimetic hydrogel promotes the repair of osteoporotic bone defects through osteogenic and angiogenic coupling

doi: 10.1126/sciadv.adq6700

Figure Lengend Snippet: ( A ) Immunofluorescence staining of F-actin in BMSCs cultured with hydrogels for 7 days (scale bar, 25 μm). ( B ) Live/dead assay of BMSCs cultured with hydrogels for 3 days (scale bar, 100 μm). Live cells appear green, and dead cells appear red. ( C ) Western blots analysis of P-eNOS and eNOS levels in BMSCs cultured with hydrogels for 7 days. ( D ) NO generation in BMSCs cultured with hydrogels for 7 days indicated by nitrite levels. ( E ) cGMP expression in BMSCs cultured with hydrogels for 7 days. ( F ) Western blots analysis of sGC and PKG levels in BMSCs cultured with hydrogels for 7 days. ( G ) Western blots analysis of Runx2 and Opn levels in BMSCs cultured with hydrogels for 7 days. ( H ) Representative of immunofluorescence staining of CD90 and Runx2 in BMSCs cultured with hydrogels for 7 days (scale bar, 25 μm). ( I ) Representative of immunofluorescence staining of CD90 and Opn in BMSCs cultured with hydrogels for 7 days (scale bar, 25 μm). ( J ) The ALP staining of BMSCs cultured with hydrogels for 7 days. ( K ) Alizarin Red S staining of BMSCs cultured with hydrogels for 7 days. ( L ) Schematic diagram of SA-MSNs@CM-Stiff activating NO/cGMP pathway to promote BMSC osteogenesis. n = 3 for each group. Error bars denote means ± SEM; * P < 0.05, ** P < 0.01, and *** P < 0.001.

Article Snippet: Following two washes in PBS, BMSCs were incubated for 6 hours in ALP staining buffer (Beyotime Institute of Biotechnology, China).

Techniques: Immunofluorescence, Staining, Cell Culture, Live Dead Assay, Western Blot, Expressing

Journal: Drug Design, Development and Therapy

Article Title: Regulation of bone formation by baicalein via the mTORC1 pathway

doi: 10.2147/DDDT.S81578

Figure Lengend Snippet: MC3T3-E1 cells increased osteogenesis in the presence of BN. Notes: ( A ) Molecular structure of BN. ( B ) CCK-8 assays showed that BN did not significantly affect the growth of MC3T3-E1 cells at the concentrations used (0.1 μmol/L, 1 μmol/L, 10 μmol/L, 100 μmol/L, and 1,000 μmol/L) after treatment for 14 days, and BN at concentrations of 0.1–100 μM did not significantly affect cell growth. ( C ) The ALP activity assay showed that BN increased intracellular ALP expression during osteoblast differentiation in cultured MC3T3-E1 cells. ( D ) ALP staining indicates that treatment with 0.1–10 μM BN increased ALP activity in MC3T3-E1 cells in osteogenic differentiation medium for 14 days. ( E ) Micrograph of ALP staining indicates that the expression of ALP shows dose-dependent with 0.1–10 μM BN treatment. * P <0.05 versus group without BN; + P <0.05 versus group with 0.1 μM BN; # P <0.05 versus group with 1 μM BN. Columns represent the mean ± standard error from six wells per group ( B , C ). Abbreviations: ALP, alkaline phosphatase; BN, baicalein; CCK-8, Cell Counting Kit-8; OD, optical density.

Article Snippet: Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, washed, incubated with ALP staining buffer (NBT-BCIP, Sigma-Aldrich) at 37°C for 30 minutes, and washed with PBS to remove excess dye.

Techniques: CCK-8 Assay, ALP Activity Assay, Expressing, Cell Culture, Staining, Activity Assay, Cell Counting