Journal: International Journal of Oral Science

Article Title: circ_0003204 regulates the osteogenic differentiation of human adipose-derived stem cells via miR-370-3p/HDAC4 axis

doi: 10.1038/s41368-022-00184-2

Figure Lengend Snippet: circ_0003204 negatively regulated hASC osteogenesis. a Images of ALP and ARS stainings after transfection (scale bar = 200 μm). b – d Western blot after 7-day osteogenic induction of hASCs detected the protein expressions and the results were quantitated. e , f The expressions of ALPL and RUNX2 after 7-day osteogenic induction of hASCs were evaluated by RT-qPCR. (* P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.000 1)

Article Snippet: After 14-day osteogenic differentiation, the mineralized nodules were measured by ARS staining using 0.1% ARS staining solution (pH = 4.2, Cyagen).

Techniques: Transfection, Western Blot, Quantitative RT-PCR

Journal: International Journal of Oral Science

Article Title: circ_0003204 regulates the osteogenic differentiation of human adipose-derived stem cells via miR-370-3p/HDAC4 axis

doi: 10.1038/s41368-022-00184-2

Figure Lengend Snippet: circ_0003204 inhibited hASC osteogenesis through sponging miR-370-3p. a Bioinformatic analysis predicted the target relationship. b RT-qPCR evaluated the expression of miR-370-3p. c The putative binding sequences. d Luciferase reporter genes were co-transfected with miR‐370‐3p mimic/mimic-NC and mut-type/wild-type circ_0003204 into cells, and the luciferase activities were measured. e RT-PCR evaluated the expression of circ_0003204 and miR-370-3p. f Images of ALP and ARS stainings after transfection. g , h ALP and ARS activity analyses were measured. i RT-qPCR evaluated the mRNA expressions after 7-day osteogenic induction of hASCs (* P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.000 1)

Article Snippet: After 14-day osteogenic differentiation, the mineralized nodules were measured by ARS staining using 0.1% ARS staining solution (pH = 4.2, Cyagen).

Techniques: Quantitative RT-PCR, Expressing, Binding Assay, Luciferase, Transfection, Reverse Transcription Polymerase Chain Reaction, Activity Assay

Journal: International Journal of Oral Science

Article Title: circ_0003204 regulates the osteogenic differentiation of human adipose-derived stem cells via miR-370-3p/HDAC4 axis

doi: 10.1038/s41368-022-00184-2

Figure Lengend Snippet: miR-370-3p regulated hASC osteogenesis by targeting HDAC4. a Bioinformatic analysis predicted the target relationship. b The expressions of HDAC4 were evaluated by RT-qPCR. c circ_0003204 siRNAs carried Cy3 reporter (red) and transfection efficiency was confirmed by fluorescence microscopy (scale bar = 100 μm). d Images of ALP and ARS stainings after transfection. e , f ALP and ARS activity analyses were detected. g RT-qPCR evaluated the mRNA expression levels of BGLAP after 7-day osteogenic induction of hASCs. h The expression of HDAC4 was regulated by circ_0003204. (* P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.000 1)

Article Snippet: After 14-day osteogenic differentiation, the mineralized nodules were measured by ARS staining using 0.1% ARS staining solution (pH = 4.2, Cyagen).

Techniques: Quantitative RT-PCR, Transfection, Fluorescence, Microscopy, Activity Assay, Expressing

Journal: International Journal of Oral Science

Article Title: circ_0003204 regulates the osteogenic differentiation of human adipose-derived stem cells via miR-370-3p/HDAC4 axis

doi: 10.1038/s41368-022-00184-2

Figure Lengend Snippet: HDAC4 inhibited hASC osteogenesis. a , b Images of ALP and ARS stainings after transfection (scale bar = 200 μm). c Western blot evaluated the protein expressions after 7-day osteogenic induction of hASCs and histograms show quantification of the band intensities. d RT-qPCR evaluated the mRNA expressions after 7-day osteogenic induction of hASCs. e , f Immunofluorescence (IF) staining of COL1A1 (green) and RUNX2 (red) was performed after 7-day osteogenic differentiation (scale bar = 40 μm). The 4’,6-diamidino-2-phenylindole (DAPI) stained nuclei (blue) (* P < 0.05, ** P < 0.01, **** P < 0.000 1)

Article Snippet: After 14-day osteogenic differentiation, the mineralized nodules were measured by ARS staining using 0.1% ARS staining solution (pH = 4.2, Cyagen).

Techniques: Transfection, Western Blot, Quantitative RT-PCR, Immunofluorescence, Staining

Journal: International Journal of Oral Science

Article Title: circ_0003204 regulates the osteogenic differentiation of human adipose-derived stem cells via miR-370-3p/HDAC4 axis

doi: 10.1038/s41368-022-00184-2

Figure Lengend Snippet: Characterization of GelMA and coculture of hASCs in GelMA. a The diameter distribution of GelMA. b , c The SEM image exhibited the three-dimensional structure of GelMA and the morphology of hASCs on the surface of GelMA. d Confocal microscopy was used to observe the morphology of hASCs encapsulated in the GelMA (scale bar = 100 μm). e Images of Calcein-AM/PI staining, where calcein-AM indicated live cells with green fluorescence and PI indicated dead cells with red fluorescence (scale bar = 200 μm). f Proliferation ability of hASCs encapsulated in GelMA scaffold was measured using CCK-8. g Images of ALP and ARS stainings after transfection

Article Snippet: After 14-day osteogenic differentiation, the mineralized nodules were measured by ARS staining using 0.1% ARS staining solution (pH = 4.2, Cyagen).

Techniques: Confocal Microscopy, Staining, Fluorescence, CCK-8 Assay, Transfection

Journal: RSC Advances

Article Title: Polydopamine-functionalized acellular fish scale scaffolds for accelerated bone tissue regeneration †

doi: 10.1039/d5ra01932j

Figure Lengend Snippet: Osteogenic differentiation ability of scaffolds in vitro . (A) ALP staining and ALP activity of BMSCs after 7 and 14 days of osteogenic induction, and ARS staining and quantitative analysis of calcium nodules after 21 days of osteogenic induction. Scale bar = 200 μm. (B and C) Fluorescence microscopy images and quantitative analysis of IF staining of RUNX2, OPN, and COL-1α in BMSCs after 14 days of osteogenic induction. Scale bar = 100 μm. (D) The mRNA expression levels of RUNX2, OPN, and COL-1α in BMSCs after 14 days of osteogenic induction. All values are presented as means ± standard deviation (ns: no significant differences, *: p < 0.05, **: p < 0.01, ***: p < 0.001, ****: p < 0.0001).

Article Snippet: After 21 days of osteogenic induction, cells were fixed with 4% paraformaldehyde and stained using an ARS kit (Solarbio, China) ( n = 3).

Techniques: In Vitro, Staining, Activity Assay, Fluorescence, Microscopy, Expressing, Standard Deviation

Journal: Journal of Nanobiotechnology

Article Title: Spatiotemporal regulation of angiogenesis/osteogenesis emulating natural bone healing cascade for vascularized bone formation

doi: 10.1186/s12951-021-01173-z

Figure Lengend Snippet: In vitro osteogenesis potential. A Representative ALP staining images on day 3. B Quantitative analysis of ALP activity of BMSCs. C Representative ARS images on day 14. D Quantitative analysis of calcium deposition of BMSCs. E , F Osteogenesis of BMSCs was measured by immunofluorescence assays for Runx2 expression on day 4, with the nucleus stained in blue and OCN stained in red. qRT-PCR analysis of osteogenesis-related gene expressions including G ALP, H Col-1, I Runx2. J Osterix and K OCN. Statistically significant differences are indicated with *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001

Article Snippet: The cells were further incubated with ARS Staining Solution (Beyotime, China) for 30 min. Stained cells were observed by an inverted light microscope (Olympus, Japan).

Techniques: In Vitro, Staining, Activity Assay, Immunofluorescence, Expressing, Quantitative RT-PCR

Journal: Heliyon

Article Title: Bone marrow fatty acids affect osteoblastic differentiation through miR-92b-3p in the early stages of postmenopausal osteoporosis

doi: 10.1016/j.heliyon.2023.e16513

Figure Lengend Snippet: The Effects of fatty acids on osteogenic differentiation of MC3T3-E1 cells (A) ARS and ALP staining after culture for 14 days. (B) Relative mRNA expression level of osteoblastic-related genes after culture for 14 days. MTT assay of palmitoleate (C), pentadecanoate (D) and palmitate (E). Relative mRNA expression level of osteoblastic-related genes after treatment with palmitoleate (F), pentadecanoate (G) and palmitate (H). *p < 0.05, **p < 0.01, ***p < 0.001. Experiments were replicated three times.

Article Snippet: After washing three times with double-distilled H 2 O, the cells were incubated with ARS staining solution (Shanghai Yuanye Biotechnology Co., Ltd., Shanghai, China) for 30 min or stained with the BCIP/NBT Alkaline Phosphatase Color Development Kit (Beyotime Institute of Biotechnology) at room temperature according to the manufacturer's instructions.

Techniques: Staining, Expressing, MTT Assay

Journal: Heliyon

Article Title: Bone marrow fatty acids affect osteoblastic differentiation through miR-92b-3p in the early stages of postmenopausal osteoporosis

doi: 10.1016/j.heliyon.2023.e16513

Figure Lengend Snippet: MiR-92b-3p regulates the osteogenic differentiation of MC3T3-E1 cells by targeting PTEN. (A) The expression of miR-92b-3p during osteogenesis. (B) Relative expression level of osteoblastic-related genes after transfection with miR-92b-3p mimic or inhibitor. (C) ARS and ALP staining after transfection with miR-92b-3p mimic or inhibitor. (D) The binding sites between miR-92b-3p and PTEN. (E) The mRNA expression of PTEN after transfection with miR-92b-3p mimic or inhibitor. (F) Luciferase reporter assay exploring the relationship between miR-92b-3p and PTEN. *p < 0.05, **p < 0.01, ***p < 0.001. Experiments were replicated three times.

Article Snippet: After washing three times with double-distilled H 2 O, the cells were incubated with ARS staining solution (Shanghai Yuanye Biotechnology Co., Ltd., Shanghai, China) for 30 min or stained with the BCIP/NBT Alkaline Phosphatase Color Development Kit (Beyotime Institute of Biotechnology) at room temperature according to the manufacturer's instructions.

Techniques: Expressing, Transfection, Staining, Binding Assay, Luciferase, Reporter Assay

Journal: Heliyon

Article Title: Bone marrow fatty acids affect osteoblastic differentiation through miR-92b-3p in the early stages of postmenopausal osteoporosis

doi: 10.1016/j.heliyon.2023.e16513

Figure Lengend Snippet: Fatty acids affected the osteoblastogenesis of MC3T3-E1 cells through miR-92b-3p. (A) The expression of miR-92b-3p after treatment with fatty acids. Relative expression level of osteoblastic-related genes after transfection with miR-92b-3p mimic or inhibitor in the palmitate (B), pentadecanoate (C) and palmitoleate (D) groups. (E) ARS and ALP staining after transfection with miR-92b-3p mimic or inhibitor in the palmitate, pentadecanoate and palmitoleate groups. *p < 0.05, **p < 0.01, ***p < 0.001. Experiments were replicated three times.

Article Snippet: After washing three times with double-distilled H 2 O, the cells were incubated with ARS staining solution (Shanghai Yuanye Biotechnology Co., Ltd., Shanghai, China) for 30 min or stained with the BCIP/NBT Alkaline Phosphatase Color Development Kit (Beyotime Institute of Biotechnology) at room temperature according to the manufacturer's instructions.

Techniques: Expressing, Transfection, Staining

Journal: Bioactive Materials

Article Title: HydroWrap for T2DM-Related Fractures: A smart H 2 S-delivery controller modulating Macrophage senescence

doi: 10.1016/j.bioactmat.2025.05.007

Figure Lengend Snippet: Macrophage Rejuvenation Enhances Angiogenesis and Osteogenesis. (a) Schematic illustration of the experimental approach to evaluate the impact of conditioned medium from macrophages on angiogenesis and osteogenesis. (b) Crystal violet staining of HUVECs in the wound healing assay, and (c) quantitative analysis of wound closure. (d) Representative images from the tube formation assay and (e) quantitative analysis of vessel area percentage. (f) Representative images of ALP and ARS staining in MC3T3-E1 cells, accompanied by (g) quantitative analysis of ALP activity and (h) mineralization levels. (i) Relative mRNA expression levels of angiogenesis-related genes in HUVECs and (j) osteogenesis-related genes in MC3T3-E1 cells (n = 6 per group). Statistical significance is indicated as follows: ∗, #, and $ denote p < 0.05 when comparing the Control, HydroWrap, and HydroWrap + NIR groups to the T2DM group, respectively.

Article Snippet: Additional kits, including the BCIP/NBT ALP Color Development Kit, ALP Assay Kit, ARS Staining Kit, and Enhanced Mitochondrial Membrane Potential Assay Kit with JC-1, were acquired from Beyotime Biotechnology (Jiangsu, China).

Techniques: Staining, Wound Healing Assay, Tube Formation Assay, Activity Assay, Expressing, Control