Journal: Materials Today Bio

Article Title: The combination of a 3D-Printed porous Ti–6Al–4V alloy scaffold and stem cell sheet technology for the construction of biomimetic engineered bone at an ectopic site

doi: 10.1016/j.mtbio.2022.100433

Figure Lengend Snippet: Gene primer sequences for RT‒PCR of BMSCs.

Article Snippet: Primary antibodies, including anti -OCN (Genetex, 1:1000), anti -OPN (Novusbio, 1:1000), anti-COL-I (Bioss, 1:1000), anti -Runx2 (Bioss, 1:1000), and anti -β-actin (Abcam, 1:1000), were added to the membranes.

Techniques:

Journal: Materials Today Bio

Article Title: The combination of a 3D-Printed porous Ti–6Al–4V alloy scaffold and stem cell sheet technology for the construction of biomimetic engineered bone at an ectopic site

doi: 10.1016/j.mtbio.2022.100433

Figure Lengend Snippet: In vitro analysis of osteogenesis of the combination of BMSCs and PT scaffolds. (A) Schematic of the experiment. BMSCs were collected from each group after 7 days and 14 days of coculturing with PT scaffolds in osteogenic induction medium. (B) Alizarin red staining images. (C) Quantification of Alizarin red staining based on the relative absorbance value. (D–G) The RT‒PCR results of osteogenesis-related gene expression of OCN, OPN, COL-I, and Runx2 in the PT, PT/LC, PT/MC, PT/HC, and PT/CS groups on days 7 and 14. These results suggested that increasing the number of initial seeding cells improved the capability of osteogenic differentiation. (H) Western blot (WB) analysis presented results similar to those of RT‒PCR. ∗ indicates significant differences (p ​< ​0.05).

Article Snippet: Primary antibodies, including anti -OCN (Genetex, 1:1000), anti -OPN (Novusbio, 1:1000), anti-COL-I (Bioss, 1:1000), anti -Runx2 (Bioss, 1:1000), and anti -β-actin (Abcam, 1:1000), were added to the membranes.

Techniques: In Vitro, Staining, Gene Expression, Western Blot

Journal: Nanomaterials

Article Title: Antibacterial and Osteogenic Activity of Titania Nanotubes Modified with Electrospray-Deposited Tetracycline Nanoparticles

doi: 10.3390/nano10061093

Figure Lengend Snippet: Osteogenic differentiation of osteoblasts on the specimens for 21 d. ( a ) The expression levels of osteopontin (OPN) and osteocalcin (OCN) assessed by real-time RT-PCR (* p < 0.05 vs. MA). ( b ) Fluorescence images of OCN (upper) and OPN (lower) (scale bar = 100 µm).

Article Snippet: To confirm the findings, immunofluorescence staining was performed with solutions of anti-OPN antibody (Santa Cruz Biotechnology, Dallas, TX, USA) and anti-OCN antibody (QED Bioscience, San Diego, CA, USA) in blocking solution (1% BSA in PBS) at 4 °C for 1 d. After the cells were rinsed with wash buffer, they were soaked in solution with goat anti-mouse lgG-FITC (Santa Cruz Biotechnology, Dallas, TX, USA) for 1 h. The stained cells were observed using CLSM (LSM700, Carl-Zeiss).

Techniques: Expressing, Quantitative RT-PCR, Fluorescence

Journal: Journal of Orthopaedic Surgery and Research

Article Title: T. gondii excretory proteins promote the osteogenic differentiation of human bone mesenchymal stem cells via the BMP/Smad signaling pathway

doi: 10.1186/s13018-024-04839-0

Figure Lengend Snippet: TgEP promoted the osteogenic differentiation of hBMSCs. ( A ) Cytotoxic effect of TgEP on hBMSCs detected using a cytotoxicity detection kit. ( B ) Effect of TgEP on cell proliferation assessed using a CCK-8 kit. ( C ) ALP staining was performed to evaluate ALP expression in the cells. Scale bar = 500 μm. ( D ) Quantification of ALP staining results using ImageJ software and statistical analysis. ( E ) Alizarin red staining was performed to measure the content of mineralized nodules in the cells. Scale bar = 500 μm. ( F ) Quantification of alizarin red staining results using ImageJ software and statistical analysis. ( G ) mRNA transcription levels of osteogenic genes were assessed using qRT‒PCR after the cells were treated for 3 or 7 days. ( H , J ) Western blotting analysis of the osteogenic proteins ALP, Runx2, Osx, and OCN in hBMSCs treated for 5 days ( H ) or 7 days ( J ). The expression levels were quantified using β-actin as a loading control ( I , K ). ( L ) Immunofluorescence staining was performed to detect the expression of Runx2 in the cells. Scale bar = 30 μm. (Data are presented as the mean ± SD; n = 3; * P < 0.05, ** P < 0.01, *** P < 0.001, ns P >0.05 compared to the control group)

Article Snippet: For immunohistochemistry analysis, tissue sections were stained according to standard protocols using anti-osteocalcin (OCN) antibodies (Cat.#: 614,487) (1:100, ZEN-BIO, China).

Techniques: CCK-8 Assay, Staining, Expressing, Software, Western Blot, Control, Immunofluorescence

Journal: Journal of Orthopaedic Surgery and Research

Article Title: T. gondii excretory proteins promote the osteogenic differentiation of human bone mesenchymal stem cells via the BMP/Smad signaling pathway

doi: 10.1186/s13018-024-04839-0

Figure Lengend Snippet: TgEP promotes the repair of bone defects in vivo. ( A ) Histological analysis of bone tissue sections at 6 weeks, including H&E staining, Masson’s trichrome staining, and osteocalcin immunohistochemical staining. Scale bar = 500 μm. ( B ) Histological analysis of bone tissue sections at 8 weeks, including H&E staining, Masson’s trichrome staining, and osteocalcin immunohistochemical staining. Scale bar = 500 μm.

Article Snippet: For immunohistochemistry analysis, tissue sections were stained according to standard protocols using anti-osteocalcin (OCN) antibodies (Cat.#: 614,487) (1:100, ZEN-BIO, China).

Techniques: In Vivo, Staining, Immunohistochemical staining

Journal: Dentistry Journal

Article Title: In Vivo Evaluation of Regenerative Osteogenic Potential Using a Human Demineralized Dentin Matrix for Dental Application

doi: 10.3390/dj12030076

Figure Lengend Snippet: Representative histological images of (H&E) ( A , D , G , J ), Masson’s trichrome ( B , E , H , K ) and osteocalcin immune staining ( C , F , I , L ) are shown at 4 weeks post-treatment. Coronal sections through the mid-point of the defects were prepared from decalcified specimens. Fibrous connective tissues were seen to be mostly filled in the defect areas in the control group ( A – C ). The group treated with SA hydrogel as the only grafted material showing fibrous and adipose tissue ( D – F ). lacuna spaces were observed in the newly formed lamellar bone in NHH with a wide bone marrow space ( G – I ). DDMH-treated groups showed considerable new bone trabeculae formation ( J – L ). BT: bone trabeculae; FT: fibrous tissue. Arrows represent MT- and OCN-positive reactions. Scale bar: 100 μm.

Article Snippet: Subsequently, the samples were embedded in paraffin; and after staining with hematoxylin and eosin (H&E), Masson’s trichrome (MT) (Cat# HT15, Sigma Aldrich), and immunohistochemical staining with a polyclonal anti-rabbit osteocalcin antibody (OCN; Cat# GB11233, diagnostic biosystems) at a 1:150 dilution, histological examination was conducted using an Olympus BX-51 optical microscope (Olympus Co., Tokyo, Japan).

Techniques: Staining, Control

Journal: Dentistry Journal

Article Title: In Vivo Evaluation of Regenerative Osteogenic Potential Using a Human Demineralized Dentin Matrix for Dental Application

doi: 10.3390/dj12030076

Figure Lengend Snippet: Representative histological images of (H&E) ( A , D , G , J ), Masson’s trichrome ( B , E , H , K ) and osteocalcin immune staining ( C , F , I , L ) are shown at 8 weeks post-treatment. Rather than the newly created bone, the defect locations in the control group were found to be primarily filled with fibrous connective tissues and adipose tissue ( A – C ). Circular new bony islands with the presence of early woven bone were observed in the group treated with SA as the only grafted material ( D – F ). Newly formed lamellar bone was observed in NHH with a wide bone marrow space ( G – I ). In contrast, DDMH-treated groups showed considerable new bone formation at 8 weeks with relatively narrow marrow spaces ( J – L ). BT means bone trabeculae; FT: fibrous tissue. Arrows represent MT- and OCN-positive reactions. Scale bar: 100 μm.

Article Snippet: Subsequently, the samples were embedded in paraffin; and after staining with hematoxylin and eosin (H&E), Masson’s trichrome (MT) (Cat# HT15, Sigma Aldrich), and immunohistochemical staining with a polyclonal anti-rabbit osteocalcin antibody (OCN; Cat# GB11233, diagnostic biosystems) at a 1:150 dilution, histological examination was conducted using an Olympus BX-51 optical microscope (Olympus Co., Tokyo, Japan).

Techniques: Staining, Control

Journal: International Journal of Nanomedicine

Article Title: Hollow Hydroxyapatite Microspheres Loaded with rhCXCL13 to Recruit BMSC for Osteogenesis and Synergetic Angiogenesis to Promote Bone Regeneration in Bone Defects

doi: 10.2147/IJN.S408905

Figure Lengend Snippet: Analysis of osteogenic differentiation in BMSCs co-cultured with scaffolds. ( A ) Quantitative RT-qPCR analysis of osteogenic gene expression (Runx2, ALPL, OCN) in BMSCs co-cultured with each group. GAPDH was used as housekeeping genes. The results were expressed as mean ± SD. ( B ) Relative protein expression of RUNX2 and OCN in BMSCs co-cultured with various treatment groups. ( C ) Representative images of Alizarin red-stained BMSCs co-cultured with various scaffolds for 21 days in 6-well plates (scale bar = 100μm). ( D ) Quantitative analysis of Alizarin Red Staining in BMSCs co-cultured with various treatment groups. Statistical analysis was performed using ANOVA and LSD tests, with results expressed as mean ± SD (*p < 0.05, **p < 0.01, ***p < 0.001, n=3).

Article Snippet: After the washing, membranes were incubated with rabbit anti-RUNX2 (Abclonal; 1:1000), rabbit anti-OCN (Abclonal; 1:1000), and rabbit anti-GAPDH (Abclonal; 1:1000) antibodies at 4◦C overnight.

Techniques: Cell Culture, Quantitative RT-PCR, Expressing, Staining

Journal: International Journal of Nanomedicine

Article Title: Hollow Hydroxyapatite Microspheres Loaded with rhCXCL13 to Recruit BMSC for Osteogenesis and Synergetic Angiogenesis to Promote Bone Regeneration in Bone Defects

doi: 10.2147/IJN.S408905

Figure Lengend Snippet: Immunofluorescence Staining Analysis of Osteogenic Differentiation in BMSCs Co-Cultured with Scaffolds. ( A ) Immunofluorescence staining of BMSCs (COLI, BMP-2, RUNX2, and OCN) when co-cultured with scaffolds (( A and B ). ( B ) Quantitative analysis of immunofluorescence staining for COLI, BMP-2, RUNX2, and OCN performed by Image J. ( a1, a2, b1 and b2 ). Statistical analysis was performed using ANOVA and LSD tests, with results expressed as mean ± SD (*p<0.05, **p<0.01, ***p<0.001, n=3).

Article Snippet: After the washing, membranes were incubated with rabbit anti-RUNX2 (Abclonal; 1:1000), rabbit anti-OCN (Abclonal; 1:1000), and rabbit anti-GAPDH (Abclonal; 1:1000) antibodies at 4◦C overnight.

Techniques: Immunofluorescence, Staining, Cell Culture

Journal: International Journal of Nanomedicine

Article Title: Hollow Hydroxyapatite Microspheres Loaded with rhCXCL13 to Recruit BMSC for Osteogenesis and Synergetic Angiogenesis to Promote Bone Regeneration in Bone Defects

doi: 10.2147/IJN.S408905

Figure Lengend Snippet: Heat Map, Protein-Protein Interaction (PPI) Network, and KEGG Up-Pathway Enrichment Analyses in Various Treatment Groups. ( A ) Heat Map of differentially expressed genes related to cellular activity, osteogenesis, angiogenesis, and BMSC recruitment in different groups comparisons. ( B1 and B2 ) PPI analysis revealing the function of DEG-encoded proteins in different groups. ( C1-C3 ) KEGG up-pathway enrichment analysis in different groups comparisons. The red symbol in ( C1 ) indicates that the KEGG up-pathway enrichment analysis of differentially expressed genes mainly concentrated in the PI3K-AKT signaling pathway. ( D ) Changes in the relative gene expression levels of RUNX2 and OCN after treatment with PI3K-AKT inhibitor BEZ235 in BMSCs from all groups. ( E ) Representative alizarin red staining image of BMSCs co-cultured with rhCXCL13-HHM/CS scaffold after 21 days of osteogenic differentiation treated with PI3K-AKT inhibitor BEZ235 (scale bar = 100μm, n=3). ( F ) Quantitative analysis of alizarin red staining of BMSCs in different treatment groups (***p<0.001, n=3).

Article Snippet: After the washing, membranes were incubated with rabbit anti-RUNX2 (Abclonal; 1:1000), rabbit anti-OCN (Abclonal; 1:1000), and rabbit anti-GAPDH (Abclonal; 1:1000) antibodies at 4◦C overnight.

Techniques: Activity Assay, Expressing, Staining, Cell Culture