Journal: Nanomaterials

Article Title: Polarization of Femtosecond Laser for Titanium Alloy Nanopatterning Influences Osteoblastic Differentiation

doi: 10.3390/nano12101619

Figure Lengend Snippet: PCR primer sequences of genes implicated in different osteogenic pathways.

Article Snippet: At 24 h post seeding, the growth medium was replaced with an osteogenic medium (MSCODM, C-28013, PromoCell).

Techniques:

Journal: Nanomaterials

Article Title: Polarization of Femtosecond Laser for Titanium Alloy Nanopatterning Influences Osteoblastic Differentiation

doi: 10.3390/nano12101619

Figure Lengend Snippet: Osteogenic-related genes analyzed by RT-qPCR for cells present on polished, linear LIPSS and radial LIPSS surfaces at 14 d post seeding. Genes are classified by biological function. Values are given as ratios to GAPDH. The graphs give p -values between each group, in bold when <0.05, Mann–Whitney U test, n = 4–6 samples/group.

Article Snippet: At 24 h post seeding, the growth medium was replaced with an osteogenic medium (MSCODM, C-28013, PromoCell).

Techniques: Quantitative RT-PCR, MANN-WHITNEY

Journal: Dose-Response

Article Title: Skeletal Stem Cells Rescue Radiation-Induced Osteogenic Precursor Cell Dysfunction via the Wnt/β-Catenin Signaling Pathway

doi: 10.1177/15593258261440983

Figure Lengend Snippet: Co-culture with SSCs rescues the function of irradiated osteogenic precursor cells. (A, B) Cell apoptosis was analyzed by flow cytometry with Annexin V-PE/7AAD double staining. (A) Representative flow cytometry plots. (B) Quantitative analysis of the apoptotic rate. (C, D) ALP activity was assessed. (C) Representative ALP staining images (Scale bar: 50 μm). (D) Quantitative analysis of the relative ALP activity. (E, F) Mineralization capacity was evaluated using Alizarin Red S staining. (E) Representative staining images of mineralized nodules (Scale bar: 100 μm). (F) Quantitative analysis of the relative mineralization level. (G, H) Cell migration was determined by a migration assay. (G) Representative images of migrated cells (Scale bar: 50 μm). (H) Quantitative analysis of the relative cell migration level. All data are presented as mean ± SD, with statistical significance determined by unpaired two-tailed Student’s t-test (* p < 0.05; ** p < 0.01).

Article Snippet: After irradiation and corresponding interventions, cells were cultured in osteogenic induction medium (iXCells Biotechnologies, San Diego, CA, Cat. No. MD-0006) for 7 days.

Techniques: Co-Culture Assay, Irradiation, Flow Cytometry, Double Staining, Activity Assay, Staining, Migration, Two Tailed Test

Journal: Dose-Response

Article Title: Skeletal Stem Cells Rescue Radiation-Induced Osteogenic Precursor Cell Dysfunction via the Wnt/β-Catenin Signaling Pathway

doi: 10.1177/15593258261440983

Figure Lengend Snippet: SSCs exert rescue effects via the Wnt/β-catenin signaling pathway. (A) Representative ALP staining images of cells in each group (Scale bar: 50 μm). (B) Quantitative analysis of ALP activity in each group. (C) Representative Alizarin Red S staining images of cells in each group (Scale bar: 100 μm). (D) Quantitative analysis of Alizarin Red S staining in each group. (E) Relative mRNA expression levels of osteogenic marker genes ( Runx2 , Col1a1 , and OCN ) detected by qRT-PCR. GAPDH was used as an internal reference gene. (F) Representative Western blot images showing the expression levels of RUNX2, COL1A1, OCN, and β-catenin in each group. GAPDH was used as a loading control. (G) Quantitative analysis of Western blot results (gray value ratio of target protein to GAPDH) in each group. All data are presented as mean ± SD, with statistical significance determined by unpaired two-tailed Student’s t-test (* p < 0.05; ** p < 0.01; *** p < 0.001).

Article Snippet: After irradiation and corresponding interventions, cells were cultured in osteogenic induction medium (iXCells Biotechnologies, San Diego, CA, Cat. No. MD-0006) for 7 days.

Techniques: Staining, Activity Assay, Expressing, Marker, Quantitative RT-PCR, Western Blot, Control, Two Tailed Test

Journal: Dose-Response

Article Title: Skeletal Stem Cells Rescue Radiation-Induced Osteogenic Precursor Cell Dysfunction via the Wnt/β-Catenin Signaling Pathway

doi: 10.1177/15593258261440983

Figure Lengend Snippet: SSCs alleviate the radiation-induced bone injury in mice. (A–G) Micro-CT analysis of bone microstructure. (A) Representative micro-CT images of femurs. Quantitative analysis of (B) bone mineral density (BMD), (C) bone volume fraction (BV/TV), (D) trabecular thickness (Tb.Th), (E) trabecular number (Tb.N), (F) connectivity density (Conn.D), and (G) trabecular separation (Tb.Sp) at 2- and 4-weeks post irradiation. (H–K) Histological analysis (Scale bar: 100 μm). (H) H&E staining showing steatosis (arrows) and (I) quantitative analysis of steatotic lesions per field. (J) TRAP staining showing osteoclasts (arrows) and (K) quantitative analysis of osteoclast number per field. (L–O) Immunohistochemical staining of osteogenic markers (Scale bar: 100 μm). (L) Osterix staining and (M) quantitative analysis of Osterix-positive area. (N) β-catenin staining and (O) quantitative analysis of β-catenin-positive area. All experiments were conducted in three groups: Control, irradiation (IR), and IR plus SSC (IR+SSC) at 2- and 4-weeks post-irradiation. All data are presented as mean ± SD, with statistical significance determined by unpaired two-tailed Student’s t-test (* p < 0.05; ** p < 0.01; *** p < 0.001)

Article Snippet: After irradiation and corresponding interventions, cells were cultured in osteogenic induction medium (iXCells Biotechnologies, San Diego, CA, Cat. No. MD-0006) for 7 days.

Techniques: Micro-CT, Irradiation, Staining, Immunohistochemical staining, Control, Two Tailed Test

Journal: The Journal of Biological Chemistry

Article Title: Periosteum-derived podoplanin-expressing stromal cells regulate nascent vascularization during epiphyseal marrow development

doi: 10.1016/j.jbc.2022.101833

Figure Lengend Snippet: SSCs generate PDPN-expressing progenies with a phenotype identical to that of epiphyseal PDPN-expressing stromal cells. A , representative flow cytometric data of in vitro SSC progenies generated from primary isolated SSCs during culture with MesenCult. B , PDPN and Sca-1 expression in in vitro generated SSC progenies. C – E , representative ICC images of the in vitro PDPN-expressing SSC progenies stained with PDPN/PDGFRβ/DAPI ( C ), PDPN/NG2/DAPI ( D ), and PDPN/αSMA/DAPI ( E ). Scale bars indicate 50 μm. αSMA, α-smooth muscle actin; DAPI, 4′,6-diamidino-2-phenylindole; ICC, immunocytochemistry; NG2, neuron-glial antigen-2; PDGFRβ, platelet-derived growth factor receptor-β; Sca-1, stem cell antigen-1; PDPN, podoplanin; SSC, skeletal stem cell.

Article Snippet: After 24 h, the culture medium was replaced with the Complete mouse MesenCult Osteogenic Medium (Stem Cell Technologies) containing L-glutamine (Wako) and penicillin/streptomycin/amphotericin B (Wako), and the cells were cultured for 12 days.

Techniques: Expressing, In Vitro, Generated, Isolation, Staining, Immunocytochemistry, Derivative Assay

Journal: Journal of Tissue Engineering

Article Title: Therapeutic potential of adipose-derived mesenchymal stem cell exosomes in tissue-engineered bladders

doi: 10.1177/20417314211001545

Figure Lengend Snippet: Identification of Ad-MSCs (a–c) and AMEs (d–f): (a) characterization of the fourth generation of Ad-MSCs was performed by flow cytometry, and the Ad-MSC histogram revealed that after purification, the cells mostly expressed mesenchymal cell markers. Differentiation assays showed that Ad-MSCs could differentiate into adipocytes (b) and osteocytes (c), which were stained with Oil Red O and Von Kossa, respectively. Scale bar = 100 μm. (d) TEM of exosomes isolated from MSCM-cultured Ad-MSCs for 3 days with 10% exosome-depleted FBS. Scale bars = 100 nm. (e) NTA found that most of these vesicles ranged in diameter from 30 to 150 nm. (f) Western blotting was performed with Ad-MSCs (MSCs) and AMEs (Exos). The expression of CD9, CD63, TSG101, and calnexin was detected. Ad-MSCs: adipose-derived mesenchymal stem cells; AMEs: adipose-derived mesenchymal stem cell exosomes; NTA: nanoparticle tracking analysis; TEM: transmission electron microscopy.

Article Snippet: The medium was changed to mesenchymal stem cell adipogenic differentiation medium (MADM, ScienCell) and mesenchymal stem cell osteogenic differentiation medium (MODM, ScienCell) when the cells reached 70–80% confluency.

Techniques: Flow Cytometry, Purification, Staining, Isolation, Cell Culture, Western Blot, Expressing, Derivative Assay, Transmission Assay, Electron Microscopy

Journal: Stem Cell Research & Therapy

Article Title: BMSC-derived exosomes promote tendon-bone healing after anterior cruciate ligament reconstruction by regulating M1/M2 macrophage polarization in rats

doi: 10.1186/s13287-022-02975-0

Figure Lengend Snippet: Morphology and characterization of BMSCs and BMSC-Exos. A Representative light microscopic images of BMSCs (Scale bar: 200 μm). The identification of osteogenic, adipogenic, and chondrogenic differentiation potential of BMSCs, and the representative microscopic image of Alizarin Red S staining ( B ) (Scale bar: 50 μm), Oil Red O staining ( C ) (Scale bar: 100 μm), and Alcian blue staining ( D ) (Scale bar: 100 μm). n = 3. E Schematic diagram of the isolation of the BMSC-Exos. F Transmission electron microscopy (TEM) image of BMSC-Exos. Scale bar: 100 nm. G NanoSight tracking analysis (NTA) for the diameters of BMSC-Exo. H Western blot analysis of CD9, CD63, TSG101, and Calnexin. n = 3. I Analysis of cellular exosome uptake by macrophages. n = 3. Scale bar: 25 μm.

Article Snippet: Osteogenic differentiation medium, adipogenic differentiation medium, and chondrogenic differentiation medium were purchased from Procell Life Science & Technology Co, Ltd (Procell, Wuhan, China).1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate (DiI) and 4′,6-diamidino-2-phenylindole (DAPI) were purchased from Beyotime (Shanghai, China).

Techniques: Staining, Isolation, Transmission Assay, Electron Microscopy, Western Blot

Journal: Clinical & Translational Immunology

Article Title: Human placenta-derived adherent cells induce tolerogenic immune responses

doi: 10.1038/cti.2014.5

Figure Lengend Snippet: PDAC cells display MSC-like characteristics. ( a ) PDAC cells from two donors show spindle-shaped fibroblast morphology under phase contrast microscope after 6 passages of culture expansion ex vivo . ( b ) Flow cytometry analysis of PDAC-cell phenotype. Top row: expression of MSC markers (CD73, CD90, CD105) and placenta marker (CD200). Bottom row: expression of hematopoietic cell surface markers. Dotted lines indicate isotype-matched control antibody staining. Numbers indicate percent positive cells. Results from one of the six independent experiments are shown. ( c ) Adipogenic (Oil red O staining), osteogenic (alizarin red staining) and chondrogenic (Alcian Blue and Sirius Red staining) differentiation of PDAC cells under different lineage differentiation culture conditions.

Article Snippet: For adipogenic and osteogenic differentiation, the PDAC cell expansion medium was changed to adipogenic (DMEM high glucose medium supplemented with 10% Fetal Calf Serum, 10 μg ml −1 insulin, 1 μ M dexmethasone, 0.1 m M indomethacin, 0.5 m M isobutylmethylxanthine and 100 IU ml −1 Pen-Strep) or osteogenic induction medium (Cambrex, East Rutherford, NJ, USA).

Techniques: Microscopy, Ex Vivo, Flow Cytometry, Expressing, Marker, Control, Staining