human bmscs #cp-h166  (Procell Inc)

Journal: Advanced Science

Article Title: Tensile Stress‐Activated and Exosome‐Transferred YAP/TAZ‐Notch Circuit Specifies Type H Endothelial Cell for Segmental Bone Regeneration

doi: 10.1002/advs.202309133

Figure Lengend Snippet: Tensile stress (TS)‐induced type H endothelial cells (THECs) phenotype and the involvement of YAP/TAZ signaling. A,B) Flow cytometry A) and quantitative analysis B) of THECs phenotype after mechanical TS stimulation on bone marrow endothelial cells (BMECs). C) Concentrations of FGF‐1 and TGF‐β1 in conditioned medium (CM) from TS‐stimulated BMECs. D,E) Alkaline phosphatase (ALP) staining D) and activity measurement E) of bone marrow mesenchymal stem cells (BMSCs) after seven days of osteogenic induction with CM treatment. F,G) Alizarin red S staining F) and quantitative analysis G) of calcium nodules after fourteen days of osteogenic induction with CM treatment. H,I) Immunofluorescence staining of OSX H) and OCN I) for the evaluation of osteogenic differentiation of BMSCs. J,K) Transwell assay images J) and quantitative analysis K) revealing the chemotaxis of BMSCs in the upper chambers towards BMECs in the lower chambers. L) The gene expression of Notch1/Dll4 and YAP/TAZ pathways in TS‐stimulated BMECs. M,N) Western blot images K) and quantitative analysis L) revealing the expression of Notch1/Dll4 and YAP/TAZ pathways in TS‐stimulated BMECs. O) Fluorescence‐activated cell sorting (FACS) for respectively isolating type L endothelial cells (TLECs) and THECs from TS‐stimulated BMECs. P,Q) Immunofluorescence staining P) and quantitative analysis Q) for evaluating the expression and nuclear translocation of YAP in FACS‐sorted TLECs and THECs. R) Concentrations of FGF‐1 and TGF‐β1 in CM from FACS‐sorted TLECs and THECs. S,T) ALP staining S) and activity measurement T) of BMSCs after osteogenic induction with CM treatment. U,V) Alizarin red S staining U) and quantitative analysis V) of calcium nodules of BMSCs in different groups. W,X) Immunofluorescence staining of OSX W) and OCN X) for the evaluation of osteogenic differentiation of BMSCs. Y,Z) Transwell assay images Y) and quantitative analysis Z) revealing the chemotaxis of BMSCs towards FACS‐sorted TLECs and THECs. ns p > 0.05, ** p < 0.01.

Article Snippet: Human bone marrow mesenchymal stem cells (BMSCs; CP‐H166, Procell) were cultured in Minimal Essential Medium Alpha (SH30265.01, HyClone) supplemented with 10% fetal bovine serum (10099‐141, Gibco) and 1% penicillin‐streptomycin (15140122, Gibco).

Techniques: Flow Cytometry, Staining, Activity Assay, Immunofluorescence, Transwell Assay, Chemotaxis Assay, Gene Expression, Western Blot, Expressing, Fluorescence, FACS, Translocation Assay

Journal: Advanced Science

Article Title: Tensile Stress‐Activated and Exosome‐Transferred YAP/TAZ‐Notch Circuit Specifies Type H Endothelial Cell for Segmental Bone Regeneration

doi: 10.1002/advs.202309133

Figure Lengend Snippet: Type H endothelial cells (THECs) phenotype and expression of Notch/Dll4 and YAP/TAZ pathways after mechano‐biological transduction disruption. A,B) Flow cytometry A) and quantitative analysis B) of THECs phenotype after GsMTx4 treatment or knockdown of YAP or Notch1 by siRNAs on tensile stress (TS)‐stimulated bone marrow endothelial cells (BMECs). C) Concentrations of FGF‐1 and TGF‐β1 in conditioned medium (CM) from TS‐stimulated BMECs in different treatment groups. D,E) Alkaline phosphatase (ALP) staining D) and activity measurement E) of bone marrow mesenchymal stem cells (BMSCs) after seven days of osteogenic induction with CM treatment. F,G) Alizarin red S staining F) and quantitative analysis G) of calcium nodules after fourteen days of osteogenic induction with CM treatment. H,I) Immunofluorescence staining of OSX H) and OCN I) for the evaluation of osteogenic differentiation of BMSCs. J,K) Transwell assay images J) and quantitative analysis K) revealing the chemotaxis of BMSCs in the upper chambers towards BMECs in the lower chambers. L) The gene expression of Notch1/Dll4 and YAP/TAZ pathways in TS‐stimulated BMECs upon different treatments. M,N) Western blot images M) and quantitative analysis N) revealing the expression of Notch1/Dll4 and YAP/TAZ pathways in TS‐stimulated BMSCs upon different treatments. ns p > 0.05, * p < 0.05, ** p < 0.01.

Article Snippet: Human bone marrow mesenchymal stem cells (BMSCs; CP‐H166, Procell) were cultured in Minimal Essential Medium Alpha (SH30265.01, HyClone) supplemented with 10% fetal bovine serum (10099‐141, Gibco) and 1% penicillin‐streptomycin (15140122, Gibco).

Techniques: Expressing, Transduction, Disruption, Flow Cytometry, Knockdown, Staining, Activity Assay, Immunofluorescence, Transwell Assay, Chemotaxis Assay, Gene Expression, Western Blot

Journal: Advanced Science

Article Title: Tensile Stress‐Activated and Exosome‐Transferred YAP/TAZ‐Notch Circuit Specifies Type H Endothelial Cell for Segmental Bone Regeneration

doi: 10.1002/advs.202309133

Figure Lengend Snippet: Tensile stress‐stimulated bone marrow endothelial cells (BMECs)‐derived exosomes (Exo TS ) for transmitting the YAP/TAZ‐Notch1/Dll4 circuit to induce type H endothelial cells (THECs) phenotype. A) Transmission electron microscopy images showing the characteristic morphology of exosomes. B) Nanoparticle tracking analysis showing the distribution of particle size of exosomes. C) Western blot analysis showing the exosomal marker expression of BMECs and their exosomes in different groups. D,E) Western blot images D) and quantitative analysis E) revealing the content of exosomal proteins involved in YAP/TAZ‐Notch1/Dll4 circuit. F) Fluorescence images of BMECs after uptake of PKH67‐labeled exosomes. G,H) Western blot images G) and quantitative analysis H) revealing the expression of YAP/TAZ‐Notch1/Dll4 circuit in exosomes‐treated BMECs. I,J) Flow cytometry I) and quantitative analysis J) of THECs phenotype on exosomes‐treated BMECs. K,L) Immunofluorescence staining K) and quantitative analysis L) for evaluating the expression and nuclear translocation of YAP in exosomes‐treated BMECs. M) Concentrations of FGF‐1 and TGF‐β1 in conditioned medium (CM) from exosomes‐treated BMECs. N,O) Alkaline phosphatase (ALP) staining N) and activity measurement O) of bone marrow mesenchymal stem cells (BMSCs) after seven days of osteogenic induction with CM treatment. P,Q) Alizarin red S staining P) and quantitative analysis Q) of calcium nodules after fourteen days of osteogenic induction with CM treatment. R,S) Immunofluorescence staining of OSX R) and OCN S) for the evaluation of osteogenic differentiation of BMSCs. T,U) Transwell assay images T) and quantitative analysis U) revealing the chemotaxis of BMSCs in the upper chambers towards BMECs in the lower chambers. ns p > 0.05, * p < 0.05, ** p < 0.01.

Article Snippet: Human bone marrow mesenchymal stem cells (BMSCs; CP‐H166, Procell) were cultured in Minimal Essential Medium Alpha (SH30265.01, HyClone) supplemented with 10% fetal bovine serum (10099‐141, Gibco) and 1% penicillin‐streptomycin (15140122, Gibco).

Techniques: Derivative Assay, Transmission Assay, Electron Microscopy, Western Blot, Marker, Expressing, Fluorescence, Labeling, Flow Cytometry, Immunofluorescence, Staining, Translocation Assay, Activity Assay, Transwell Assay, Chemotaxis Assay