cat n dy460 uk  (R&D Systems)

Journal: Stem Cell Research & Therapy

Article Title: G protein-coupled receptor kinase 3 modulates mesenchymal stem cell proliferation and differentiation through sphingosine-1-phosphate receptor regulation

doi: 10.1186/s13287-022-02715-4

Figure Lengend Snippet: Grk3-/- BmMSCs have higher levels of CXCL12 detected in culture and proliferate more in comparison with WT BmMSCs but secrete similar amounts of CXCL12 per cell. ( A ) Quantification of CXCL12 protein concentration from supernatant of BmMSC cultures. Lower limit of detection at 46.9 ρg/mL. Data represent mean ± SEM, n = 4, passages 11 and 12. ( B ) BmMSC proliferation determined by increased formazan dye production from viable cells over time using CCK-8 proliferation assay. Data represent mean ± SEM, n = 3, except day 0 n = 2, passage 7. * P ≤ 0.05 ** P ≤ 0.01 *** P ≤ 0.001, **** P ≤ 0.0001 ( C ) CXCL12 per MSC determined by trypsinized cell count using trypan blue exclusion of individual wells divided by the amount of CXCL12 (pg/ml) measured by ELISA. Data represent mean ± SEM, n = 11 Grk3-/-, n = 10 WT, passages 10–13

Article Snippet: Supernatants of undifferentiated BmMSC monolayers were collected and analyzed for CXCL12 protein using the CXCL12 DuoSet ELISA kit (R&D Systems, DY460), as per instructions.

Techniques: Comparison, Protein Concentration, CCK-8 Assay, Proliferation Assay, Cell Counting, Enzyme-linked Immunosorbent Assay

Journal: Stem Cell Research & Therapy

Article Title: G protein-coupled receptor kinase 3 modulates mesenchymal stem cell proliferation and differentiation through sphingosine-1-phosphate receptor regulation

doi: 10.1186/s13287-022-02715-4

Figure Lengend Snippet: Proposed model for GRK3-deficient BmMSCs within the niche. ( A ) Our studies suggest BmMSCs deficient in GRK3 secrete more CXCL12 and proliferate more rapidly, which may increase support to hematopoietic stem and progenitor cell (HSPC) development. ( B , top) We propose GRK3 desensitizes S1PR and recruits β-arrestin to affect ERK1/2 signaling and receptor internalization. ( B , bottom) Grk3-/- BmMSCs have enhanced functions of proliferation and osteogenic differentiation, and this may be mediated through decreased S1PR1 internalization (i.e., more surface expression) and prolonged late-phase ERK1/2 activation

Article Snippet: Supernatants of undifferentiated BmMSC monolayers were collected and analyzed for CXCL12 protein using the CXCL12 DuoSet ELISA kit (R&D Systems, DY460), as per instructions.

Techniques: Expressing, Activation Assay