nf κb activator prostratin  (MedChemExpress)

Journal: Stem Cell Reviews and Reports

Article Title: Equine Embryonic Stem Cell-Derived Tenocytes are Insensitive to a Combination of Inflammatory Cytokines and Have Distinct Molecular Responses Compared to Primary Tenocytes

doi: 10.1007/s12015-024-10693-8

Figure Lengend Snippet: Experimental study design outlining ESC-tenocyte transcriptomic analysis in response to IL-1β ( a ), effect of inflammatory cytokine stimulation on equine ESC-tenocytes ( b ), inflammatory pathway activation ( c ), inflammatory cytokine receptor expression analysis ( d ), activation of NF-κB through pharmaceuticals ( e ), and ESC-tenocyte conditioned media rescue ( f ). Created with BioRender.com

Article Snippet: The plate with attached lid was then sealed with parafilm and incubated at 37 °C for 60–90 min. Once set, the 3-D tendon constructs were cultured in growth media alone (unstimulated control) or supplemented with recombinant human TNFα (10 ng/mL), recombinant human IL-1β (17 ng/mL), and/or recombinant equine IFN-γ (100 ng/mL; all PeproTech) [ ], or NF-κB activators Prostratin (2 μM) and Phorbol 12-myristate 13-acetate (PMA) (10 ng/mL; both Abcam, Cambridge, UK).

Techniques: Activation Assay, Expressing

Journal: Stem Cell Reviews and Reports

Article Title: Equine Embryonic Stem Cell-Derived Tenocytes are Insensitive to a Combination of Inflammatory Cytokines and Have Distinct Molecular Responses Compared to Primary Tenocytes

doi: 10.1007/s12015-024-10693-8

Figure Lengend Snippet: The localisation and activation of inflammatory pathway proteins in response to inflammatory cytokine stimulation. Immunofluorescence staining of NF-κB P65 (a-a’’’’’), STAT1 (b-b’’’’’), and JNK (c–c’’’’’) in ESC-tenocytes following stimulation with IFN-γ, TNFα, and/or IL-1β. Unstimulated cells served as a control. Nuclear staining is shown by DAPI in blue. All images are representative of three independent biological replicates. Scale bar = 50 μm. Semi-quantitative analysis of the relative nuclear fluorescent intensity of NF-κB P65 (d), STAT1 (e), and JNK (f) following inflammatory cytokine stimulation is also shown. Data is displayed as fold change compared to the no cytokine control. Error bars depict the S.E.M of three measurements from three independent biological replicates of ESC-tenocytes. The asterisk (*) denotes the fold change is significantly different to the no cytokine control at p < 0.05. Cells in these experiments were between P9 and P21

Article Snippet: The plate with attached lid was then sealed with parafilm and incubated at 37 °C for 60–90 min. Once set, the 3-D tendon constructs were cultured in growth media alone (unstimulated control) or supplemented with recombinant human TNFα (10 ng/mL), recombinant human IL-1β (17 ng/mL), and/or recombinant equine IFN-γ (100 ng/mL; all PeproTech) [ ], or NF-κB activators Prostratin (2 μM) and Phorbol 12-myristate 13-acetate (PMA) (10 ng/mL; both Abcam, Cambridge, UK).

Techniques: Activation Assay, Immunofluorescence, Staining

Journal: Stem Cell Reviews and Reports

Article Title: Equine Embryonic Stem Cell-Derived Tenocytes are Insensitive to a Combination of Inflammatory Cytokines and Have Distinct Molecular Responses Compared to Primary Tenocytes

doi: 10.1007/s12015-024-10693-8

Figure Lengend Snippet: NF-κB signalling can be activated in ESC-tenocytes through NF-κB activator pharmaceuticals. Immunofluorescence staining of NF-κB P65 in adult (a-a’’’’’) and ESC-tenocytes (b-b’’’’’) following 30 min of stimulation with Prostratin (a’’ and b’’) and 1 h stimulation with PMA (a’’’ and b’’’), A23187 (a’’’’ and b’’’’), or Betulinic Acid (a’’’’’ and b’’’’’). DAPI staining of the nucleus is shown in blue. Images are representative of three biological replicates. Scale bar = 50 μm. Semi-quantitative analysis of the relative nuclear fluorescent intensity of NF-κB P65 following NF-κB activator stimulation in adult (c) and ESC-tenocytes (d) is shown. Data is displayed as fold change compared to the unstimulated control. Error bars represent the S.E.M of three measurements from three independent biological replicates. The asterisk (*) denotes the fold change is significantly different to the unstimulated at p < 0.05. Adult tenocytes in these experiments were between P3 and P6 and ESC-tenocytes between P9 and P33

Article Snippet: The plate with attached lid was then sealed with parafilm and incubated at 37 °C for 60–90 min. Once set, the 3-D tendon constructs were cultured in growth media alone (unstimulated control) or supplemented with recombinant human TNFα (10 ng/mL), recombinant human IL-1β (17 ng/mL), and/or recombinant equine IFN-γ (100 ng/mL; all PeproTech) [ ], or NF-κB activators Prostratin (2 μM) and Phorbol 12-myristate 13-acetate (PMA) (10 ng/mL; both Abcam, Cambridge, UK).

Techniques: Immunofluorescence, Staining

Journal: Stem Cell Reviews and Reports

Article Title: Equine Embryonic Stem Cell-Derived Tenocytes are Insensitive to a Combination of Inflammatory Cytokines and Have Distinct Molecular Responses Compared to Primary Tenocytes

doi: 10.1007/s12015-024-10693-8

Figure Lengend Snippet: NF-κB activators cause changes to 2-D gene expression but not 3-D collagen gel contraction in adult and ESC-tenocytes. Fold change in gene expression of adult ( a ) and ESC-tenocytes ( b ) following 72 h stimulation with Prostratin and PMA compared to the unstimulated control. Secretion of IL-6 is significantly increased by Prostratin and PMA in adult tenocytes ( c ) but not in ESC-tenocytes ( d ). Error bars represent the S.E.M of three biological replicates. In 3-D culture, neither Prostratin or PMA inhibited the degree of collagen gel contraction by adult ( e ) or ESC-tenocytes ( f ). Error bars here represent the S.E.M of five biological replicates of adult tenocytes and three biological replicates of ESC-tenocytes. The asterisk (*) denotes a significant difference to the unstimulated control at p < 0.05. Adult tenocytes in these experiments were used between P2 and P8 and ESC-tenocytes between P9 and P31

Article Snippet: The plate with attached lid was then sealed with parafilm and incubated at 37 °C for 60–90 min. Once set, the 3-D tendon constructs were cultured in growth media alone (unstimulated control) or supplemented with recombinant human TNFα (10 ng/mL), recombinant human IL-1β (17 ng/mL), and/or recombinant equine IFN-γ (100 ng/mL; all PeproTech) [ ], or NF-κB activators Prostratin (2 μM) and Phorbol 12-myristate 13-acetate (PMA) (10 ng/mL; both Abcam, Cambridge, UK).

Techniques: Expressing