thp 1 dual cells  (InvivoGen)

Journal: eLife

Article Title: Suppression of interferon signaling via small-molecule modulation of TFAM

doi: 10.7554/eLife.108742

Figure Lengend Snippet: ( A ) Flowchart of the CXCL-10 experiment. ( B ) Compounds 2 , 3 , and 15 repressed ISG signaling in CXCL-10 production. THP-1 cells were pre-treated with individual compounds 48 h prior to 100 ng/mL TNFa stimulation for 48 h. CXCL-10 levels were measured. ( C ) Flowchart of the ISRE experiment. ( D ) Compounds 2 , 3 , and 15 repressed ISG signaling in IRF3 production. THP-1-Dual cells were pre-treated with individual compounds 24 h prior to 100 ng/mL TNFa stimulation for 48 h. The inhibition of type I IFN response was monitored via ISRE reporter activation. Figures are representatives of at least two independent experiments. Graph shows one representative experiment of two independent experiments. Error bars represent ± SD from n=3 biological replicates. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001. Source data for this figure are available in (raw data and analysis). Figure 2—source data 1. Raw data and analysis results to generate the graphs shown in .

Article Snippet: THP-1-Dual cells (thpd-nifs, 1 × 10 6 /mL) treated with 100 ng/mL TNF (Biolegend, 575204) for 48 h, or cGAMP (InvivoGen, tlrl-nacga23-02) for 24 h to induce a cGAS-STING-dependent interferon response.

Techniques: Inhibition, Activation Assay

Journal: eLife

Article Title: Suppression of interferon signaling via small-molecule modulation of TFAM

doi: 10.7554/eLife.108742

Figure Lengend Snippet: ( A ) Chemical structures of hit compounds and related analogs. ( B ) TFAM modulators were profiled in the ISRE assay and display dose-dependent suppression of ISG signaling. THP-1-Dual cells were pre-treated with individual compounds 24 h prior to 100 ng/mL TNFa stimulation for 48 h. ISRE reporter activation was measured. Figures are representatives of at least two independent experiments. Graph shows one representative experiment of two independent experiments. Error bars represent ± SD from n=3 biological replicates. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001. Source data for this figure are available in (raw data and analysis). Figure 3—source data 1. Raw data and analysis results to generate the graphs shown in .

Article Snippet: THP-1-Dual cells (thpd-nifs, 1 × 10 6 /mL) treated with 100 ng/mL TNF (Biolegend, 575204) for 48 h, or cGAMP (InvivoGen, tlrl-nacga23-02) for 24 h to induce a cGAS-STING-dependent interferon response.

Techniques: Activation Assay

Journal: eLife

Article Title: Suppression of interferon signaling via small-molecule modulation of TFAM

doi: 10.7554/eLife.108742

Figure Lengend Snippet: ( A ) ISRE reporter activation of THP-1 dual cells stimulated with 100 ng/mL TNFa for 48 h. Compounds were added to the cells 5 min before the assay detection. Figures are representatives of at least two independent experiments. Graph shows one representative experiment of two independent experiments. Error bars represent ± SD from n=3 biological replicates. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001. Source data for this figure are available in (raw data and analysis).

Article Snippet: THP-1-Dual cells (thpd-nifs, 1 × 10 6 /mL) treated with 100 ng/mL TNF (Biolegend, 575204) for 48 h, or cGAMP (InvivoGen, tlrl-nacga23-02) for 24 h to induce a cGAS-STING-dependent interferon response.

Techniques: Activation Assay

Journal: eLife

Article Title: Suppression of interferon signaling via small-molecule modulation of TFAM

doi: 10.7554/eLife.108742

Figure Lengend Snippet: ( A ) Compounds 2 , 3 , and 15 do not repress cGAMP induced ISG signaling. THP-1-Dual cells were pre-treated with individual compounds 48 h prior to 10 ug/mL cGAMP stimulation for 24 h. ISRE reporter activation was measured. ( B ) Compounds 2 , 3 , and 11 impart a dose-dependent increase in TFAM protein levels. Immunoblot analysis of TFAM from T47D cells treated with indicated compounds for 5 days. ( C ) Compounds exhibit minimal impact on TFAM mRNA levels. ( D ) Downregulation of TFAM attenuates the function of compounds 2 and 3 in repression of ISG signaling. THP-1-Dual cells were treated with individual compounds 24 h after siRNA transfection. After incubation for 24 h, THP-1-Dual cells were stimulated with 100 ng/mL TNFa for another 48 h. ISRE reporter activation was measured and normalized to protein concentration. ( E ) Compound 3 inhibits mtDNA cytosolic leakage. THP-1 cells were pre-treated with individual compounds 72 h prior to 100 ng/mL TNFa stimulation for 48 h. Cytosolic mtDNA was extracted and quantified using a qPCR assay. Figures are representatives of at least two independent experiments. Graph shows one representative experiment of two independent experiments. Error bars represent ± SD from n=3 biological replicates. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001. Source data for this figure are available in (original uncropped blots) and (annotated uncropped blots), and (raw data and analysis). Figure 4—source data 1. Original uncropped western blot images for . Figure 4—source data 2. Annotated uncropped western blot images for , with treatment conditions and protein identities indicated. Figure 4—source data 3. Raw data and analysis results to generate the graphs shown in .

Article Snippet: THP-1-Dual cells (thpd-nifs, 1 × 10 6 /mL) treated with 100 ng/mL TNF (Biolegend, 575204) for 48 h, or cGAMP (InvivoGen, tlrl-nacga23-02) for 24 h to induce a cGAS-STING-dependent interferon response.

Techniques: Activation Assay, Western Blot, Transfection, Incubation, Protein Concentration