human cgas inhibitor g140  (InvivoGen)

Journal: Journal of Translational Medicine

Article Title: Piezo1 mediates hypoxia-induced endometriosis fibrosis via the mtDNA -dependent cGAS-STING pathway

doi: 10.1186/s12967-026-08255-2

Figure Lengend Snippet: Hypoxic activation of the cGAS-STING pathway relies on mtDNA leakage. (A) Mitochondrial morphological alterations in HESCs under varying durations of hypoxia, shown by immunofluorescence (Scale bar: 20 μm). (B) JC-1 staining of mitochondrial membrane potential in HESCs following hypoxic treatment (Scale bar: 25 μm). (C , D) Quantitative flow cytometric analysis of JC-1 staining, showing changes in mitochondrial membrane potential in HESCs under varying durations of hypoxia. (E) Immunofluorescence co-staining for TOM20 (mitochondria) and dsDNA was performed to visualize mtDNA leakage in control versus hypoxic HESCs. Shown are representative images (Scale bar:20 μm). (F) qRT-PCR analysis of cytosolic mtDNA in control and hypoxic HESCs. (G–I) Western blotting and qRT-PCR analysis was performed to detect the expression levels of HIF-1α, cGAS and STING in the NC, EtBr, Hypoxia and Hypoxia-EtBr groups, along with quantitative analysis. (J , K) Western blotting analysis of TBK1 and IRF3 phosphorylation in HESCs from the NC, EtBr, Hypoxia, and Hypoxia-EtBr groups, along with quantitative analysis. (L , M) Mitochondrial ROS levels assessed by MitoSOX staining in HESCs from Normal, Hypoxia, Hypoxia-EtBr and Hypoxia-NAC groups. Shown are representative images and quantification (Scale bar: 20 μm). (N , O) Immunofluorescence showing α-SMA expression in HESCs from the Normal, Hypoxia, Hypoxia-EtBr and Hypoxia-NAC groups. Representative images and quantification are shown (Scale bar: 100 μm). *P<0.05; **P<0.01; ***P<0.001; ****P<0.0001

Article Snippet: At 60-70% confluency, cells were treated for 24 h with the specified compounds: the Piezo1 agonist Yoda1 (5 μM; MCE, HY-18723); the Piezo1 inhibitor GsMTx4 (5 μM; MCE, HY-P1410); Ethidium bromide (0.2 μg/ml; Macklin, 1239-45-8) to deplete mitochondrial DNA; the calcium chelator BAPTA-AM (4 μM; MCE, HY-100168); the cGAS inhibitor RU.521(2.5μM, MCE, HY-114180 ); the STING inhibitor C-176 (2.5 μM; MCE, HY-112906); the anti-IL6 antibody (2.5 μg/ml, Sigma-Aldrich, I7901); or the ROS inhibitor NAC (2.0mM, MCE, HY-B0215).

Techniques: Activation Assay, Immunofluorescence, Staining, Membrane, Control, Quantitative RT-PCR, Western Blot, Expressing, Phospho-proteomics

Journal: Journal of Translational Medicine

Article Title: Piezo1 mediates hypoxia-induced endometriosis fibrosis via the mtDNA -dependent cGAS-STING pathway

doi: 10.1186/s12967-026-08255-2

Figure Lengend Snippet: Piezo1-mediated Ca²⁺ influx promotes endometrial fibrosis via activation of the cGAS-STING pathway. (A) Immunofluorescence images showing cytosolic and mitochondrial Ca²⁺ levels in HESCs under the indicated conditions: Normoxic control (NC), Hypoxia, Hypoxia-GsMTx4, and Hypoxia-BAPTA AM (Scale bar: 100 μm). (B) Immunofluorescence analysis of mitochondrial morphology in HESCs from the NC, Hypoxia, and Hypoxia with GsMTx4 groups (Scale bar: 20 μm). (C) qRT-PCR analysis quantifying the levels of cytoplasmic mtDNA in NC and Hypoxic HESCs. (D) Representative immunofluorescence images of HESCs co-immunostained for the mitochondrial marker TOM20 and dsDNA, showing mtDNA leakage in the NC, Hypoxia, and Hypoxia-GsMTx4 groups (Scale bar: 20 μm). (E) Immunofluorescence staining analysis of the mitochondrial ROS levels in HESC cell (Scale bar: 20 μm). (F , G) Western blotting and qRT-PCR analysis of HIF-1α, Piezo1, cGAS and STING expression in HESCs Quantitative data are presented as mean ± SD. (H , I) Representative immunofluorescence images and quantification of the fibrosis marker α-SMA in HESCs (Scale bar: 100 μm). *P<0.05; **P<0.01; ***P<0.001; ****P<0.0001

Article Snippet: At 60-70% confluency, cells were treated for 24 h with the specified compounds: the Piezo1 agonist Yoda1 (5 μM; MCE, HY-18723); the Piezo1 inhibitor GsMTx4 (5 μM; MCE, HY-P1410); Ethidium bromide (0.2 μg/ml; Macklin, 1239-45-8) to deplete mitochondrial DNA; the calcium chelator BAPTA-AM (4 μM; MCE, HY-100168); the cGAS inhibitor RU.521(2.5μM, MCE, HY-114180 ); the STING inhibitor C-176 (2.5 μM; MCE, HY-112906); the anti-IL6 antibody (2.5 μg/ml, Sigma-Aldrich, I7901); or the ROS inhibitor NAC (2.0mM, MCE, HY-B0215).

Techniques: Activation Assay, Immunofluorescence, Control, Quantitative RT-PCR, Marker, Staining, Western Blot, Expressing

Journal: bioRxiv

Article Title: cGAS–STING induced IFN-β acts as a dual regulator of osteoclastogenesis via direct and osteoblast-mediated mechanisms

doi: 10.64898/2026.05.09.724040

Figure Lengend Snippet: (A) Effect of cGAS activation by G3-YSD complexed in LyoVec™ (YSD/LV; BMDMs: 250 ng/mL, RAW 264.7: 500 ng/mL) on RANKL-mediated osteoclast formation. Representative images of osteoclasts derived from BMDMs (left) and quantification of relative osteoclast numbers per well in BMDMs and RAW 264.7 cells (right). (B+C) Gene expression analysis of interferon-related genes (B) and osteoclast-associated genes (C) 48 h after stimulation with G3-YSD complexed in LyoVec™ (YSD/LV; BMDMs: 250 ng/mL, RAW 264.7: 500 ng/mL) in the presence or absence of 50 ng/mL RANKL. Data are normalized to the unstimulated control. (D–G) Effect of cGAS inhibition using RU.521 (10 µg/mL in DMSO) on osteoclast formation in RAW 264.7 cells. (D) Quantification of relative osteoclast numbers per well. (E) Gene expression analysis of interferon-related and osteoclast-associated genes 48 h after cGAS inhibition in the presence of 50 ng/mL RANKL. Data are normalized to the unstimulated control. (F) Time-dependent effects of cGAS inhibition, with inhibitor (RU.521, 10 µg/mL in DMSO) added throughout differentiation (“both”), during early stages (first 3 days) or during late stages (days 3–5/6). (G) Pre-inhibition of cGAS by treatment with RU.521 (10 µg/mL in DMSO) 24 h prior to RANKL stimulation. The inhibitor was removed before 50 ng/mL RANKL was added. Left: relative osteoclast numbers per well. Right: gene expression analysis of interferon- and macrophage-related genes and osteoclast-associated genes after 24 h cGAS inhibition followed by 48 h RANKL treatment. Data are normalized to the DMSO pre-treated RANKL control. (A-G) BMDMs were cultured in the presence of 25 ng/mL recombinant mouse M-CSF throughout all experiments. Osteoclast numbers per well are shown relatively to the RANKL control. Heatmaps display mean values, and bar graphs show mean ± SEM with individual data points. Statistical analysis was performed using one-way ANOVA with Bonferroni post hoc test (n = 3). RL: RANKL; LV: LyoVec™ transfection agent.

Article Snippet: Where indicated, cells were treated with: cGAS agonist G3-YSD (RAW 264.7: 500 ng/mL; BMDMs: 250 ng/mL) complexed with LyoVecTM (1:100, 15 min pre-incubation), cGAS inhibitor RU.521 (10 μg/mL in DMSO) added 3 h prior to stimulation; STING agonists 2′3′-cGAMP (RAW: 10 μg/mL; BMDMs: 5 μg/mL) or diABZI (0.01–10 μg/mL), STING inhibitor H-151 (40 or 400 ng/mL in DMSO) added 2 h prior to stimulation (all InvivoGen, USA).

Techniques: Activation Assay, Derivative Assay, Gene Expression, Control, Inhibition, Cell Culture, Recombinant, Transfection