Journal: International immunopharmacology

Article Title: Oxidative stress induces ferroptosis in tendon stem cells by regulating mitophagy through cGAS-STING pathway.

doi: 10.1016/j.intimp.2024.112652

Figure Lengend Snippet: Fig. 1. Ferroptosis is involved in the oxidative damage of TSCs induced by H2O2. (A-B) Through Western blot and RT-PCR, the levels of GPX4, SLC7A11 protein and mRNA were assessed with or without the ferroptosis inhibitor Fer-1 (10 μM). (C) Detection of mitochondrial ultrastructure by TEM. (D) TSC cells were stained with Prussian blue. The levels of (E) Iron, (F) GSH, (G) MDA in TSC cells was examined using the appropriate test kits. By using ELISA assays, the level of (H) 4-HNE and (I) LPO were determined. (J) The ROS intensity in TSC cells was evaluated using ROS probe. (K) Western blot detected the protein levels of tenomodulin (n = 3); ***p < 0.001 and ###p < 0.001. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: To evaluate intracellular 4-HNE, the ELISA kit (CUSA BIO; # CSBEQ027232RA) was utilized.

Techniques: Western Blot, Reverse Transcription Polymerase Chain Reaction, Staining, Enzyme-linked Immunosorbent Assay

Journal: International immunopharmacology

Article Title: Oxidative stress induces ferroptosis in tendon stem cells by regulating mitophagy through cGAS-STING pathway.

doi: 10.1016/j.intimp.2024.112652

Figure Lengend Snippet: Fig. 3. Inhibition of mitophagy attenuates ferroptosis induced by H2O2 in TSCs. TSCs cells were pretreated with Mdivi-1 (1 μM) for 2 h and then subjected to the prescribed concentration of H2O2 (100 μM) for a period of 24 h. (A) MTT experiment were used to test cell viability. (B) Using the relevant kits, the levels of GSH in TSC cells were measured. (C–D) To measure the level of intracellular iron, Prussian blue staining and an Iron Colorimetric Assay Kit were utilized. (E) MDA levels in TSC cells were determined using the relevant kits. (F-G) To measure the amount of 4-HNE and LPO in TSC cells, ELISA assays were employed. (H) The ROS con centration in TSC cells was measured using a ROS probe. (I) Western blot detected the protein levels of Tenomodulin (n = 3); **p < 0.01, ***p < 0.001, #p < 0.05, ##p < 0.01, and ###p < 0.001. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: To evaluate intracellular 4-HNE, the ELISA kit (CUSA BIO; # CSBEQ027232RA) was utilized.

Techniques: Inhibition, Concentration Assay, Staining, Colorimetric Assay, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: International immunopharmacology

Article Title: Oxidative stress induces ferroptosis in tendon stem cells by regulating mitophagy through cGAS-STING pathway.

doi: 10.1016/j.intimp.2024.112652

Figure Lengend Snippet: Fig. 6. Interfering with cGAS attenuates ferroptosis induced by H2O2 in TSCs. TSC cells were treated with H2O2 for 24 h after receiving cGAS siRNA transfection for 12 h. (A) MTT experiment were used to test cell viability. (B) By using appropriate kits, the levels of GSH in TSC cells were determined. (C-D) To measure the level of intracellular iron, Prussian blue staining and an Iron Colorimetric Assay Kit were utilized. (E) To measure the amount of ROS present in TSC cells, a ROS probe was employed. (F) MDA levels in TSC cells were determined using the relevant kits. (G-H) To measure the amount of 4-HNE and LPO in TSC cells, ELISA assays were employed. (I) Western blot detected the protein levels of Tenomodulin. (n = 3); *p < 0.05, **p < 0.01, ***p < 0.001, #p < 0.05, ##p < 0.01, ###p < 0.001, ####p < 0.0001. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: To evaluate intracellular 4-HNE, the ELISA kit (CUSA BIO; # CSBEQ027232RA) was utilized.

Techniques: Transfection, Staining, Colorimetric Assay, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: PloS one

Article Title: Increased oxidative stress impairs adipose tissue function in sphingomyelin synthase 1 null mice.

doi: 10.1371/journal.pone.0061380

Figure Lengend Snippet: Figure 3. SMS1-KO WAT is severely damaged by oxidative stress. (A) Proteins were extracted from epiWAT and liver of 10-week-old wild- type (WT, n = 3) or SMS1-KO (KO, n = 3) mice and carbonylated proteins were detected by immunoblot analysis using anti-DNP antibody. (B) Samples in (A) were subjected to immunoblot analysis using anti-4-HNE antibody to detect protein modification by ROS. Hsc70 staining served as a standard. (C) Carbonylated proteins in epiWAT of 20-week-old mice were immunohistochemically detected using an anti-DNP antibody (Left panel), and DNP signal intensity in the plasma membrane area was quantified (WT, n = 3; KO, n = 3) (Right panel). (D) mRNAs were extracted from WAT of 10-week-old mice. mRNA expression levels of genes encoding apoptotic factors were assessed by quantitative RT-PCR and normalized to b–actin expression. The wild-type group average was set to 1. n = 6–9 samples per group. (E) Activity of caspase-3 was spectrophotometrically assessed. n = 6 samples per

Article Snippet: Immunoblotting was performed with antiHsc70 antibody (Santa Cruz Biotechnology, Santa Cruz, California, USA) or anti-4-hydroxy-2-nonenal (4-HNE) antibody (R&D Systems, Minneapolis, Minnesota, USA).

Techniques: Western Blot, Modification, Staining, Clinical Proteomics, Membrane, Expressing, Quantitative RT-PCR, Activity Assay