calcimycin  (MedChemExpress)

Journal: Advanced Science

Article Title: Succinate Dehydrogenase Subunit A (SDHA) Mediated Microglia Extracellular Traps Formation Participating in Cerebral Ischemic Reperfusion Injury

doi: 10.1002/advs.202411873

Figure Lengend Snippet: OGD/R induced MiETosis in vitro. A) Cell viability of micoglia was detected by CCK‐8 assay after OGD (4 h) following re‐oxygenation for 6, 12, 24 h, and B) incubation with A23187 (5 µM) for 1, 2, 4, 6 h. C) Representative images of microglia following incubation with the Hoechst 33342 show the presence of nuclear decondensation at 40 × magnification. Scale bar = 10 µm. D) Quantification of percentage of microglia undergoing nuclear decondensation. E) Representative images of Iba1, citH3 and MPO triple‐stained microglia, and colocalization analysis of MPO, citH3 and DNA in three group. Scale bar = 50 µm F,G) Quantitative analysis of MPO and citH3 mean immunofluorescence intensities (MFI) in each group of microglia. H) MPO, I) ds‐DNA) concentrations in microglia culture supernatant, as detected by ELISA. J) Scanning electron microscopy (SEM) results demonstrated that microglia released porous, protein‐rich extracellular structures following OGD/R treatment. K,L) Representative immunoblot and statistical analysis of citH3, MPO, MMP9, PAD2, PAD4 and β‐actin after OGD/R exposure. M) Representative images of microglia following incubation with the fluorescent probe mitoSOX show the production of mtROS at 40 × magnification. Scale bar = 10 µm. N) Single‐color histogram of mitoSOX fluorescence intensity. All images are representative and were chosen at random. O) Quantification of mean intensities of mitoSOX. All experiments were performed at least three independent times. All data are presented as mean ± SD, * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: A23187 (10 mM, 52665‐69‐7; MedChemExpress) stock solutions were dissolved in DMSO at a storage concentration.

Techniques: In Vitro, CCK-8 Assay, Incubation, Staining, Immunofluorescence, Enzyme-linked Immunosorbent Assay, Electron Microscopy, Western Blot, Fluorescence

Journal: Advanced Science

Article Title: Succinate Dehydrogenase Subunit A (SDHA) Mediated Microglia Extracellular Traps Formation Participating in Cerebral Ischemic Reperfusion Injury

doi: 10.1002/advs.202411873

Figure Lengend Snippet: Inhibition of MiETs formation protected neuron from injury in vitro. A,E) Schematic illustration of the experimental design framework for this section. B) Cell viability of neurons was detected by CCK‐8 assay after incubation with supernatant from microglia in Con, A23187 and OGD/R groups. C) PI‐Hoechst double‐stained neurons treated with four types of microglia culture medium. D) Quantification of PI/Hoechst double‐positive cells. Scale bar = 50 µm. F) Cell viability of neurons was detected by CCK‐8 assay after incubation with supernatant from microglia in Con, OGD/R, BB‐Cl, and DMM groups. G) PI‐Hoechst double‐stained neurons treated with five types of microglia culture medium. H) Quantification of PI/Hoechst double‐positive cells in these five groups. Scale bar = 50 µm. All experiments were performed at least three independent times. All data are presented as mean ± SD, * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: A23187 (10 mM, 52665‐69‐7; MedChemExpress) stock solutions were dissolved in DMSO at a storage concentration.

Techniques: Inhibition, In Vitro, CCK-8 Assay, Incubation, Staining