Journal: iScience

Article Title: Accumulated mtDNA mutations are linked to specific impairments in NADH-linked respiration

doi: 10.1016/j.isci.2026.115184

Figure Lengend Snippet: PolG Mut mitochondria possess selective impairments to NAD-linked Respiration (A) Schematic depiction of the substrates and inhibitors added during the OxPhos kinetics assay. Mitochondrial oxygen consumption ( J O 2 ) across OxPhos kinetics assay in mitochondria isolated from (B) BAT, (C) brain, (D) colon, (E) heart, (F) kidney, (G) Liver, (H) and lung tissue. Ratio of maximal complex I (CI) versus CII-supported mitochondrial J O 2 (I). N = 4–6 per group. Data are presented as mean ± SEM and analyzed using multiple unpaired t tests ( B–5H) or unpaired t test ( I), ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. Substrates utilized are indicated as follows: creatine kinase (CK; 20 U/mL), ATP (5 mM), phospho-creatine (PCr; 1 mM), cytochrome c (Cyt C; 10 μM; pyruvate (Pyr; 5 mM), malate (Mal; 1 mM), octanoyl-carnitine (Oct; 0.2 mM), glutamate (Glut; 5 mM), rotenone (Rot; 0.5 μM), succinate (Succ; 5 mM) oligomycin (Oligo; 0.02 μM), malonate (Malo; 20 mM), calcium chloride (CaCl 2 ; 0.6 mM) glycerol-3-phosphate (G3P; 10 mM), and antimycin A (Ant A; 0.5 μM). Graphics were generated using BioRender.

Article Snippet: Following freeze fracture, 20μg of mitochondria were added to Respiration Buffer in the Oroboros O2k system followed by Cyt C (10μM).

Techniques: Isolation, Generated

Journal: iScience

Article Title: Accumulated mtDNA mutations are linked to specific impairments in NADH-linked respiration

doi: 10.1016/j.isci.2026.115184

Figure Lengend Snippet: Respiratory capacity within the electron transport system remains intact in PolG Mut mice (A–H) Schematic representing the maximal respiratory capacity protocol. Real-time oxygen consumption ( J O 2 ) in mitochondria isolated from (B) BAT, (C) brain, (D) colon, (E) heart, (F) kidney, (G) Liver, (H) and lung following serial titration of the mitochondrial uncoupling agent, carbonyl cyanide- p -trifluoromethoxyphenylhydrazone (FCCP, FC; 0.25 μM). (I) Maximal mitochondrial respiration achieved during FCCP titration. N = 5–6 per group, Data are presented as mean ± SEM. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗∗ p < 0.0001 depict significant post hoc LSD by two-way ANOVA; main effect PolG ( B–4H) or unpaired t test ( I). Substrates utilized are indicated as follows: pyruvate (Pyr; 5 mM), malate (Mal; 1 mM), octanoyl-carnitine (Oct; 0.2 mM), succinate (Succ; 5 mM) cytochrome c (Cyt C, 10 μM). Graphics were generated using BioRender.

Article Snippet: Following freeze fracture, 20μg of mitochondria were added to Respiration Buffer in the Oroboros O2k system followed by Cyt C (10μM).

Techniques: Isolation, Titration, Generated

Journal: iScience

Article Title: Accumulated mtDNA mutations are linked to specific impairments in NADH-linked respiration

doi: 10.1016/j.isci.2026.115184

Figure Lengend Snippet: Mitochondrial respiratory phenotypes are maintained in permeabilized tissue (A) Tissue slices were permeabilized in saponin before assessment of oxygen consumption ( J O 2 ). (B) Intact colon, (C) heart, and (D) liver J O 2 under multiple substrate conditions. (E) Ratio of complex I (CI) versus CII-supported respiration. (F) Dose-response to FCCP titration and (G) maximal respiratory capacity in intact bone marrow-derived mononuclear cells (BMMCs). (H) Mitochondrial J O 2 during OxPhos kinetics technique in permeabilized BMMCs. (I) Ratio of CI versus CII-supported respiration in permeabilized BMMCs. N = 5 per group. Data are presented as mean ± SEM, p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001,∗∗∗∗ p < 0.00001 depict significant multiple unpaired t tests ( A–6C and 6G), and unpaired t test ( D–6F and 6H). Substrates utilized are indicated as follows: creatine kinase (CK; 20 U/mL), ATP (5 mM), phospho-creatine (PCr; 1 mM), cytochrome c (Cyt C; 10 μM), pyruvate (Pyr; 5 mM), malate (Mal; 1 mM), octanoyl-carnitine (Oct; 0.2 mM), glutamate (Glut; 5 mM), rotenone (Rot; 0.5 μM), succinate (Succ; 5 mM) oligomycin (Oligo; 0.02 μM), malonate (Malo; 20 mM), calcium chloride (CaCl 2 ; 0.6 mM) glycerol-3-phosphate (G3P; 10 mM) antimycin A (Ant A; 0.5 μM), and carbonyl cyanide- p -trifluoromethoxyphenylhydrazone (FCCP, FC; 0.25 μM). Graphics were generated using BioRender.

Article Snippet: Following freeze fracture, 20μg of mitochondria were added to Respiration Buffer in the Oroboros O2k system followed by Cyt C (10μM).

Techniques: Titration, Derivative Assay, Generated

Journal: Redox Biology

Article Title: Unveiling a novel function of Aconitase-2: attenuating lung ischemia-reperfusion injury via inhibition of pulmonary endothelial apoptosis

doi: 10.1016/j.redox.2026.104016

Figure Lengend Snippet: ACO2 overexpression protected against LIRI in mice. (A) Schematic of AAV9-mediated ACO2-OE in I/R mice. (B–C) ACO2 expression levels assessed by RT-qPCR and western blot. (D) Blood gas parameters (PaO 2 and PaCO 2 ) following I/R injury. (E) Representative H&E-stained lung sections (scale bars = 20/50 μm, Black arrow: Alveolar expansion; Red arrow: Hemorrhage; Green arrow: Alveolar septum thickening and inflammatory cell infiltration). (F–G) Quantitative assessment of lung injury via histopathological scoring and wet/dry weight ratio. (H) Total cells count and protein concentration in BALF. (I) The mRNA expression of pro-inflammatory cytokines ( IL-6 , IL-1β , and TNF-α ). (J–K) Oxidative stress markers: T-AOC and MDA. (L) Apoptosis-related gene expression ( caspase 9 , Bcl-2, BAX , and Cyt-c ). (M) Mitochondrial respiratory function measured by OCR. (N) Serum Cyt-c release. (O) TUNEL staining for apoptosis detection (scale bar = 20 μm). Data are presented as the mean ± SD; n = 6; ∗ P < 0.05, ∗∗ P < 0.01, and ∗∗∗ P < 0.001.

Article Snippet: Contents of MDA (Cat. No: A003-1-2 ), MPO (Cat. No: A044-1-1), GSH (Cat. No: A006-2-1), T-AOC (Cat. No: A015-2-1), SOD (Cat. No: A001-3-2), and Cyt-c (Cat. No: H190-1-2) were quantified using commercial kits (Nanjing Jiancheng Bioengineering Institute) with a UV-VIS spectrophotometer.

Techniques: Over Expression, Expressing, Quantitative RT-PCR, Western Blot, Staining, Protein Concentration, Gene Expression, TUNEL Assay

Journal: Redox Biology

Article Title: Unveiling a novel function of Aconitase-2: attenuating lung ischemia-reperfusion injury via inhibition of pulmonary endothelial apoptosis

doi: 10.1016/j.redox.2026.104016

Figure Lengend Snippet: ACO2-KO exacerbated mitochondrial damage and apoptosis in HUVECs. (A–B) Validation of ACO2-KO efficiency by RT-qPCR and western blot. (C) Representative TEM images showing mitochondrial ultrastructural changes. The red arrows indicate the disappearance of mitochondrial cristae. (D) Cellular ATP contents measured by enzymatic assay. (E) The mtDNA copy number assessed via quantification of mitochondrial genes (ND-1, COX I, and COX IV). (F) Mitochondrial superoxide production detected by MitoSOX Red staining (scale bar = 20 μm). (G) Mitochondrial respiratory function measured by OCR. (H–I) Metabolomic analysis of TCA cycle intermediates: itaconate and isocitrate. (J) The mRNA expression of apoptosis-related genes ( Bcl-2, BAX, and Caspase-9 ). (K) Cyt-c release from mitochondria. (L) Apoptosis evaluation by TUNEL staining (scale bar = 20 μm). Data are presented as the mean ± SD; n = 3; ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001, and ∗∗∗∗ P < 0.0001.

Article Snippet: Contents of MDA (Cat. No: A003-1-2 ), MPO (Cat. No: A044-1-1), GSH (Cat. No: A006-2-1), T-AOC (Cat. No: A015-2-1), SOD (Cat. No: A001-3-2), and Cyt-c (Cat. No: H190-1-2) were quantified using commercial kits (Nanjing Jiancheng Bioengineering Institute) with a UV-VIS spectrophotometer.

Techniques: Biomarker Discovery, Quantitative RT-PCR, Western Blot, Enzymatic Assay, Staining, Metabolomic, Expressing, TUNEL Assay

Journal: Redox Biology

Article Title: Unveiling a novel function of Aconitase-2: attenuating lung ischemia-reperfusion injury via inhibition of pulmonary endothelial apoptosis

doi: 10.1016/j.redox.2026.104016

Figure Lengend Snippet: 4-OI rescued mitochondrial dysfunction after ACO2-KO in HUVECs. (A) Mitochondrial mass assessed by MitoTracker staining (scale bar = 20 μm). (B) Mitochondrial respiratory function measured by OCR. (C) Mitochondrial membrane potential evaluated using JC-1 staining (scale bar = 20 μm). (D) Activities of mitochondrial ETC complexes. (E) Protein contents of ETC complex subunits. (F–G) Metabolic ratios of NAD + /NADH and ATP/ADP. (H–I) The mRNA expression of apoptosis-related genes ( caspase-9 and BAX ). (J) Apoptosis-related protein contents (cleaved caspase-3, Bcl-2, and BAX) analyzed by western blot with semi-quantitative analysis. (K) Cyt-c release from mitochondria. (L) Apoptosis evaluation by TUNEL staining (scale bar = 20 μm). Data are presented as the mean ± SD; n = 3; ∗ P < 0.05, ∗∗ P < 0.01, and ∗∗∗ P < 0.001.

Article Snippet: Contents of MDA (Cat. No: A003-1-2 ), MPO (Cat. No: A044-1-1), GSH (Cat. No: A006-2-1), T-AOC (Cat. No: A015-2-1), SOD (Cat. No: A001-3-2), and Cyt-c (Cat. No: H190-1-2) were quantified using commercial kits (Nanjing Jiancheng Bioengineering Institute) with a UV-VIS spectrophotometer.

Techniques: Staining, Membrane, Expressing, Western Blot, TUNEL Assay