Journal: Journal of Cellular and Molecular Medicine

Article Title: Arachidonic Acid Disrupts PKM2 ‐Linked Glycolysis‐Related Metabolism and Exacerbates PAN ‐Induced Podocyte Injury

doi: 10.1111/jcmm.71404

Figure Lengend Snippet: Aa aggravated PAN‐induced podocyte injury in vitro. Control podocytes were treated with PBS. PAN‐treated podocytes received 250 μM PAN for 24 h. PAN+AA‐treated podocytes received PAN (250 μM) and AA (5 × 10 −7 M) for 24 h. AA‐treated podocytes received AA (5 × 10 −7 M) for 24 h. (A) Representative phalloidin staining images of F‐Actin (green) and nuclei (blue), with quantification of F‐Actin mean intensity per cell ( n = 3). Scale bar, 20 μm. (B) Western blot analysis and quantification of nephrin protein expression in cultured podocytes ( n = 6). Data are expressed as the mean ± SD and analysed by one‐way ANOVA with Bonferroni correction. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001; ns, not significant.

Article Snippet: Membranes were incubated overnight at 4°C with primary antibodies against β‐actin (1:2000, Sigma‐Aldrich, MO, USA), nephrin (1:500, PROGEN), PKM1 (1:1000, Abcam), PKM2 (1:1000, Proteintech), phospho‐PKM2 (Tyr105 1:1000, Proteintech), β‐actin (1:1000, CST), and TOMM20 (1:1000, CST).

Techniques: In Vitro, Control, Staining, Western Blot, Analysis, Expressing, Cell Culture

Journal: Journal of Cellular and Molecular Medicine

Article Title: Arachidonic Acid Disrupts PKM2 ‐Linked Glycolysis‐Related Metabolism and Exacerbates PAN ‐Induced Podocyte Injury

doi: 10.1111/jcmm.71404

Figure Lengend Snippet: PKM2 Activation attenuated PAN + AA‐induced podocyte injury and mitochondrial dysfunction in vitro. PAN+AA‐treated podocytes were exposed to PAN (250 μM) and AA (5 × 10 −7 M) for 24 h. PAN+AA+TEPP‐46‐treated podocytes were exposed to TEPP‐46 (10 μM) under PAN+AA conditions. T denotes TEPP‐46 in the figure. (A) Representative phalloidin staining images of F‐Actin and quantification of F‐Actin mean intensity per cell ( n = 3). Scale bar, 20 μm. (B) Representative TUNEL staining images and quantification of apoptotic podocytes ( n = 3). Scale bar, 50 μm. (C, D) Representative immunofluorescence images and quantification of PKM2 and phospho‐PKM2. Nephrin is shown in red, PKM2 or phospho‐PKM2 in green, and DAPI in blue ( n = 3). Scale bar, 20 μm. (E) Western blot analysis and quantification of PKM2 protein expression ( n = 6). (F) Western blot analysis and quantification of phospho‐PKM2 (p‐PKM2) protein expression ( n = 6). (G) Western blot analysis and quantification of TOM20 protein expression ( n = 6). (H) Representative immunofluorescence images and quantification of TOM20 ( n = 3). Scale bar, 20 μm. (I) Transmission electron microscopy images and quantification of mean mitochondrial area, cristae‐damage rate and mitochondrial damage score ( n = 3). Scale bar, 2.0 μm. (J) MitoTracker staining and quantification of mitochondrial fluorescence intensity ( n = 3). Scale bar, 20 μm. (K) JC‐1 staining and quantification of mitochondrial membrane potential ( n = 3). Scale bar, 20 μm. (L) Representative DCFH‐DA staining images and quantification of ROS levels ( n = 3). Scale bar, 20 μm. (M) PK activity, lactate production, pyruvate production and ATP levels ( n = 3). Data are expressed as mean ± SD. Statistical significance was determined by unpaired Student's t ‐test. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: Membranes were incubated overnight at 4°C with primary antibodies against β‐actin (1:2000, Sigma‐Aldrich, MO, USA), nephrin (1:500, PROGEN), PKM1 (1:1000, Abcam), PKM2 (1:1000, Proteintech), phospho‐PKM2 (Tyr105 1:1000, Proteintech), β‐actin (1:1000, CST), and TOMM20 (1:1000, CST).

Techniques: Activation Assay, In Vitro, Staining, TUNEL Assay, Immunofluorescence, Western Blot, Analysis, Expressing, Transmission Assay, Electron Microscopy, Fluorescence, Membrane, Activity Assay

Journal: Journal of Cellular and Molecular Medicine

Article Title: Arachidonic Acid Disrupts PKM2 ‐Linked Glycolysis‐Related Metabolism and Exacerbates PAN ‐Induced Podocyte Injury

doi: 10.1111/jcmm.71404

Figure Lengend Snippet: Proposed working model of AA‐associated aggravation of PAN‐induced podocyte injury. Based on our previous study, AA may activate the PKA‐c‐Abl‐nephrin axis and contribute to nephrin phosphorylation and actin cytoskeletal remodelling. In the present study, PAN+AA was associated with reduced PKM2 abundance, increased PKM2 Tyr105 phosphorylation, decreased PK activity, lower pyruvate/lactate/ATP levels, mitochondrial injury, ROS accumulation and apoptosis. These cytoskeletal and metabolic stress pathways may converge to aggravate podocyte injury and proteinuria. Potential crosstalk between these pathways is proposed, but the sequence and causal relationship remain to be experimentally validated.

Article Snippet: Membranes were incubated overnight at 4°C with primary antibodies against β‐actin (1:2000, Sigma‐Aldrich, MO, USA), nephrin (1:500, PROGEN), PKM1 (1:1000, Abcam), PKM2 (1:1000, Proteintech), phospho‐PKM2 (Tyr105 1:1000, Proteintech), β‐actin (1:1000, CST), and TOMM20 (1:1000, CST).

Techniques: Phospho-proteomics, Cytoskeletal, Activity Assay, Sequencing

Journal: Science Advances

Article Title: Canonical Wnt induction by OTULIN prevents keratinocyte death and skin inflammation

doi: 10.1126/sciadv.adz0181

Figure Lengend Snippet: List of antibodies.

Article Snippet: 1/10000 , Santa Cruz Biotechnology Biotechnology , sc-47778 , Western blotting.

Techniques: Concentration Assay, Cell-Signaling, Immunohistochemistry, Immunofluorescence, Western Blot

Journal: Neurochemical Research

Article Title: Recurrent Sleep Restriction During Aging Induces Early Recognition Memory Impairment, Neuroinflammation, and Blood–Brain Barrier Dysfunction in Female Wistar Rats

doi: 10.1007/s11064-026-04909-1

Figure Lengend Snippet: Recurrent sleep restriction decreases levels of the ZO-1 protein in the cerebral cortex during aging. A Representative blots of each protein in the cortex using β-actin as a load control. B Normalized levels expressed as relative fold change of the ZO-1 and C occludin proteins. D Representative blots of each protein in the hippocampus using β-actin as a load control. E Normalized levels expressed as relative fold change of the ZO-1 and F occludin proteins. Data are presented as mean ± SEM. Two-way ANOVA, followed by Tukey’s test for multiple comparisons ( n = 4 per group, in duplicate). * p < 0.05, ** p < 0.001, *** p < 0.0001

Article Snippet: Membranes were then incubated overnight at 4 °C with the following primary antibodies: ZO-1 (Invitrogen, Cat. No. 40–2200, 1:500), occludin (Invitrogen, Cat. No. 40–4700, 1:500), and β-actin (Santa Cruz Biotechnology, Cat. No. sc-69879, 1:2500).

Techniques: Control