Journal: Bioactive Materials

Article Title: A multimodal ROS logic-gated therapeutic platform disrupts the vicious cycle of senescence to promote aged bone defect repair

doi: 10.1016/j.bioactmat.2026.02.002

Figure Lengend Snippet: Transcriptomic and molecular analysis of the potential pathways involved in MMBOx-mediated BMSCs rejuvenation. (A) Circular heatmap showing differentially expressed genes (DEGs) associated with cell senescence, inflammation, and osteogenesis in senescent BMSCs treated with MMBOx@GPP compared to GPP. (B) Gene Ontology (GO) enrichment analysis of upregulated DEGs. (C) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of upregulated DEGs. (D) Gene Set Enrichment Analysis (GSEA) plots of the glutathione metabolic process (ES: enrichment score; NES: normalized enrichment score; FDR: false discovery rate). (E) Heatmap of DEGs enriched in aging-related GO terms. (F) Western blot analysis of Keap1, Nrf2, Nqo1, Gclc, and GAPDH protein expression in BMSCs. (G) Quantitative analysis of protein band intensities ( n = 3). (H) Representative flow cytometry plots of ThiolTracker™ fluorescence staining indicating intracellular glutathione levels. (I) Quantification of intracellular GSH/GSSG ratio in BMSCs ( n = 3). (J) Schematic diagram illustrating the proposed mechanism by which MMBOx attenuates BMSCs senescence via Nrf2 pathway activation and glutathione metabolism enhancement. Data are expressed as mean ± SD; ∗ P < 0.05, ∗∗∗ P < 0.001.

Article Snippet: Western blotting was employed to evaluate protein expression of key targets, including Keap1 (Affinity, AF5266; 1:1000), Nrf2 (Proteintech, 16396-1-AP; 1:1000), Nqo1 (Abcam, ab80588; 1:10,000), Gclc (Proteintech, 12601-1-AP; 1:25000) and GAPDH (Proteintech, 60004-1-Ig; 1:50,000).

Techniques: Western Blot, Expressing, Flow Cytometry, Fluorescence, Staining, Activation Assay

Journal: bioRxiv

Article Title: Systematic Evaluation Defines the Limits of Ferroptosis in Cancer Therapy

doi: 10.64898/2026.03.11.711115

Figure Lengend Snippet: A , Left, correlation between the reported number of damaging mutations in each gene (DepMap) and nAUC measurement for BSO sensitivity (+Fer-1 condition, ) and associated P value, TXNRD1 highlighted. Right, correlation between number of number of damaging mutations in TXNRD1 in each cell line and corresponding nAUC for BSO sensitivity (+Fer-1), SKMES1 cell line highlighted. B , Immunoblot for indicated proteins from lysates of A549 cells and two clones in which TXNDR1 is deleted. MW markers indicated. C , Viability (ATP) of a TXNRD1 KO clones from B upon treatment with BSO, with or without βMe (50 μM), or Fer-1 (10 μM), 3 days. Data report mean ± SEM of n=3 biological replicates. D , Viability (ATP) of A549 cells, A549 TXNRD1 knockout clones, or SKMES1 cells expressing the indicated TXNRD1 cDNAs and selenocysteine (U498C) or SEICS (Δ3’UTR) mutants, treated with BSO as indicated, 3 days. Data report mean ± SEM of n=3 biological replicates. E , Schematic of glutathione synthesis and BSO sensitivity for GCLC versus GshF. F , Viability (ATP) of HT-1080, A549, A549 TXNRD1 knockout clones, or SKMES1 cells expressing S. thermophilus GshF, treated with BSO as indicated, 3 days. Data report mean ± SEM of n=3 biological replicates. G , Viability (ATP) of SKMES1 cells expressing exogenous GCLC cDNA and GCLC mutants resistant to BSO identified in , treated with BSO as indicated, 3 days. Data report mean ± SEM of n=3 biological replicates. H , Schematic of BSO (L-buthionine sulfoximine) and sulfoxide or methionine derivatives. Differences in buthionine vs methionine (blue circle) or sulfoximine vs sulfoxide (red circle) indicated. I , GSH content of HT-1080 cells upon treatment with BSO (red line) and indicated related species, 3 days. Data are mean of two technical replicates. J-K , Viability (ATP) upon treatment with BSO (red line) and indicated related species for A549 or A549 TXNRD1 knockout clones ( J ), or HT-1080 and SKMES1 cells, including additional Fer-1 treatment (10 μM, K ), 3 days. Data are mean of two technical replicates. L , Viability (ATP) of HT-1080, A549, A549 TXNRD1 knockout clones, or SKMES1 cells expressing the indicated GPX4 cDNAs and selenocysteine (U46C) or SEICS (Δ3’UTR) mutants, treated with BSO as indicated, 3 days. Data report mean ± SEM of n=3 biological replicates.

Article Snippet: TXNRD1 (SC-28321), GCLC (SC-166345) from Santa Cruz Goat Anti-mouse IgG (H+L), HRP (Invitrogen; 62-6520) Goat Anti-rabbit IgG (H+L), HRP (Invitrogen; 31460).

Techniques: Western Blot, Clone Assay, Knock-Out, Expressing

Journal: Scientific Reports

Article Title: Cycloastragenol attenuates osteoarthritis by restoring chondrocyte senescence via the NRF2/NF-κB signaling axis

doi: 10.1038/s41598-026-43064-z

Figure Lengend Snippet: CAG inhibiting NF-κB pathway through NRF2. ( A ) The structure model of CAG. ( B ) The ribbon model of the KEAP1. ( C ) The interaction between CAG and KEAP1; interaction energy is -9.1 kcal/mol. ( D ) The binding model between CAG and KEAP1, and CAG interacts with multiple residues within the KEAP1 Kelch domain. ( E , F ) Western blot analysis and quantification of NRF2 protein in chondrocytes under indicated treatments. ( G , H ) Western blot analysis and quantification of NRF2 downstream targets HO-1, NQO1, and GCLC proteins. ( I , J ) Representative immunofluorescence images and quantification of NRF2 protein localization, bar = 50 μm. n = 3. ( K , L ) Immunofluorescence staining and quantification of the effect of CAG on the mitochondrial membrane potential, bar = 50 μm, n = 3. ( M , N ) Effects of CAG on ROS levels in chondrocytes measured by flow cytometry. n = 3. ( O , P ) Western blot analysis and quantification of NRF2 protein expression in cells transfected with NRF2-targeting siRNAs (siNRF2-1, siNRF2-2, siNRF2-3). ( Q , R ) Immunofluorescence staining and quantification of the effect of si-NRF2 on the mitochondrial membrane potential, bar = 50 μm, n = 3. ( S , T ) Effects of si-NRF2 on ROS levels in chondrocytes measured by flow cytometry. n = 3. ( U , V ) Representative immunofluorescence images of p65 protein localization in chondrocytes under NRF2 inhibition treatment, bar = 50 μm. n = 3. ( W – Y ) Western blot analysis and quantification of proteins involving NF-κB pathway in chondrocytes under si-NRF2 transfection treatment (See Supplementary Materials for original blots). n = 3. * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001.

Article Snippet: Primary antibodies against COL2A1 (1:1000, YT1022, Immunoway), ADAMTS5 (1:1000, A2836, ABclonal), Aggrecan (1:500, A11691, ABclonal), MMP3 (1:1000, A1202, ABclonal), CDKN2A (1:1000, ab51243, Abcam), p53 (1:1000, ab32049, Abcam), CDKN1A (1:1000, ab109520, Abcam), NRF2 (1:1000, A3577, ABclonal), IKBα (1:1000, A19714, ABclonal), p-IKBα (1:1000, AP0707, ABclonal), p65 (1:1000, A19653 ABclonal), p-p65 (1:1000, AP1294, ABclonal), HO-1(1:5000, A28140PM, ABclonal), NQO1 (1:1000, A19586, ABclonal), GCLC (1:1000, A4499, ABclonal), Histone H3 (1:1000, A2348, ABclonal), and β-actin (1:1000, AC006, ABclonal) were used.

Techniques: Binding Assay, Western Blot, Immunofluorescence, Staining, Membrane, Flow Cytometry, Expressing, Transfection, Inhibition