Journal: Communications Biology

Article Title: Astragalin promotes HSCs ferroptosis through NCOA4 mediated ferritinophagy to alleviate liver fibrosis in zebrafish and mice

doi: 10.1038/s42003-025-08421-0

Figure Lengend Snippet: A Morphological changes of zebrafish larvae in 0–200 μM Ag at 24, 48, 72, and 96 hpf. B Survival rate of zebrafish larvae in 0–200 μM Ag from 24 to 96 hpf. C Hatching rate of zebrafish larvae in 0–200 μM Ag from 24 to 72 hpf. D Heat rate of zebrafish larvae in 0–200 μM Ag at 96 hpf. E Body length of zebrafish larvae in 0–200 μM Ag at 96 hpf. F H&E staining of liver in zebrafish larvae treated with 12.5, 25, and 50 μM Ag. Scale bar = 50 μm. G , H Immunohistochemical analysis of Desmin expression in zebrafish live and the quantitative analysis. Scale bar = 50 μm. I , J Immunofluorescence analysis of α-SMA expression in zebrafish and the quantitative analysis. Scale bar = 50 μm. K Fe 2+ probe staining of zebrafish liver. The quantitative analysis of staining was measured by imaged J software. All data presented as means ± SEM. n = 10 per group. (ns means p > 0.05, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001).

Article Snippet: Primary antibodies including α-SMA (1:400, proteintech), Desmin (HuaBio, 1:200), Collagen-1 (Abcam, 1:400), LC3B II (Abcam, 1:200), P62 (Abmart, 1:200), NCOA4 (DF4255, SAB, 1:100), GPX4 (ab125066, Abcam, 1:500), COX2 (ab283574, Abcam, 1:500), FTH (1:100, Abcam), ASCL4 (HY- P80977 , MCE, 1:100).

Techniques: Staining, Immunohistochemical staining, Expressing, Immunofluorescence, Software

Journal: Communications Biology

Article Title: Astragalin promotes HSCs ferroptosis through NCOA4 mediated ferritinophagy to alleviate liver fibrosis in zebrafish and mice

doi: 10.1038/s42003-025-08421-0

Figure Lengend Snippet: A , B Expression of α-SMA by immunofluorescence staining in HSC-T6 cells treated by Ag or/and CQ, and the quantitative analysis. C , D Expression of P62 by immunofluorescence staining in HSC-T6 cells treated by Ag or/and CQ, and the quantitative analysis. E – H Western blot analyses of LC3B II, NCOA4, and FTH protein in HSC-T6 cells, and the relative expression quantitative results. I , J Expression of NCOA4 by immunofluorescence staining in HSC-T6 cells treated by Ag or/and CQ, and the quantitative analysis. K , L Expression of α-SMA, LC3B II, and P62 by immunofluorescence staining in activated JS-1 cells treated by Ag or/and CQ, and the quantitative analysis. M Expression of α-SMA , LC3B II , and P62 mRNA in activated JS-1 cells. Scale bar = 10 μm. All data presented as means ± SEM. n = 3–6 per group. (ns means p > 0.05, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001).

Article Snippet: Primary antibodies including α-SMA (1:400, proteintech), Desmin (HuaBio, 1:200), Collagen-1 (Abcam, 1:400), LC3B II (Abcam, 1:200), P62 (Abmart, 1:200), NCOA4 (DF4255, SAB, 1:100), GPX4 (ab125066, Abcam, 1:500), COX2 (ab283574, Abcam, 1:500), FTH (1:100, Abcam), ASCL4 (HY- P80977 , MCE, 1:100).

Techniques: Expressing, Immunofluorescence, Staining, Western Blot

Journal: Communications Biology

Article Title: Astragalin promotes HSCs ferroptosis through NCOA4 mediated ferritinophagy to alleviate liver fibrosis in zebrafish and mice

doi: 10.1038/s42003-025-08421-0

Figure Lengend Snippet: A , B Western blot analyses of NCOA4 protein in HSC-T6 cells in NC or siNCOA4 group. C , D Expression of α-SMA, Desmin, and FTH by immunofluorescence staining in NC or siNCOA4 HSC-T6 cells treated with Ag, and the quantitative analysis. Scale bar = 10 or 50 μm. E HSC-T6 cells captured by TEM. The green arrows indicate the damaged mitochondria. The yellow arrows indicate the healthy mitochondria. The red arrows stand for autolysosome. F Quantitative PCR analyses of NCOA4 mRNA expression in activated JS-1 cells in NC or siNCOA4 group. G , H Western blot analyses of P62 and LC3B II protein in NC or siNCOA4 JS-1 cells treated with TGF-β1 or/and Ag, and the quantitative analysis. I and J Western blot analyses of ASCL4, XCT, and GPX4 protein in NC or siNCOA4 JS-1 cells treated with TGF-β1 or/and Ag, and the quantitative analysis. K JS-1 cells captured by TEM. L , M Expression of α-SMA by immunofluorescence staining in JS-1 cells in different groups, and the quantitative analysis. Scale bar = 20 μm. N Quantitative PCR analyses of FTH and FTL mRNA expression in JS-1 cells. All data presented as means ± SEM. n = 3–6 per group. (ns means p > 0.05, * p < 0.05, ** p < 0.01, ***p < 0.001, **** p < 0.0001).

Article Snippet: Primary antibodies including α-SMA (1:400, proteintech), Desmin (HuaBio, 1:200), Collagen-1 (Abcam, 1:400), LC3B II (Abcam, 1:200), P62 (Abmart, 1:200), NCOA4 (DF4255, SAB, 1:100), GPX4 (ab125066, Abcam, 1:500), COX2 (ab283574, Abcam, 1:500), FTH (1:100, Abcam), ASCL4 (HY- P80977 , MCE, 1:100).

Techniques: Western Blot, Expressing, Immunofluorescence, Staining, Real-time Polymerase Chain Reaction

Journal: Communications Biology

Article Title: Astragalin promotes HSCs ferroptosis through NCOA4 mediated ferritinophagy to alleviate liver fibrosis in zebrafish and mice

doi: 10.1038/s42003-025-08421-0

Figure Lengend Snippet: A , B Immunofluorescence staining of α-SMA, TFR, DMT1, and GPX4 in HSC-T6 cells treated with Ag as well as vehicle or Fer-1, and the quantitative analysis. Scale bar = 10 μm. C , D ROS probe staining and Fe 2+ probe staining of HSC-T6 cells in different groups, and the quantitative analysis. Scale bar= 10 or 100 μm. E MDA assay of HSC-T6 cells in different groups. F , G Immunofluorescence staining of α-SMA in JS-1 cells in different groups, and the quantitative analysis. Scale bar = 20 μm. H Quantitative PCR analyses of GPX4 , COX2 and TFR mRNA expression in JS-1 cells. I , J Western blot analyses of ASCL4 and DMT1 protein in JS-1 cells, and the quantitative analysis. K , L Western blot analyses of LC3B II, P62, FTH and FTL protein in JS-1 cells, and the quantitative analysis. All data presented as means ± SEM. n = 3–5 per group. (ns means p > 0.05, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001).

Article Snippet: Primary antibodies including α-SMA (1:400, proteintech), Desmin (HuaBio, 1:200), Collagen-1 (Abcam, 1:400), LC3B II (Abcam, 1:200), P62 (Abmart, 1:200), NCOA4 (DF4255, SAB, 1:100), GPX4 (ab125066, Abcam, 1:500), COX2 (ab283574, Abcam, 1:500), FTH (1:100, Abcam), ASCL4 (HY- P80977 , MCE, 1:100).

Techniques: Immunofluorescence, Staining, Multiple Displacement Amplification, Real-time Polymerase Chain Reaction, Expressing, Western Blot

Journal: Communications Biology

Article Title: Astragalin promotes HSCs ferroptosis through NCOA4 mediated ferritinophagy to alleviate liver fibrosis in zebrafish and mice

doi: 10.1038/s42003-025-08421-0

Figure Lengend Snippet: A The schematic diagram of experiments in mice. B Western blot analysis of Desmin and α-SMA proteins in different groups. C H&E, Masson, Sirius Red staining, and immumohistochemical staining of Collagen-1 in different groups. D , E The quantitative analysis of ( B ). F – H The quantitative analysis the quantitative analysis of ( C ). Scale bar = 100 μm. All data presented as means ± SEM. n = 3–5 per group. (ns means p > 0.05, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001).

Article Snippet: Primary antibodies including α-SMA (1:400, proteintech), Desmin (HuaBio, 1:200), Collagen-1 (Abcam, 1:400), LC3B II (Abcam, 1:200), P62 (Abmart, 1:200), NCOA4 (DF4255, SAB, 1:100), GPX4 (ab125066, Abcam, 1:500), COX2 (ab283574, Abcam, 1:500), FTH (1:100, Abcam), ASCL4 (HY- P80977 , MCE, 1:100).

Techniques: Western Blot, Staining

Journal: Communications Biology

Article Title: Astragalin promotes HSCs ferroptosis through NCOA4 mediated ferritinophagy to alleviate liver fibrosis in zebrafish and mice

doi: 10.1038/s42003-025-08421-0

Figure Lengend Snippet: A , B Western blot analysis of NCOA4 and FTH proteins in different groups and the quantitative analysis. C Quantitative PCR analyses of NCOA4 , FTL , FTH , COX2 , GPX4 , and TFR mRNA expression in HSCs sorted from the liver of mice in each group. D – F Immunofluorescence co-staining of COX2 and α-SMA of liver in different groups and the analysis of co-expression. Scale bar = 100 μm. G The quantitative analysis of COX2 and α-SMA. H Quantitative PCR analyses of COX2 , GPX4 , NCOA4 , FTL and FTH mRNA expression in hepatocytes sorted from the liver of mice in each group. I Quantitative PCR analyses of COX2 , GPX4 , NCOA4 , FTL and FTH mRNA expression in macrophages sorted from the liver of mice in each group. All data presented as means ± SEM. n = 3–5 per group. (ns means p > 0.05, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001).

Article Snippet: Primary antibodies including α-SMA (1:400, proteintech), Desmin (HuaBio, 1:200), Collagen-1 (Abcam, 1:400), LC3B II (Abcam, 1:200), P62 (Abmart, 1:200), NCOA4 (DF4255, SAB, 1:100), GPX4 (ab125066, Abcam, 1:500), COX2 (ab283574, Abcam, 1:500), FTH (1:100, Abcam), ASCL4 (HY- P80977 , MCE, 1:100).

Techniques: Western Blot, Real-time Polymerase Chain Reaction, Expressing, Immunofluorescence, Staining

Journal: Journal of Cachexia, Sarcopenia and Muscle

Article Title: The impact of senescence on muscle wasting in chronic kidney disease

doi: 10.1002/jcsm.13112

Figure Lengend Snippet: Cellular senescence appeared in the muscle of chronic kidney disease mice. A partial nephrectomy (5/6Nx) or sham operation (S) was performed in two‐month‐old C57BL6 mice; sham and CKD mice were harvested 2‐month later. (A) Blood urea nitrogen (BUN) serum and creatinine and (B) body weight and hindlimb muscle grip strength were measured. (C) Changes of muscle weight in gastrocnemius, tibialis anterior (TA), extensor digitorum longus (EDL) and soleus were evaluated in sham and CKD group. (D) the amount of p53, γ‐H2A.X, p21, and 16 INK4a were measured by immunoblot analysis in gastrocnemius muscle. The results are reported in the bar graph as the fold change of each protein, normalized to the GAPDH (mean ± SE; n = 6/cohort; * P < 0.05; ** P < 0.01; *** P < 0.001 vs. sham). (E) Muscles were stained with SA‐β‐gal (scale bar 50 μm), and the staining was quantified using ImageJ software. The results are reported in the bar/point graph as percentage of positive area (mean ± SE; n = 6/group; * P < 0.05; ** P < 0.01; *** P < 0.001 vs. sham). (F) The gastrocnemius muscle weight was plotted versus the quantified senescence markers γ‐H2A.X, p21, and p16 and the data were analysed by linear regression. Each point represents one animal ( n = 12).

Article Snippet: A FOXO1a plasmid was a gift from Domenico Accili (Addgene plasmid #17551; Watertown, MA) ; a p16 INK4a plasmid (SR403922) was from OriGene Technologies (Rockville, MD); p21 Waf1/Cip1 (sc‐29427) siRNA was from Santa Cruz, (Dallas, TX).

Techniques: Western Blot, Muscles, Staining, Software

Journal: Journal of Cachexia, Sarcopenia and Muscle

Article Title: The impact of senescence on muscle wasting in chronic kidney disease

doi: 10.1002/jcsm.13112

Figure Lengend Snippet: Limiting senescence with the senolytics ameliorated muscle wasting in CKD mice. CKD mice were fed a cocktail of dasatinib and quercetin (D&Q) two times per week and skeletal muscles were harvested 8 weeks later. (A) γ‐H2A.X + , Rb + , and SA‐β‐gal + were stained in gastrocnemius (scale bar 50 μm). The positive cells were counted using ImageJ software. The results are reported in the bar/point graphs as the percentage positive cells or area for sham (S), CKD (Nx) CKD plus vehicle (V), and CKD plus D&Q (DQ) ( n = 6/group; mean ± SE; * P < 0.05; ** P < 0.01; *** P < 0.001). (B) The protein level of senescence markers, γ‐H2A.X, p21, 16 INK4a , cyclin‐dependent kinases (CDK)1/2, and CDK4 were measured by immunoblot analysis. The graphs show the normalized fold changes of the indicated proteins compared with the levels of sham group (presented as one‐fold) ( n = 6/group; mean ± SE; * P < 0.05; ** P < 0.01; *** P < 0.001). (C) Muscle function was measured using a mouse grip strength meter with dual computerized sensors to detect and record the grip force in each group of mice. Data are presented as mean ± SE ( n = 6/group; KGF = kilogram force, *** P < 0.001). (D) Representative images of muscle cross‐sections immunostained with anti‐laminin antibody are presented. Cross‐sectional areas of 700–900 fibres per cohort were measured and the frequency distribution of fibre CSA are reported in the bar graph: Sham (blue), 5/6Nx (red), 5/6Nx + vehicle (green), and 5/6Nx + D&Q (purple) (mean ± SE; n = 6/group; scale bars = 50 μm). (E) The muscle atrophy markers atrogin‐1, MuRF1, and myostatin in the gastrocnemius muscle were measured by immunoblot analysis and normalized data are reported in arbitrary units in the bar/point graph ( n = 6/group; mean ± SE; * P < 0.05; ** P < 0.01; *** P < 0.001).

Article Snippet: A FOXO1a plasmid was a gift from Domenico Accili (Addgene plasmid #17551; Watertown, MA) ; a p16 INK4a plasmid (SR403922) was from OriGene Technologies (Rockville, MD); p21 Waf1/Cip1 (sc‐29427) siRNA was from Santa Cruz, (Dallas, TX).

Techniques: Muscles, Staining, Software, Western Blot

Journal: Journal of Cachexia, Sarcopenia and Muscle

Article Title: The impact of senescence on muscle wasting in chronic kidney disease

doi: 10.1002/jcsm.13112

Figure Lengend Snippet: Senolytics suppress uraemic serum‐induced senescence in MPCs. (A) MPCs were isolated from the skeletal muscle of normal C57BL6 mice and cultured in 2% control (CS) or uraemic serum (US) for 48 h and the amount of γ‐H2A.X, p21, 16 INK4a , and ki67 immunofluorescence was measured. DAPI staining was used to determine the number of nuclei (total cell population). The percentages of positive cells for each marker are reported in the bar/point graphs (scale bar = 50 μm). (B) MPCs were treated in 2% control or uraemic serum for 48 h. siRNAs corresponding to control (ctrlsi), p16 (p16si), or p21 (p21si) were transfected into the cells and cell proliferation was measured 48 h later. (C) MPCs were treated with 2% control or uraemic serum for 48 h, followed by transduction with ad‐ctrl, ad‐CDK1 or ad‐CDK4. After 48 h, cell proliferation was measured. Results are reported in the bar/point graph; data represent the absorbance in arbitrary units for each group. N = 6/group; bar: mean ± SE; * P < 0.05; ** P < 0.01; *** P < 0.001.

Article Snippet: A FOXO1a plasmid was a gift from Domenico Accili (Addgene plasmid #17551; Watertown, MA) ; a p16 INK4a plasmid (SR403922) was from OriGene Technologies (Rockville, MD); p21 Waf1/Cip1 (sc‐29427) siRNA was from Santa Cruz, (Dallas, TX).

Techniques: Isolation, Cell Culture, Control, Immunofluorescence, Staining, Marker, Transfection, Transduction

Journal: Journal of Cachexia, Sarcopenia and Muscle

Article Title: The impact of senescence on muscle wasting in chronic kidney disease

doi: 10.1002/jcsm.13112

Figure Lengend Snippet: Senolytics induced apoptosis in senescent MPCs. MPCs were cultured in 2% control or uraemic serum and treated with vehicle or D&Q for 48 h. (A) MPCs were evaluated for apoptosis by TUNEL and for p16 INK4a by immunostaining; DAPI staining was used to determine the number of nuclei. TUNEL and p16 INK4a positive cells are indicated by white arrows. The percentages of TUNEL positive cells that were also p16 INK4a positive cells were calculated and summarized in the bar/point graph (bars: mean ± SE; scale bar = 50 μm.; * P < 0.05; ** P < 0.01; *** P < 0.001). (B) MPCs were evaluated for apoptosis by TUNEL and for p21 by immunostaining; DAPI staining was used to determine the number of nuclei. Both TUNEL and p21 + cells were indicated by white arrows. The percentages of TUNEL + cells that were also p21 + cells were calculated and summarized in the bar/point graph (bars: mean ± SE; scale bar = 50 μm.; * P < 0.05; ** P < 0.01; *** P < 0.001).

Article Snippet: A FOXO1a plasmid was a gift from Domenico Accili (Addgene plasmid #17551; Watertown, MA) ; a p16 INK4a plasmid (SR403922) was from OriGene Technologies (Rockville, MD); p21 Waf1/Cip1 (sc‐29427) siRNA was from Santa Cruz, (Dallas, TX).

Techniques: Cell Culture, Control, TUNEL Assay, Immunostaining, Staining

Journal: Journal of Cachexia, Sarcopenia and Muscle

Article Title: The impact of senescence on muscle wasting in chronic kidney disease

doi: 10.1002/jcsm.13112

Figure Lengend Snippet: Mechanisms responsible for MPC senescence in CKD. CKD induces DNA damage in MPCs and activates the FOXO1 transcription factor. DNA damage increases p16 INK4a whereas the activation of FOXO1 transcription factor increases p21. Both p16 INK4a and p21 are cyclin‐dependent kinase (CDKs) inhibitors, which prevent cell cycle progression and lead to cell senescence. MPC senescence prevents muscle progenitor proliferation and induces the SASP in MPCs which leads to their production of inflammatory mediators and subsequent muscle wasting. Administration of dasatinib plus quercetin (D&Q) reduces CKD‐induced muscle atrophy by downregulation of FoxO1 and P16 INK4a as well as inducing apoptosis of senescent cells. This reduction of senescent cells limits inflammation and attenuates muscle wasting and dysfunction.

Article Snippet: A FOXO1a plasmid was a gift from Domenico Accili (Addgene plasmid #17551; Watertown, MA) ; a p16 INK4a plasmid (SR403922) was from OriGene Technologies (Rockville, MD); p21 Waf1/Cip1 (sc‐29427) siRNA was from Santa Cruz, (Dallas, TX).

Techniques: Activation Assay

Journal: Biochemistry and Biophysics Reports

Article Title: Biochemical characterization of a unique cytokinin and nucleotide phosphoribohydrolase Lonely Guy protein from Dictyostelium discoideum

doi: 10.1016/j.bbrep.2024.101756

Figure Lengend Snippet: Structural and conserved features of Dd Log. A) The predicted monomeric structure of Dd Log (Alphafold ID: AF-Q54UC5–F1) [ , ]. B) Proposed dimeric structure of Dd Log generated through SWISS-Model. The conserved PGG x GT xx E motif is highlighted in yellow in A and B, and both structures were visualized using Mol* (RCSB PDB) . C) MUSCLE alignments were performed using D. discoideum and 12 representative species with characterized or putative LOG orthologs ( Corynebacterium glutamicum , Mycobacterium tuberculosis , Bordetella pertussis , Sulfolobus islandicus , Chlorella variabilis , Marchantia polymorpha , Physcomitrium patens , Selaginella mollendorfii , Arabidopsis thaliana , Oryza sativa , Dictyostelium discoideum , Saccharomyces cerevisiae , and Claviceps purpurea ). The alignments were made and visualized with DNASTAR MegAlignPro and conserved functional residues across all 13 species are highlighted. The PGG x GT xx E motif is indicated by the dashed red box and red line. The red, teal, and purple triangles represent residues involved in catalysis, AMP binding, and prenyl-binding respectively . Dashes indicate gaps within the sequence alignment between orthologs. A full sequence alignment can be viewed in . D) Sequence logo for the multiple alignment. The figure was generated with DNASTAR MegAlignPro, and colors are representative of amino acid chemistry.

Article Snippet: Multiple sequence alignment was conducted using MUSCLE software [ ] in Lasergene MegAlignPro v. 17.4 (DNASTAR, Madison, Wisconsin, USA).

Techniques: Generated, Functional Assay, Binding Assay, Sequencing