Journal: Nature Communications

Article Title: ARMH4 accelerates aging by maintaining a positive-feedback growth signaling circuit

doi: 10.1038/s41467-025-67505-x

Figure Lengend Snippet: a Schematic diagram of IGF1R and FGFR1 activation pathways. b , c Co-IP ( b ) and immunofluorescence ( c ) to detect the interaction between ARMH4 and IGF1R/FGFR1 in HEK293 cells co-transfected with pcDNA3.1-h Armh4 -3FLAG and pcDNA3.1-h Igf1r -HA, as well as pcDNA3.1-h Armh4 -3FLAG and pcDNA3.1-h Fgfr1 -HA, Scale bar, 10 μm. All experiments were performed twice independently with comparable results, and representative data from one experiment are shown. d , e Validation of ARMH4-IGF1R ( d ) and ARMH4-FGFR1 ( e ) interaction using endogenous IP in heart tissues. All experiments were performed twice independently with comparable results, and representative data from one experiment are shown. f AlphaFold 3 simulation of the effect of ARMH4 on ligand-receptor binding for IGF1/IGF1R and FGF2/FGFR1. g AlphaFold3 simulation of the specific binding region of ARMH4 within the IGF1/IGF1R ligand-receptor complex. h Schematic diagram of the designed plasmid construct targeting the predicted binding sequence between ARMH4 and IGF1R. i Co-IP analysis of the interaction between IGF1R and ARMH4, including its isoforms, in HEK293 cells. All experiments were performed twice independently with comparable results, and representative data from one experiment are shown. Source data are provided as a Source Data file.

Article Snippet: The pcDNA3.1-h Igf1r -HA and pcDNA3.1-h Fgfr1 -HA recombinant plasmids were constructed by cloning the coding region of h Igf1r and h Fgfr1 into the NheI and XbaI (NEB, #R0145) sites of the pcDNA3.1-HA plasmid. pLV3-CMV- Myc (mouse)-3FLAG-Fluc-Puro was purchased from MIAOLING BIOLOGY (#P58001).

Techniques: Activation Assay, Co-Immunoprecipitation Assay, Immunofluorescence, Transfection, Biomarker Discovery, Binding Assay, Plasmid Preparation, Construct, Sequencing

Journal: Nature Communications

Article Title: ARMH4 accelerates aging by maintaining a positive-feedback growth signaling circuit

doi: 10.1038/s41467-025-67505-x

Figure Lengend Snippet: a , b Adenoviral overexpression of Armh4 with PPP ( a ) and FA ( b ) treatment in NRVMs followed by Western blot analysis of phosphorylated mTOR, p70S6K, Akt and ERK. Representative blot of n = 3 biological replicates. Data are presented as mean values +/- SEM. Black circles represent individual data points for n = 3 biological replicates. c , d Adenoviral overexpression of Armh4 with PPP ( c ) and FA ( d ) treatment in NRVMs followed by western blot to detect the autophagy-related markers. Representative blot of n = 3 biological replicates. Data are presented as mean values +/- SEM. Black circles represent individual data points for n = 3 biological replicates. e , f Adenoviral overexpression of Armh4 with siRNA-mediated knockdown of Igf1r ( e ) and Fgfr1 ( f ) in NRVMs followed by western blot analysis of phosphorylated p70S6K, Akt, ERK and LC3. Representative blot of n = 3 biological replicates. Data are presented as mean values +/- SEM. Black circles represent individual data points for n = 3 biological replicates. g , h Adenoviral overexpression of Armh4 with IGF1 ( g ) and FGF2 ( h ) treatment in NRVMs followed by western blot analysis of phosphorylated p70S6K, Akt, ERK and LC3. n = 3. Representative blot of n = 3 biological replicates. Data are presented as mean values +/- SEM. Black circles represent individual data points for n = 3 biological replicates. Statistical significance was determined by two-way ANOVA, and the exact P values are reported directly on the figures. Source data are provided as a Source Data file.

Article Snippet: The pcDNA3.1-h Igf1r -HA and pcDNA3.1-h Fgfr1 -HA recombinant plasmids were constructed by cloning the coding region of h Igf1r and h Fgfr1 into the NheI and XbaI (NEB, #R0145) sites of the pcDNA3.1-HA plasmid. pLV3-CMV- Myc (mouse)-3FLAG-Fluc-Puro was purchased from MIAOLING BIOLOGY (#P58001).

Techniques: Over Expression, Western Blot, Knockdown

Journal: Nature Communications

Article Title: ARMH4 accelerates aging by maintaining a positive-feedback growth signaling circuit

doi: 10.1038/s41467-025-67505-x

Figure Lengend Snippet: a Western blot analysis of FGFR1, IGFR1 protein expression in the hearts of young and aged WT and KO mice. Representative blot of n = 3 independent mice. Data are presented as mean values +/- SEM. Black circles represent individual data points for n = 3 independent mice. Statistical significance was determined by two-way ANOVA. b Western blot analysis of FGFR1, IGFR1 protein expression in the livers of young and aged WT and KO mice. Representative blot of n = 3 independent mice. Data are presented as mean values +/- SEM. Black circles represent individual data points for n = 3 independent mice. Statistical significance was determined by two-way ANOVA. c Western blot analysis of FGFR1, IGFR1 protein expression in Armh4 -deficient MEFs. Representative blot of n = 3 biological replicates. Data are presented as mean values +/- SEM. Black circles represent individual data points for n = 3 biological replicates. Statistical significance was determined by a two-tailed unpaired t -test. d, e Western blot analysis of FGFR1 and IGFR1 protein expression in NRVMs with/without siRNA-mediated knockdown of Armh4 ( d ) and adenoviral overexpression of Armh4 ( e ). Representative blot of n = 3 biological replicates. Data are presented as mean values +/- SEM. Black circles represent individual data points for n = 3 biological replicates. Statistical significance was determined by a two-tailed unpaired t -test. f Western blot analysis of FGFR1 and IGFR1 protein expression in NRVMs treated with recombinant ARMH4. n = 6. Representative blot of n = 6 biological replicates. Data are presented as mean values +/- SEM. Black circles represent individual data points for n = 6 biological replicates. Statistical significance was determined by a two-tailed unpaired t -test. The exact P values are reported directly on the figures. Source data are provided as a Source Data file.

Article Snippet: The pcDNA3.1-h Igf1r -HA and pcDNA3.1-h Fgfr1 -HA recombinant plasmids were constructed by cloning the coding region of h Igf1r and h Fgfr1 into the NheI and XbaI (NEB, #R0145) sites of the pcDNA3.1-HA plasmid. pLV3-CMV- Myc (mouse)-3FLAG-Fluc-Puro was purchased from MIAOLING BIOLOGY (#P58001).

Techniques: Western Blot, Expressing, Two Tailed Test, Knockdown, Over Expression, Recombinant

Journal: Nature Communications

Article Title: ARMH4 accelerates aging by maintaining a positive-feedback growth signaling circuit

doi: 10.1038/s41467-025-67505-x

Figure Lengend Snippet: a TF-Target Finder predicted the transcription factors that co-regulated by IGF1R and FGFR1 screening diagram. b Western blot analysis of c-Myc protein expression in NRVMs upon Armh4 knockdown and overexpression. Representative blot of n = 6 biological replicates. Data are presented as mean values +/- SEM. Black circles represent individual data points for n = 6 biological replicates. Statistical significance was determined by a two-tailed unpaired t -test. c, d qRT-PCR of Myc expression in NRVMs upon Armh4 knockdown ( c ) and overexpression ( d ). Data are presented as mean values +/- SEM. Black circles represent individual data points for n = 3 biological replicates. Statistical significance was determined by a two-tailed unpaired t -test. e Schematic diagram of the dual-luciferase reporter constructs. f Dual-luciferase reporter assay in HEK293 cells. Data are presented as mean values +/- SEM. Black circles represent individual data points for n = 3 biological replicates. Statistical significance was determined by a two-tailed unpaired t -test. g Screening the c-Myc binding motifs by JASPAR. h Computational prediction of c-Myc binding motifs in IGF1R/FGFR1 regulatory regions using JASPAR. i Western blot analysis of c-Myc, IGF1R, FGFR1 expression in NRVMs upon Myc knockdown by siRNA. Representative blot of n = 6 biological replicates. Data are presented as mean values +/- SEM. Black circles represent individual data points for n = 6 biological replicates. Statistical significance was determined by a two-tailed unpaired t -test. j–m Western blot analysis of IGF1R and FGFR1 expression ( j ), phosphorylated p70S6K, Akt, ERK ( k ), puromycin incorporation ( l ), the autophagy-related marker ( m ) in NRVMs with Myc knockdown followed by Armh4 overexpression. Representative blot of n = 4 biological replicates ( j , k , m ) and n = 3 biological replicates ( l ). Data are presented as mean values +/- SEM. Black circles represent individual data points for n = 4 biological replicates ( j , k , m ) and n = 3 biological replicates ( l ). Statistical significance was determined by two-way ANOVA. The exact P values are reported directly on the figures. Source data are provided as a Source Data file.

Article Snippet: The pcDNA3.1-h Igf1r -HA and pcDNA3.1-h Fgfr1 -HA recombinant plasmids were constructed by cloning the coding region of h Igf1r and h Fgfr1 into the NheI and XbaI (NEB, #R0145) sites of the pcDNA3.1-HA plasmid. pLV3-CMV- Myc (mouse)-3FLAG-Fluc-Puro was purchased from MIAOLING BIOLOGY (#P58001).

Techniques: Western Blot, Expressing, Knockdown, Over Expression, Two Tailed Test, Quantitative RT-PCR, Luciferase, Construct, Reporter Assay, Binding Assay, Marker

Journal: British Journal of Cancer

Article Title: Fibroblast growth factor signals drive the metastatic behavior in small cell lung cancer

doi: 10.1038/s41416-025-03276-y

Figure Lengend Snippet: a Pooled data of sprouting inhibition compared to DMSO controls after 120 h of sprouter cell lines (HLHE, H196, H372, H1341) after treatment with FGFR inhibitor erdafitinib, EGFR inhibitor erlotinib, TGFβR inhibitor galunisertib and SB-431542, EZH2 inhibitor GSK2816126A (GSK126), VEGFR1-3 inhibitor KRN-633, YAP1/TAZ and TEAD inhibitor K-975, VEGFR1-3, FGFR1-3, PDGFR inhibitor nintedanib, PDGFRα, β inhibitor seralutinib, MST1/2 inhibitor XMU-MP-1 and PORCN inhibitor Wnt-C59. Each dot represents one cell line. Statistical evaluation was performed using one-way ANOVA and DUNN´s multiple comparison test. * p < 0.05, ** p < 0.01, *** p < 0.001 b Representative images of sprouting ability of HLHE cells with control (DMSO, upper panel) and nintedanib (0.5 µM, lower panel) treatment after 24 h, 96 h, and 120 h. Scale bar: 500 µm. c Quantification of sprouting based on spheroid sprouting assays with reduced serum (2.5% FBS). Spheroids were treated with 10 ng/ml and 50 ng/ml recombinant FGF2 for 96 h. Mean sprout length over time is shown as mean ± SEM of at least 10 individual spheroids. Two-way ANOVA and Tukey´s multiple comparison test. *** p < 0.001. d RNA expression of FGFRs and FGFs in pooled non-sprouter (ochre) and non-sprouter (blue) cell lines ( n = 4), determined by qPCR. Data is shown as mean ± SEM. Mann-Whitney test. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: TaqMan assay IDs used in this study comprised FGF-1, Hs01106908_m1, FGF-2, Hs00266645_m1, FGF-5, Hs00170454_m1, FGF-18, Hs00826077_m1, FGFR1, Hs00915142_m1, FGFR2, Hs01552918_m1, FGFR3, Hs00179829_m1, FGFR4 and Hs01106908_m1.

Techniques: Inhibition, Comparison, Control, Recombinant, RNA Expression, MANN-WHITNEY

Journal: Nature Communications

Article Title: FGF4-FGFR1 signaling promotes podocyte survival and glomerular function to ameliorate diabetic kidney disease in male mice

doi: 10.1038/s41467-025-65978-4

Figure Lengend Snippet: a Relative mRNA levels of Fgfr1-4 in total renal tissue lysates from C57BL/6 mice ( n = 16). b , c Immunofluorescence images depicting FGFR1 distribution (green) in normal renal tissues from ( b ) C57BL/6 mice and ( c ) human samples. White dashed lines delineate glomerular boundaries. d , e High-magnification images showing co-localization of FGFR1 (green) with the podocyte marker podocin (red) in ( d ) mouse and ( e ) human glomeruli. f Schematic of podocyte-specific Fgfr1 knockout ( Fgfr1 -PKO), and Western blot of isolated glomeruli confirms loss of FGFR1 in Fgfr1 -PKO versus littermate controls. g , h Changes in ( g ) blood glucose levels and ( h ) BUN and UACR across treatment groups ( n = 6). i Renal excretion kinetics, GFR, and elimination half-life ( t 1/2 ) in mice from each group ( n = 3). j , k Histological examination and quantification of renal sections using H&E, PAS, and Masson’s trichrome staining ( n = 6). l , m immunohistochemistry and quantification of renal sections for the podocyte marker Nephrin from the indicated groups ( n = 6). n , o Representative images of isolated mouse glomeruli stained with WT-1 (green), TUNEL (green), and DHE (red), with quantification shown in ( o ) ( n = 6). Nuclei were counterstained with DAPI (blue). Data are presented as mean ± s.e.m. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 as determined by ordinary one-way ANOVA followed by Tukey’s multiple comparisons test ( a − o ); ns, not significant.

Article Snippet: The sections were then washed three times with PBS and incubated with Alexa Fluor 488- or 647-conjugated secondary antibodies (1:300, Cat. No. ab150073 or ab150115, Abcam) for 1 h. The sections were then incubated with DAPI (Southern Biotech, Birmingham) for 10 min. Co-staining for FGFR1 (1:1000, Cat. No. 60325, Proteintech) and NPHS2 (1:1000, Cat. No. 20384, Proteintech) was performed using a multiplex staining kit (Cat. No. AFIHC034, AiFang Biological) based on Tyramide Signal Amplification (TSA) technology.

Techniques: Immunofluorescence, Marker, Knock-Out, Western Blot, Isolation, Staining, Immunohistochemistry, TUNEL Assay

Journal: Nature Communications

Article Title: FGF4-FGFR1 signaling promotes podocyte survival and glomerular function to ameliorate diabetic kidney disease in male mice

doi: 10.1038/s41467-025-65978-4

Figure Lengend Snippet: a , b Human podocytes were isolated from the urine of DKD patients and exposed to high glucose conditions with or without rFGF4 treatment. a Representative images of TUNEL staining (green) and quantification of TUNEL-positive cells from the indicated groups ( n = 6). b Immunofluorescence analysis of tubulin (green) and quantification of the aspect ratio (width to height ratio) of human podocytes ( n = 6). White dashed lines mark the edge of the cells. c Representative images and quantification of isolated human glomeruli stained for WT-1 (green), TUNEL (green), DHE (red), and Nrf-2 (pink). Nuclei were counterstained with DAPI (blue). d Western blot and quantitative analysis of c-Cas3, CAT, SOD-2, and Nrf-2 protein levels in human glomeruli from the indicated groups ( n = 4). β-actin served as a loading control. e Western blot analysis of the activation of AMPK-FOXO1 pathway components in total glomerular tissue lysate. The ratios of phosphorylated proteins to total proteins are shown as scatter plots ( n = 4). f Schematic showing that FGF4 protects against DKD by activating the FGFR1-AMPK-FOXO1 signaling axis. Data are presented as mean ± s.e.m. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 as determined by unpaired two-tailed Student’s t -test ( a , b ) or ordinary one-way ANOVA followed by Tukey’s multiple comparisons test ( c − e ); ns, not significant. c-Cas3, cleaved-Caspase3; CAT, Catalase.

Article Snippet: The sections were then washed three times with PBS and incubated with Alexa Fluor 488- or 647-conjugated secondary antibodies (1:300, Cat. No. ab150073 or ab150115, Abcam) for 1 h. The sections were then incubated with DAPI (Southern Biotech, Birmingham) for 10 min. Co-staining for FGFR1 (1:1000, Cat. No. 60325, Proteintech) and NPHS2 (1:1000, Cat. No. 20384, Proteintech) was performed using a multiplex staining kit (Cat. No. AFIHC034, AiFang Biological) based on Tyramide Signal Amplification (TSA) technology.

Techniques: Isolation, TUNEL Assay, Staining, Immunofluorescence, Western Blot, Control, Activation Assay, Two Tailed Test