Journal: bioRxiv

Article Title: Membrane-associated polymerases deliver most of the actin subunits to a lamellipodial network

doi: 10.1101/2025.03.24.645090

Figure Lengend Snippet: A. Schematic representation of the route actin takes into filaments through the actin-binding protein profilin. The covalently linked actin-profilin probe can enter filaments directly from the cytosol and by first binding to the proline-rich region of NPFs like WAVE1. A similar probe that contains a modified profilin that can still incorporate into filaments but cannot bind proline-rich domains (actin-profilin MUT ) only enters the filament directly from the cytosol. B. Illustration of actin-profilin orientations for linked constructs using previously published structures (PDB:2BTF) with either 10, 16 or 30 alternating glycine-serine residues between the N terminus of β-actin and the C-terminus of profilin. EGFP is linked to the N terminus of β-actin for probe visualization. C. Fluorescence of B16F1 cells transiently expressing indicated constructs. Scale bar represents 5µm. D. Averaged linescans at the leading edge of cells expressing actin-profilin probes with different linker lengths, where the cell edge = 0. The bands depict 95% confidence intervals. For all conditions, n = 100 lines drawn from 10 cells. Lines are normalized to the cell mean intensity. Only the probe with the longest linker shows a similar increase at the leading edge to eGFP-actin. E. Quantification of mobile fraction of recovery curve after photobleaching per cell indicating that both actin and actinGS 30 profilin readily encorporate into filaments, as they both exhibit substantial nonmobile fractions. Number of cells per experiment per round for both A and B are as follows with cell number indicated in parenthesis: eGFP (37,23,22) eGFP-actin WT (22,27), eGFP-profilin1 (11,9,26), β-actinGS 10 profilin (23,18), β-actinGS 16 profilin (21,12,23), β-actinGS 30 profilin (16,35,14). One-way ANOVA and Tukey’s multiple comparison tests are comparisons with eGFP.

Article Snippet: B16F1 (ATCC-CRL-6323) mouse melanoma cells were cultured in DMEM/F12 with 4.5 g/mL glucose and supplemented with 10% fetal bovine serum (FBS, Life Technologies Certified, US) and penicillin/streptomycin.

Techniques: Binding Assay, Modification, Construct, Fluorescence, Expressing, Comparison

Journal: bioRxiv

Article Title: Membrane-associated polymerases deliver most of the actin subunits to a lamellipodial network

doi: 10.1101/2025.03.24.645090

Figure Lengend Snippet: A. Cartoon illustrating actin polymerization with the WAVE1 regulatory complex with the proline-rich region intact (left) and a WAVE1 with its proline-rich region mutated so that profilin cannot bind (right). B. Diagram of the PVCA domain of WAVE1 depicting modifications made to the proline-rich domain to alter binding of profilin-actin (profilin-binding deficient). C. Fluorescence of B16F1 depleted of both WAVE1 and 2 isoforms (WAVE1/2 double knockout) and rescues transiently expressing indicated constructs. B16F1 wild-type cells are shown for comparison. EGFP-WAVE1 with a modified proline-rich domain (WAVE1 MUT ) is shown in comparison to the intact domain (WAVE1 WT ), with magnified insets of the leading edge. The wildtype WAVE1 rescues the typical lamellipodia phenotype, while cells with the WAVE1 MUT that cannot bind profilin exhibit many filopodia, more similar to the double knockout cells. Scale bar represents 5 µm for inset, 10 µm for whole image. D. Principal component analysis of cell shape for various cell lines. The cells rescued with wild type WAVE1 appear more similar to wild type B16F1 cells, while cells transiently expressing only the WAVE1 MUT construct are intermediate between the wild type and double-knockout cells. The first principal component (PC1, x-axis) explains 84.1% of the variation in the data, while the second principal component (PC2, y-axis) represents 15.9% of the variation. Confidence ellipses are drawn at 95% for each group. Means for each group are displayed by larger sizes and representative outlines of cell shape for each mean are displayed in the corners. E. Filopodia number per cell. B16F1 wildtype cells (16, 14, 15), double-knockout cells (16, 37), and double-knockout cells transiently expressing either WAVE1 MUT (10,10,12,23) or WAVE1 WT (8,15,20). F. Filopodia length in microns, n= filopodia number per round and are as follows: (58,88,69), (290,536), (79,99,11), and (52,20,60), respectively. Replicate means are represented by large dots superimposed on cell-level data. One-way ANOVA and Tukey’s multiple comparison tests were performed on replicate means.

Article Snippet: B16F1 (ATCC-CRL-6323) mouse melanoma cells were cultured in DMEM/F12 with 4.5 g/mL glucose and supplemented with 10% fetal bovine serum (FBS, Life Technologies Certified, US) and penicillin/streptomycin.

Techniques: Binding Assay, Fluorescence, Double Knockout, Expressing, Construct, Comparison, Modification

Journal: Cellular & Molecular Biology Letters

Article Title: Orally administered endoxifen inhibits tumor growth in melanoma-bearing mice

doi: 10.1186/s11658-017-0068-7

Figure Lengend Snippet: Therapeutic efficacy of endoxifen in the B16F10 melanoma tumor model in mice. Endoxifen or tamoxifen significantly reduced nodule counts when compared to the control group ( p < 0.05). At an equal dose of 8 mg/kg body weight, endoxifen exhibited significantly stronger activity in reducing melanoma nodule counts than tamoxifen ( p < 0.05). The data represent means ± SD, ( n = 5)

Article Snippet: Mouse melanoma cell line B16F10 (ATCC) was maintained in DMEM supplemented with 10% FBS, 2 mM glutamine, 100 units/ml penicillin and 100 μg/ml streptomycin at 37 °C in a humidified atmosphere of 5% CO 2 .

Techniques: Drug discovery, Control, Activity Assay

Journal: British Journal of Cancer

Article Title: In vivo evolution of tumour cells after the generation of double-strand DNA breaks

doi: 10.1038/sj.bjc.6600959

Figure Lengend Snippet: Histological characteristics of the untreated murine fibrosarcoma tumour LPB ( A and B ) and murine melanoma tumour B16F0 ( C and D ). Small arrows: mitotic cells; large arrows: apoptotic cells; arrowheads: atypical cells.

Article Snippet: The LPB cell line, a methylcholanthrene-induced C57Bl/6 mouse sarcoma cell line ( Belehradek et al , 1972 ), and the B16 F0 melanoma cells (ATCC CRL 6322) were cultured using classical procedures and minimum essential medium culture medium (Gibco BRL, Cergy-Pontoise, France) supplemented with 100 U ml −1 penicillin, 100 mg ml −1 streptomycin (Sarbach, France) and 8% fetal calf serum (Gibco).

Techniques:

Journal: British Journal of Cancer

Article Title: In vivo evolution of tumour cells after the generation of double-strand DNA breaks

doi: 10.1038/sj.bjc.6600959

Figure Lengend Snippet: Kinetics of the fraction of ( I ) mitotic cells, ( II ) apoptotic cells and ( III ) atypical cells in LPB and B16F0 tumours after ECT using 1 mg BLM (panels A) or 10 μ g BLM (panels B). Statistical analysis: all groups were compared to the respective controls by means of Student's t -test. All groups were significantly different from the controls (at least P <0.05) except in panels IIA LPB (at 1 and 8 h), IIA B16F0 (at 5 min and 1 h), IIB LPB (before 20 h), IIB B16F0 before 25 h, IIIA B16F0 (at 5 min and 1 h), IIIA LPB (20 min, 1 h and 2 h), IIIB B16F0 at 50 h and IIIB LPB at 15, 20, 25 and after 70 h.

Article Snippet: The LPB cell line, a methylcholanthrene-induced C57Bl/6 mouse sarcoma cell line ( Belehradek et al , 1972 ), and the B16 F0 melanoma cells (ATCC CRL 6322) were cultured using classical procedures and minimum essential medium culture medium (Gibco BRL, Cergy-Pontoise, France) supplemented with 100 U ml −1 penicillin, 100 mg ml −1 streptomycin (Sarbach, France) and 8% fetal calf serum (Gibco).

Techniques:

Journal: British Journal of Cancer

Article Title: In vivo evolution of tumour cells after the generation of double-strand DNA breaks

doi: 10.1038/sj.bjc.6600959

Figure Lengend Snippet: Histological changes observed in tumours removed 30 min after ECT using 1 mg BLM ( A – D ) and in tumours removed 50 h after ECT using 10 μ g BLM ( E and F ); ( A , C and E ): LPB tumours; ( B , D and F ): B16F0 tumours.

Article Snippet: The LPB cell line, a methylcholanthrene-induced C57Bl/6 mouse sarcoma cell line ( Belehradek et al , 1972 ), and the B16 F0 melanoma cells (ATCC CRL 6322) were cultured using classical procedures and minimum essential medium culture medium (Gibco BRL, Cergy-Pontoise, France) supplemented with 100 U ml −1 penicillin, 100 mg ml −1 streptomycin (Sarbach, France) and 8% fetal calf serum (Gibco).

Techniques:

Journal: British Journal of Cancer

Article Title: In vivo evolution of tumour cells after the generation of double-strand DNA breaks

doi: 10.1038/sj.bjc.6600959

Figure Lengend Snippet: Immunohistochemical detection of DNA fragmentation in tumour cells. Fluorescein detection ( A – C ); alkaline phosphatase detection ( D – F ); ( A – C ): LPB fibrosarcoma; ( D – F ): B16F0 melanoma; ( A and D ): stained untreated controls; ( B and E ): staining of tumours removed 30 min after ECT using 1 mg BLM; ( C and F ): staining of tumours removed 50 h after ECT using 10 μ g BLM.

Article Snippet: The LPB cell line, a methylcholanthrene-induced C57Bl/6 mouse sarcoma cell line ( Belehradek et al , 1972 ), and the B16 F0 melanoma cells (ATCC CRL 6322) were cultured using classical procedures and minimum essential medium culture medium (Gibco BRL, Cergy-Pontoise, France) supplemented with 100 U ml −1 penicillin, 100 mg ml −1 streptomycin (Sarbach, France) and 8% fetal calf serum (Gibco).

Techniques: Immunohistochemical staining, Staining

Journal: Experimental & Molecular Medicine

Article Title: H3K18 lactylation in cancer-associated fibroblasts drives malignant pleural effusion progression via TNFR2 + T reg recruitment

doi: 10.1038/s12276-025-01557-3

Figure Lengend Snippet: a KEGG pathway enrichment of DEGs between FACS-sorted TNFR2 + and TNFR2 − CD4 + T cells from MPE ( n = 3). b , c Volcano plot ( b ) and heatmap ( c ) of DEGs related to the chemokine signaling pathway. d , e Flow cytometry histograms ( d ) and comparisons of chemokine receptor expression (CXCR6, CCR4 and CCR6) ( e ) on TNFR2 + T reg cells and TNFR2 − T reg cells ( n = 13). f Schematic of Transwell chemotaxis assay testing TNFR2 − T reg chemotaxis toward MPE supernatant (by Figdraw). g , h Transwell chemotaxis assay comparing TNFR2 + T reg frequencies between freshly isolated cells and cells that migrated toward MPE supernatant after 4 h incubation. i Concentrations of CXCL16, CCL17, CCL22 and CCL20 in MPE and PB were quantified using ELISA ( n = 16). j Schematic of chemotaxis assay to investigate the chemotatic axis to attract TNFR2 + T reg cells in MPE. k , l Flow cytometry histograms and comparisons of chemotaxis of TNFR2 + T reg cells in response to MPE in the presence of anti-CXCL16, anti-CCL17, anti-CCL22 or anti-CCL20 mAbs. Data shown in d , e , g – i , k and l are representative of at least three independent experiments (mean ± s.d.). Statistical analysis was performed using paired two-tailed Student’s t -test ( e and h ), Wilcoxon test ( i ) or one-way ANOVA ( l ). * P < 0.05, ** P < 0.01, **** P < 0.0001. ns not significant, mAbs monoclonal antibodies.

Article Snippet: The membrane was blocked with 5% skim milk in Tris-buffered saline containing 0.05% Tween 20 (TBST) at room temperature for 2 h, followed by overnight incubation with primary antibodies at 4 °C, including anti-Pan-Kla antibody (#PTM-1401, PTM BIO), anti-H3K18la antibody (#PTM-1406RM, PTM BIO), anti-H3K9la antibody (#PTM-1419RM, PTM BIO), anti-H3K27la antibody (#PTM-1428, PTM BIO), anti-H4K12la antibody (#PTM-1411RM, PTM BIO), anti-H4K8la antibody (#PTM-1415RM, PTM BIO), anti-CXCL16 antibody (#60123-1-Ig, Proteintech), anti-FOXO3 antibody(#10849-1-AP, Proteintech), anti-GAPDH antibody (#60004-1-Ig, Proteintech) and anti-histone H3 antibody (#17168-1-AP, Proteintech).

Techniques: Flow Cytometry, Expressing, Chemotaxis Assay, Isolation, Incubation, Enzyme-linked Immunosorbent Assay, Two Tailed Test, Bioprocessing

Journal: Experimental & Molecular Medicine

Article Title: H3K18 lactylation in cancer-associated fibroblasts drives malignant pleural effusion progression via TNFR2 + T reg recruitment

doi: 10.1038/s12276-025-01557-3

Figure Lengend Snippet: a Flow cytometry analysis comparing of CXCL16 expression on CD45 + and CD45 − cells from MPE ( n = 9). b CXCL16 expression on major cell populations of MPE: CAFs (CD45 − FAP + ), neutrophils (CD45 + CD11b + Ly6G + ), macrophages (CD45 + CD68 + ), T lymphocytes (CD45 + CD3 + ) and B lymphocytes (CD45 + CD19 + ) from MPE ( n = 8). c Schematic of CAF purification from MPE using MACS (by Figdraw). d Schematic of Transwell assay testing TNFR2 + T reg chemotaxis toward CXCL16-supplemented medium or CAF culture supernatant, with or without anti-CXCL16 mAbs (by Figdraw). e , f Representative flow cytometry plots ( e ) and comparisons ( f ) of TNFR2 + T reg frequencies recruited under different conditions ( n = 3). g Schematic diagram of the MPE mouse model. h Representative images showing mouse MPE and pleural cavity tumors. i – k Bioluminescence images depicting the growth ( n = 5) ( i ), MPE volume ( n = 5) ( j ) and Kaplan–Meier survival plot ( k ) of MPE mice ( n = 9–10 per group). l Concentrations of CXCL16 in MPE from mouse models ( n = 6). m , n Frequencies of TNFR2 + cells among T reg cells in murine MPE of each group. o , p Frequencies of CD8 + T cells from mouse MPE in each group. Data shown in a , b , e , f and h – p are representative of at least three independent experiments (mean ± s.d.). Statistical analysis was performed using paired two-tailed Student’s t -test ( a ), unpaired two-tailed Student’s t -test ( l ), one-way ANOVA ( b , f , j , n and p ) or log-rank test ( k ). * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. ns not significant, CS culture supernatant, MACS magnetic-activated cell sorting, α-CXCL16 CXCL16 neutralizing antibody.

Article Snippet: The membrane was blocked with 5% skim milk in Tris-buffered saline containing 0.05% Tween 20 (TBST) at room temperature for 2 h, followed by overnight incubation with primary antibodies at 4 °C, including anti-Pan-Kla antibody (#PTM-1401, PTM BIO), anti-H3K18la antibody (#PTM-1406RM, PTM BIO), anti-H3K9la antibody (#PTM-1419RM, PTM BIO), anti-H3K27la antibody (#PTM-1428, PTM BIO), anti-H4K12la antibody (#PTM-1411RM, PTM BIO), anti-H4K8la antibody (#PTM-1415RM, PTM BIO), anti-CXCL16 antibody (#60123-1-Ig, Proteintech), anti-FOXO3 antibody(#10849-1-AP, Proteintech), anti-GAPDH antibody (#60004-1-Ig, Proteintech) and anti-histone H3 antibody (#17168-1-AP, Proteintech).

Techniques: Flow Cytometry, Expressing, Purification, Transwell Assay, Chemotaxis Assay, Two Tailed Test, FACS

Journal: Experimental & Molecular Medicine

Article Title: H3K18 lactylation in cancer-associated fibroblasts drives malignant pleural effusion progression via TNFR2 + T reg recruitment

doi: 10.1038/s12276-025-01557-3

Figure Lengend Snippet: a GSVA of signaling pathways enriched in CXCL16 + CAFs compared with CXCL16 − CAFs. b , c Gene set enrichment analysis (GSEA) was performed on gene sets related to the hypoxia signaling pathway ( b ) and the glycolysis signaling pathway ( c ). A positive NES indicates higher expression in CXCL16 + CAFs. d , e Lactate levels in CAFs cultured under normoxia (21% oxygen) or hypoxia (1% oxygen) for 48 h or at indicated time points ( n = 3). f , g CXCL16 concentrations in CAFs supernatant under normoxia or hypoxia for 48 h or at indicated time points ( n = 3). h Western blot of CXCL16 expression in CAFs under hypoxia at designated times. i Schematic of metabolic modulators targeting glycolysis or lactate production. j , k Lactate levels ( j ) and CXCL16 concentrations ( k ) in CAFs treated with indicated glycolysis modulators for 48 h ( n = 3). l – n Dose-dependent effects of DCA ( l ), oxamate ( m ) or rotenone ( n ) on lactate and CXCL16 levels in CAFs supernatant. o Western blot analysis of CXCL16 expression in CAFs treated with the indicated concentrations of glycolysis modulators for 48 h. Data shown in d – h and j – o are representative of at least three independent experiments (mean ± s.d.). Statistical analysis was performed using unpaired two-tailed Student’s t -test ( d and f ) or one-way ANOVA ( e , g and j – n ). * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. ns not significant, NES normalized enrichment score, DMSO dimethylsulfoxide, DCA dichloroacetate.

Article Snippet: The membrane was blocked with 5% skim milk in Tris-buffered saline containing 0.05% Tween 20 (TBST) at room temperature for 2 h, followed by overnight incubation with primary antibodies at 4 °C, including anti-Pan-Kla antibody (#PTM-1401, PTM BIO), anti-H3K18la antibody (#PTM-1406RM, PTM BIO), anti-H3K9la antibody (#PTM-1419RM, PTM BIO), anti-H3K27la antibody (#PTM-1428, PTM BIO), anti-H4K12la antibody (#PTM-1411RM, PTM BIO), anti-H4K8la antibody (#PTM-1415RM, PTM BIO), anti-CXCL16 antibody (#60123-1-Ig, Proteintech), anti-FOXO3 antibody(#10849-1-AP, Proteintech), anti-GAPDH antibody (#60004-1-Ig, Proteintech) and anti-histone H3 antibody (#17168-1-AP, Proteintech).

Techniques: Protein-Protein interactions, Expressing, Cell Culture, Western Blot, Two Tailed Test

Journal: Experimental & Molecular Medicine

Article Title: H3K18 lactylation in cancer-associated fibroblasts drives malignant pleural effusion progression via TNFR2 + T reg recruitment

doi: 10.1038/s12276-025-01557-3

Figure Lengend Snippet: a Western blot analysis of the indicated proteins in CAFs treated with increasing concentrations of exogenous lactate or glucose for 48 h. b CXCL16 concentrations in CAFs supernatant after exogenous lactate treatment. c , d Lactate ( c ) and CXCL16 ( d ) levels in CAFs supernatant after treatment with glucose for 48 h. e Western blot analysis of the indicated proteins in CAFs treated with DCA, oxamate or rotenone for 48 h. f Schematic of LDHA knockdown in CAFs. g Lactate in the culture supernatant of LDHA-knockdown CAFs. h Western blot analysis of the indicated proteins in LDHA-knockdown CAFs. i CXCL16 levels in the supernatant of LDHA-knockdown CAFs. j Western blot analysis of the indicated proteins in CAFs treated with increasing concentrations of β-alanine, lactylation modulator, for 48 h. k CXCL16 concentrations in CAFs supernatant after β-alanine treatment. Data shown in a – e and g – k are representative of at least three independent experiments (mean ± s.d.). Statistical analysis was performed using one-way ANOVA ( b – d , g , i and k ). * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. ns not significant, KD knockdown, sh-LDHA shRNA against LDHA, sh-NC negative control.

Article Snippet: The membrane was blocked with 5% skim milk in Tris-buffered saline containing 0.05% Tween 20 (TBST) at room temperature for 2 h, followed by overnight incubation with primary antibodies at 4 °C, including anti-Pan-Kla antibody (#PTM-1401, PTM BIO), anti-H3K18la antibody (#PTM-1406RM, PTM BIO), anti-H3K9la antibody (#PTM-1419RM, PTM BIO), anti-H3K27la antibody (#PTM-1428, PTM BIO), anti-H4K12la antibody (#PTM-1411RM, PTM BIO), anti-H4K8la antibody (#PTM-1415RM, PTM BIO), anti-CXCL16 antibody (#60123-1-Ig, Proteintech), anti-FOXO3 antibody(#10849-1-AP, Proteintech), anti-GAPDH antibody (#60004-1-Ig, Proteintech) and anti-histone H3 antibody (#17168-1-AP, Proteintech).

Techniques: Western Blot, Knockdown, shRNA, Negative Control

Journal: Experimental & Molecular Medicine

Article Title: H3K18 lactylation in cancer-associated fibroblasts drives malignant pleural effusion progression via TNFR2 + T reg recruitment

doi: 10.1038/s12276-025-01557-3

Figure Lengend Snippet: a IGV tracks presenting H3K18la enrichment at the CXCL16 gene locus in CAFs from MPE by CUT&Tag analysis. b ChIP–qPCR validation of H3K18la binding at the CXCL16 promoter. c – e Predicted FOXO3 binding sites at the CXCL16 promoter using JASPAR ( http://jaspar.genereg.net ). f ChIP–qPCR validation of FOXO3 binding at the CXCL16 gene promoter in CAFs. g Western blot analysis of FOXO3 and CXCL16 expression following FOXO3 knockdown or overexpression in CAFs. h , i CXCL16 levels in CAFs supernatants after FOXO3 knockdown ( h ) or overexpression ( i ). j Schematic of dual-luciferase reporter assay, drawn by Figdraw. k Luciferase assays assessing FOXO3-mediated regulation of CXCL16 transcription in CAFs. l , m CUT&Tag ( l ) and ChIP–qPCR ( m ) analyses showing H3K18la enrichment at the FOXO3 promoter. n Western blot analysis of the indicated proteins in CAFs treated with glucose (0–20 mm/l), DCA (0–20 mm/l), oxamate (0–20 mm/l), retenone (0–50 nm/l) or β-alanine (0–20 mm/l) for 48 h. o Western blot analysis of the indicated proteins in LDHA-knockdown CAFs. Data shown in b , f – i , k and m – o are representative of at least three independent experiments (mean ± s.d.). Statistical analysis was performed using unpaired two-tailed Student’s t -test ( h and i ) or one-way ANOVA ( b , f , k and m ). * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. ns not significant, IGV Integrative Genomics Viewer, OE overexpression.

Article Snippet: The membrane was blocked with 5% skim milk in Tris-buffered saline containing 0.05% Tween 20 (TBST) at room temperature for 2 h, followed by overnight incubation with primary antibodies at 4 °C, including anti-Pan-Kla antibody (#PTM-1401, PTM BIO), anti-H3K18la antibody (#PTM-1406RM, PTM BIO), anti-H3K9la antibody (#PTM-1419RM, PTM BIO), anti-H3K27la antibody (#PTM-1428, PTM BIO), anti-H4K12la antibody (#PTM-1411RM, PTM BIO), anti-H4K8la antibody (#PTM-1415RM, PTM BIO), anti-CXCL16 antibody (#60123-1-Ig, Proteintech), anti-FOXO3 antibody(#10849-1-AP, Proteintech), anti-GAPDH antibody (#60004-1-Ig, Proteintech) and anti-histone H3 antibody (#17168-1-AP, Proteintech).

Techniques: ChIP-qPCR, Biomarker Discovery, Binding Assay, Western Blot, Expressing, Knockdown, Over Expression, Luciferase, Reporter Assay, Two Tailed Test

Journal: Experimental & Molecular Medicine

Article Title: H3K18 lactylation in cancer-associated fibroblasts drives malignant pleural effusion progression via TNFR2 + T reg recruitment

doi: 10.1038/s12276-025-01557-3

Figure Lengend Snippet: a Schematic illustrating the the generation of LDHA-knockout NIH/3T3 fibroblasts (LDHA −/− 3T3) using CRISPR–Cas9, drawn by Figdraw. b Western blot validation of LDHA knockout in LDHA −/− 3T3 fibroblasts. c , d Lactate ( c ) and CXCL16 ( d ) levels in the culture supernatant of LDHA −/− 3T3 fibroblasts. e Western blot analysis of the indicated proteins in LDHA −/− 3T3 fibroblasts cultured with or without glucose. f Schematic of MPE mouse model. g Representative images showing mouse MPE and thoracic tumors in each group. h – j, Bioluminescence images depicting the growth ( n = 5) ( h ), MPE volume ( n = 6) ( i ) and Kaplan–Meier survival curves (n = 10 per group) ( j ) of MPE-bearing mice. Data shown in c – e and g – j are representative of at least three independent experiments (mean ± s.d.). Statistical analysis was performed using unpaired two-tailed Student’s t -test ( c and d ), one-way ANOVA ( i ) and log-rank test ( j ). * P < 0.05, ** P < 0.01, *** P < 0.001, ns not significant, LDHA−/−3T3 LDHA-knockout NIH/3T3 fibroblasts.

Article Snippet: The membrane was blocked with 5% skim milk in Tris-buffered saline containing 0.05% Tween 20 (TBST) at room temperature for 2 h, followed by overnight incubation with primary antibodies at 4 °C, including anti-Pan-Kla antibody (#PTM-1401, PTM BIO), anti-H3K18la antibody (#PTM-1406RM, PTM BIO), anti-H3K9la antibody (#PTM-1419RM, PTM BIO), anti-H3K27la antibody (#PTM-1428, PTM BIO), anti-H4K12la antibody (#PTM-1411RM, PTM BIO), anti-H4K8la antibody (#PTM-1415RM, PTM BIO), anti-CXCL16 antibody (#60123-1-Ig, Proteintech), anti-FOXO3 antibody(#10849-1-AP, Proteintech), anti-GAPDH antibody (#60004-1-Ig, Proteintech) and anti-histone H3 antibody (#17168-1-AP, Proteintech).

Techniques: Knock-Out, CRISPR, Western Blot, Biomarker Discovery, Cell Culture, Two Tailed Test

Journal: Experimental & Molecular Medicine

Article Title: H3K18 lactylation in cancer-associated fibroblasts drives malignant pleural effusion progression via TNFR2 + T reg recruitment

doi: 10.1038/s12276-025-01557-3

Figure Lengend Snippet: a , b Lactate ( a ) and CXCL16 ( b ) levels measured in MPE from mice treated with LDHA −/− 3T3 fibroblasts or control fibroblasts. c – j Flow cytometry plots ( c , e , g and i ) and comparisons ( d , f , h and j ) of the frequencies of TNFR2 + T reg cells ( c and d ), IFN-γ + CD8 + T cells ( e and f ), granzyme B + CD8 + T cells ( g and h ) and perforin + CD8 + T cells ( i and j ) in MPE. k Schematic diagram illustrating the proposed mechanism: CAFs in MPE undergo glycolysis, leading to elevated endogenous lactate levels. This increase in lactate induces H3K18 lactylation modification at the promoter regions of both the CXCL16 gene and its transcription factor FOXO3, thereby promoting CXCL16 expression. TNFR2 + T reg cells, which express high levels of CXCR6, the only known receptor for CXCL16, are efficiently recruited into MPE. This recruitment dampens the antitumor response generated by CD8 + T cells, leading to the immunosuppression and progression of MPE (by Figdraw). Data shown in a – j are representative of at least three independent experiments (mean ± s.d.). Statistical analysis was performed using unpaired two-tailed Student’s t -test ( a and b ) and one-way ANOVA ( d , f , h and j ). ** P < 0.01, *** P < 0.001, **** P < 0.0001. ns not significant.

Article Snippet: The membrane was blocked with 5% skim milk in Tris-buffered saline containing 0.05% Tween 20 (TBST) at room temperature for 2 h, followed by overnight incubation with primary antibodies at 4 °C, including anti-Pan-Kla antibody (#PTM-1401, PTM BIO), anti-H3K18la antibody (#PTM-1406RM, PTM BIO), anti-H3K9la antibody (#PTM-1419RM, PTM BIO), anti-H3K27la antibody (#PTM-1428, PTM BIO), anti-H4K12la antibody (#PTM-1411RM, PTM BIO), anti-H4K8la antibody (#PTM-1415RM, PTM BIO), anti-CXCL16 antibody (#60123-1-Ig, Proteintech), anti-FOXO3 antibody(#10849-1-AP, Proteintech), anti-GAPDH antibody (#60004-1-Ig, Proteintech) and anti-histone H3 antibody (#17168-1-AP, Proteintech).

Techniques: Control, Flow Cytometry, Modification, Expressing, Generated, Two Tailed Test

Journal: Hepatology Communications

Article Title: CXCR6 recruits Th2 cells and promotes liver fibrosis in schistosomiasis japonica

doi: 10.1097/HC9.0000000000000784

Figure Lengend Snippet: CXCL16 was primarily derived from macrophages in schistosome-associated fibrotic livers. (A) tSNE plot showing the distribution of different cell clusters. (B) Bubble plot of Cellchat analysis between different cell clusters in the CXCL signaling network. The closer to red, the greater the proportion of interaction. (C) Representative western blot images for CXCL16 in schistosome-associated fibrotic liver (SJ) and normal mouse liver (NC). (D) Violin plots of Cxcl16 in different cell clusters. (E) Proportions of different cell clusters in single-cell sequencing. (F–H) Expression of CXCL16 in macrophages. Gating strategy for macrophages (F4/80 + CD11b + CD45 + ). Statistical significance was determined using an unpaired t test, and the data are presented as mean ± SD (* p <0.05; ** p <0.01; *** p <0.001; **** p <0.0001). Abbreviation: MP, mononuclear phagocytes.

Article Snippet: Primary antibodies against α-SMA (1:1000, Zenbio) and CXCL16 (1:500, Proteintech) were incubated overnight at 4 °C, with GAPDH and β-tubulin serving as loading controls.

Techniques: Derivative Assay, Western Blot, Sequencing, Expressing