Journal: Nature Communications

Article Title: Targeting hepatocytic TβRI ameliorates liver metastatic outcomes by revitalizing stem-like CD8 + Tex subsets

doi: 10.1038/s41467-025-65615-0

Figure Lengend Snippet: a IHC staining with TβRI-specific antibody to detect the distribution of TβRI + cells in colorectal liver metastases ( n = 36 patients). Representative IHC image. Scale bar, 50 μm. The bar chart in the right represents the percentage of high and low TβRI expression cases in liver metastasis, adjacent and distal liver tissues. p < 0.001 by two-sided Pearson’s chi-square test. b Overall survival curves based on adjacent liver TβRI expressed level in CRLM patients using the multivariate COX regression analysis, a two-sided P -value of < 0.05 was considered statistically significant. c Representative images depicting multicolor immunohistochemical co-staining of TβRI with specific cell type markers in CRLM adjacent liver tissues (DAPI: cell nucleus, HNF4α: hepatocytes, α-SMA: hepatic stellate cells, CD68: macrophages, PAN-CK: tumor cells). Scale bar, 25 μm. d Experimental scheme for generating TβRI flox/flox Alb-Cre +/- ( TβRI-hKO ) mice. TβRI flox/flox Alb-Cre -/- and C57BL/6 J were used as TβRI-hWT mice. e Representative bioluminescent image and Quantification of TβRI-hWT versus TβRI-hKO mice inoculated with MC38-Luc cells ( n = 6 mice/group). f Representative macroscopic image of ( e ) and Quantification of Liver to body weight ratio (LBR). g Representative bioluminescent image of TβRI-hWT versus TβRI-hKO mice inoculated with PAN02-Luc cells ( n = 5 mice/group). h Quantification of ( g ). i Representative macroscopic image of ( g ) and Quantification of LBR. j Representative bioluminescent image of TβRI-hWT versus TβRI-hKO mice inoculated with B16F10-Luc cells ( n = 5 mice/group). k Quantification of ( j ). l Representative macroscopic image of ( j ) and Quantification of LBR. Data are presented as the mean ± SD, and were analyzed with an unpaired two-sided Student’s t test ( e–l ). Consistent results were observed across three biological replicates in ( e–l ). Source data are provided as a Source Data file.

Article Snippet: The above findings prompted us to explore whether therapeutic interventions, specifically Galectin-9 neutralization (αGal-9) or TβRI inhibition (Galunisertib) , would similarly alleviate liver metastasis by reshaping CD8 + T cell responses in our mouse model.

Techniques: Immunohistochemistry, Expressing, Immunohistochemical staining, Staining

Journal: Nature Communications

Article Title: Targeting hepatocytic TβRI ameliorates liver metastatic outcomes by revitalizing stem-like CD8 + Tex subsets

doi: 10.1038/s41467-025-65615-0

Figure Lengend Snippet: a , b Frequency of T cell subsets based on the expression of Ly108 and CX3CR1 within CD8 + PD1 + CD44 + tumor-reactive T cells in different anatomical locations of TβRI-hWT and TβRI-hKO mice ( n = 6 mice/group). Flow cytometry representative plots ( a ). Statistical analysis of these subsets in liver metastases across multiple tumor types are shown ( b ). Statistical analysis of these subsets in the spleen and PBMC are not shown. c–f Flow cytometry was used to detect the phenotypes, transcriptional profiles, and effector functions of Ly108 + CX3CR1 - , Ly108 + CX3CR1 + , Ly108 - CX3CR1 + , Ly108 - CX3CR1 - subsets in liver metastases of TβRI-hWT and TβRI-hKO mice inoculated with PAN02 on day 28. c Representative histograms showing the expression of PD-1, LAG3, TIGIT, TIM3, and 2B4 among these T subsets ( n = 6 mice/group). MFI analysis of TIM3 and 2B4 was summarized in the right statistical chart. d Representative flow cytometry plots and statistical analysis depicting the TOX/TCF1 co-expression among these T subsets ( n = 4 mice/group). e Representative flow cytometry plots and statistical analysis depicting the TNF and IFNγ co-production by these T subsets upon ex vivo CD3/CD28 stimulation ( n = 5 mice/group). f Representative flow cytometry plots and statistical analysis depicting the percentages of these T subsets degranulating (CD107a) and producing IL2 upon ex vivo CD3/CD28 stimulation ( n = 5 mice/group). Data are presented as the mean ± SD. Data were analyzed with unpaired two-sided Student’s t test ( b ) and two-way ANOVA with Tukey’s multiple comparisons test ( c–f ). Consistent results were observed across three biological replicates in ( a–f ). Source data are provided as a Source Data file.

Article Snippet: The above findings prompted us to explore whether therapeutic interventions, specifically Galectin-9 neutralization (αGal-9) or TβRI inhibition (Galunisertib) , would similarly alleviate liver metastasis by reshaping CD8 + T cell responses in our mouse model.

Techniques: Expressing, Flow Cytometry, Ex Vivo

Journal: Nature Communications

Article Title: Targeting hepatocytic TβRI ameliorates liver metastatic outcomes by revitalizing stem-like CD8 + Tex subsets

doi: 10.1038/s41467-025-65615-0

Figure Lengend Snippet: a Experimental design. b Representative bioluminescent image of pre-adoptive and post-adoptive C57BL/6 J mice receiving Ly108 + CX3CR1 - , Ly108 + CX3CR1 + , Ly108 - CX3CR1 + , Ly108 - CX3CR1 - cells transfer or without transfer ( n = 5 mice/group). c Quantification of ( b ). d Survival curve of tumor-bearing mice for indicated groups described in ( b ). Log-rank test for survival comparison. e Flow cytometry depicted the phenotype of adoptively transferred cells before and 14 days after transfer. All subsets shown were gated on CD8 + PD1 + CD44 + CD45.2 + CD45.1 - cells. f Representative bioluminescent image of TβRI-hWT and TβRI-hKO mice inoculated with TGFβ1 OE , TGFβ1 KD , or TGFβ1 CTL PAN02-Luc cells ( n = 6 mice/group). g Quantification of ( f ). Data are presented as the mean ± SD, and were analyzed with one-way ( g ) or two-way ( c ) ANOVA with Tukey’s multiple comparison test. Consistent results were observed across three biological replicates in ( b–g ). Source data are provided as a Source Data file.

Article Snippet: The above findings prompted us to explore whether therapeutic interventions, specifically Galectin-9 neutralization (αGal-9) or TβRI inhibition (Galunisertib) , would similarly alleviate liver metastasis by reshaping CD8 + T cell responses in our mouse model.

Techniques: Comparison, Flow Cytometry

Journal: Nature Communications

Article Title: Targeting hepatocytic TβRI ameliorates liver metastatic outcomes by revitalizing stem-like CD8 + Tex subsets

doi: 10.1038/s41467-025-65615-0

Figure Lengend Snippet: a Experimental setup to assess the indispensable role of TAMs or CD8 + T cells in improving metastatic outcomes of PAN02-bearing mice resulting from hepatocytic-TβRI ablation. b The M1-to-M2 ratio. c Representative bioluminescent image of TβRI-hWT and TβRI-hKO mice treated with vehicle (veh), Clodronate (CLD) liposome or αCD8 (n = 5 mice/group). d Survival curve of tumor-bearing mice for the indicated groups described in ( c ). Log-rank test for survival comparison. e Quantification of ( c ). f Representative flow cytometry plots and Statistical analysis depicting the percentages of Tex subsets in liver metastases of veh or CLD liposome-treated TβRI-hWT and TβRI-hKO mice on days 28. g Representative bioluminescent image and liver macroscopic image of TβRI-hWT and TβRI-hKO mice inoculated with PAN02 + BMDM or PAN02 alone ( n = 5 mice/group). h Quantification of ( g ). i Statistical analysis depicting the percentages of Tex subsets about the indicated groups. Data are presented as the mean ± SD, and were analyzed with one-way ( h ) or two-way ( b , e , f , i ) ANOVA with Tukey’s multiple comparison test. Consistent results were observed across three biological replicates in ( b–i ). Source data are provided as a Source Data file.

Article Snippet: The above findings prompted us to explore whether therapeutic interventions, specifically Galectin-9 neutralization (αGal-9) or TβRI inhibition (Galunisertib) , would similarly alleviate liver metastasis by reshaping CD8 + T cell responses in our mouse model.

Techniques: Comparison, Flow Cytometry

Journal: Nature Communications

Article Title: Targeting hepatocytic TβRI ameliorates liver metastatic outcomes by revitalizing stem-like CD8 + Tex subsets

doi: 10.1038/s41467-025-65615-0

Figure Lengend Snippet: a Luminex showing the protein levels of inflammatory cytokines in tumor interstitial fluid of TβRI-hWT and TβRI-hKO mice ( n = 3 mice/group). Data are shown as a heatmap. b Experimental scheme for generating Gal-9 flox/flox Alb-Cre +/- (Gal9-hKO) mice. Gal-9 flox/flox Alb-Cre -/- and C57BL/6 J were used as Gal9-hWT mice. c Representative bioluminescent image of Gal9-hWT versus Gal9-hKO mice inoculated with TGFβ1 OE or TGFβ1 CTL -PAN02 cells in the portal vein on day 28 ( n = 6 mice/group). d Bioluminescence quantification of ( c ). e Statistical analysis depicting the percentages of CD39 + TOX + cells within CD8 + PD1 + CD44 + tumor-reactive T cells in the indicated groups. f Statistical analysis depicting the percentages of Tex subsets based on TCF1 and CX3CR1 within CD8 + PD1 + CD44 + tumor-reactive T cells. g statistical analysis depicting the TNF and IFNγ co-production by CD8 + CD44 + T subsets in the indicated groups upon ex vivo CD3/CD28 stimulation. h The diagram of experimental design. i Representative bioluminescent image of Gal9-hWT versus Gal9-hKO mice treated with IgG, αCD4, αCD8 or αPD-1( n = 5 mice/group). j Representative macroscopic image of ( i ). k Bioluminescence quantification of ( i ). l Statistical analysis depicting the percentages of Tex subsets based on Ly108 and CX3CR1 within CD8 + PD1 + CD44 + tumor-reactive T cells. Data are presented as the mean ± SD, and were analyzed with unpaired two-sided Student’s t test ( k ) or two-way ( d , e , f , g , l ) ANOVA with Tukey’s multiple comparison test. Consistent results were observed across three biological replicates in ( a , c–g , i–l ). Source data are provided as a Source Data file.

Article Snippet: The above findings prompted us to explore whether therapeutic interventions, specifically Galectin-9 neutralization (αGal-9) or TβRI inhibition (Galunisertib) , would similarly alleviate liver metastasis by reshaping CD8 + T cell responses in our mouse model.

Techniques: Luminex, Ex Vivo, Comparison

Journal: Nature Communications

Article Title: Targeting hepatocytic TβRI ameliorates liver metastatic outcomes by revitalizing stem-like CD8 + Tex subsets

doi: 10.1038/s41467-025-65615-0

Figure Lengend Snippet: a Experimental setup to assess the effects of αGal-9 or Galunisertib monotherapy and combination therapy in improving outcomes in B16F10-bearing C57BL/6 J mice with liver metastases. ( n = 6 mice, Vehicle + IgG; n = 6 mice, Galunisertib + IgG; n = 6 mice, Vehicle + αGal-9, n = 5 mice, Galunisertib + αGal-9). b Representative bioluminescent and macroscopic image of B16F10-bearing C57BL/6 J mice for indicated groups describe in ( a ). c Quantification of bioluminescence and LBR. d Survival curve of tumor-bearing mice for the indicated groups described in ( b ). Log-rank test for survival comparison. e Representative flow cytometry plots depicting the percentages of Tex subsets based on TCF1and CX3CR1. f Statistical analysis of ( e ). g Representative flow cytometry plots depicting the percentages of Tex subsets based on TOX/CX3CR1. h Statistical analysis of ( g ). i Representative flow cytometry plots based depicting the percentages of Tex subsets on Ki-67 and TCF1. j Statistical analysis of ( i ). k–m Representative flow cytometry plots ( k ) and statistical analysis ( l , m ) depicting the TNF and IFNγ co-production by CD8 + T cells upon ex vivo CD3/CD28 stimulation for the indicated groups. Data are presented as the mean ± SD, and were analyzed with one-way ( c , l , m ) or two-way ( f , h , g ) ANOVA with Sidak’s multiple comparison test. Consistent results were observed across three biological replicates in ( b–m ). Source data are provided as a Source Data file.

Article Snippet: The above findings prompted us to explore whether therapeutic interventions, specifically Galectin-9 neutralization (αGal-9) or TβRI inhibition (Galunisertib) , would similarly alleviate liver metastasis by reshaping CD8 + T cell responses in our mouse model.

Techniques: Comparison, Flow Cytometry, Ex Vivo

Journal: Nature Communications

Article Title: Targeting hepatocytic TβRI ameliorates liver metastatic outcomes by revitalizing stem-like CD8 + Tex subsets

doi: 10.1038/s41467-025-65615-0

Figure Lengend Snippet: a IHC staining with TβRI-specific antibody to detect the distribution of TβRI + cells in colorectal liver metastases ( n = 36 patients). Representative IHC image. Scale bar, 50 μm. The bar chart in the right represents the percentage of high and low TβRI expression cases in liver metastasis, adjacent and distal liver tissues. p < 0.001 by two-sided Pearson’s chi-square test. b Overall survival curves based on adjacent liver TβRI expressed level in CRLM patients using the multivariate COX regression analysis, a two-sided P -value of < 0.05 was considered statistically significant. c Representative images depicting multicolor immunohistochemical co-staining of TβRI with specific cell type markers in CRLM adjacent liver tissues (DAPI: cell nucleus, HNF4α: hepatocytes, α-SMA: hepatic stellate cells, CD68: macrophages, PAN-CK: tumor cells). Scale bar, 25 μm. d Experimental scheme for generating TβRI flox/flox Alb-Cre +/- ( TβRI-hKO ) mice. TβRI flox/flox Alb-Cre -/- and C57BL/6 J were used as TβRI-hWT mice. e Representative bioluminescent image and Quantification of TβRI-hWT versus TβRI-hKO mice inoculated with MC38-Luc cells ( n = 6 mice/group). f Representative macroscopic image of ( e ) and Quantification of Liver to body weight ratio (LBR). g Representative bioluminescent image of TβRI-hWT versus TβRI-hKO mice inoculated with PAN02-Luc cells ( n = 5 mice/group). h Quantification of ( g ). i Representative macroscopic image of ( g ) and Quantification of LBR. j Representative bioluminescent image of TβRI-hWT versus TβRI-hKO mice inoculated with B16F10-Luc cells ( n = 5 mice/group). k Quantification of ( j ). l Representative macroscopic image of ( j ) and Quantification of LBR. Data are presented as the mean ± SD, and were analyzed with an unpaired two-sided Student’s t test ( e–l ). Consistent results were observed across three biological replicates in ( e–l ). Source data are provided as a Source Data file.

Article Snippet: TβRI flox/flox (Cat JAX: 028701), Alb-Cre +/+ (Cat JAX: 003574) mice were obtained from the Jackson Laboratory (USA).

Techniques: Immunohistochemistry, Expressing, Immunohistochemical staining, Staining

Journal: Nature Communications

Article Title: Targeting hepatocytic TβRI ameliorates liver metastatic outcomes by revitalizing stem-like CD8 + Tex subsets

doi: 10.1038/s41467-025-65615-0

Figure Lengend Snippet: a , b Frequency of T cell subsets based on the expression of Ly108 and CX3CR1 within CD8 + PD1 + CD44 + tumor-reactive T cells in different anatomical locations of TβRI-hWT and TβRI-hKO mice ( n = 6 mice/group). Flow cytometry representative plots ( a ). Statistical analysis of these subsets in liver metastases across multiple tumor types are shown ( b ). Statistical analysis of these subsets in the spleen and PBMC are not shown. c–f Flow cytometry was used to detect the phenotypes, transcriptional profiles, and effector functions of Ly108 + CX3CR1 - , Ly108 + CX3CR1 + , Ly108 - CX3CR1 + , Ly108 - CX3CR1 - subsets in liver metastases of TβRI-hWT and TβRI-hKO mice inoculated with PAN02 on day 28. c Representative histograms showing the expression of PD-1, LAG3, TIGIT, TIM3, and 2B4 among these T subsets ( n = 6 mice/group). MFI analysis of TIM3 and 2B4 was summarized in the right statistical chart. d Representative flow cytometry plots and statistical analysis depicting the TOX/TCF1 co-expression among these T subsets ( n = 4 mice/group). e Representative flow cytometry plots and statistical analysis depicting the TNF and IFNγ co-production by these T subsets upon ex vivo CD3/CD28 stimulation ( n = 5 mice/group). f Representative flow cytometry plots and statistical analysis depicting the percentages of these T subsets degranulating (CD107a) and producing IL2 upon ex vivo CD3/CD28 stimulation ( n = 5 mice/group). Data are presented as the mean ± SD. Data were analyzed with unpaired two-sided Student’s t test ( b ) and two-way ANOVA with Tukey’s multiple comparisons test ( c–f ). Consistent results were observed across three biological replicates in ( a–f ). Source data are provided as a Source Data file.

Article Snippet: TβRI flox/flox (Cat JAX: 028701), Alb-Cre +/+ (Cat JAX: 003574) mice were obtained from the Jackson Laboratory (USA).

Techniques: Expressing, Flow Cytometry, Ex Vivo

Journal: Nature Communications

Article Title: Targeting hepatocytic TβRI ameliorates liver metastatic outcomes by revitalizing stem-like CD8 + Tex subsets

doi: 10.1038/s41467-025-65615-0

Figure Lengend Snippet: a Experimental design. b Representative bioluminescent image of pre-adoptive and post-adoptive C57BL/6 J mice receiving Ly108 + CX3CR1 - , Ly108 + CX3CR1 + , Ly108 - CX3CR1 + , Ly108 - CX3CR1 - cells transfer or without transfer ( n = 5 mice/group). c Quantification of ( b ). d Survival curve of tumor-bearing mice for indicated groups described in ( b ). Log-rank test for survival comparison. e Flow cytometry depicted the phenotype of adoptively transferred cells before and 14 days after transfer. All subsets shown were gated on CD8 + PD1 + CD44 + CD45.2 + CD45.1 - cells. f Representative bioluminescent image of TβRI-hWT and TβRI-hKO mice inoculated with TGFβ1 OE , TGFβ1 KD , or TGFβ1 CTL PAN02-Luc cells ( n = 6 mice/group). g Quantification of ( f ). Data are presented as the mean ± SD, and were analyzed with one-way ( g ) or two-way ( c ) ANOVA with Tukey’s multiple comparison test. Consistent results were observed across three biological replicates in ( b–g ). Source data are provided as a Source Data file.

Article Snippet: TβRI flox/flox (Cat JAX: 028701), Alb-Cre +/+ (Cat JAX: 003574) mice were obtained from the Jackson Laboratory (USA).

Techniques: Comparison, Flow Cytometry

Journal: Nature Communications

Article Title: Targeting hepatocytic TβRI ameliorates liver metastatic outcomes by revitalizing stem-like CD8 + Tex subsets

doi: 10.1038/s41467-025-65615-0

Figure Lengend Snippet: a Experimental setup to assess the indispensable role of TAMs or CD8 + T cells in improving metastatic outcomes of PAN02-bearing mice resulting from hepatocytic-TβRI ablation. b The M1-to-M2 ratio. c Representative bioluminescent image of TβRI-hWT and TβRI-hKO mice treated with vehicle (veh), Clodronate (CLD) liposome or αCD8 (n = 5 mice/group). d Survival curve of tumor-bearing mice for the indicated groups described in ( c ). Log-rank test for survival comparison. e Quantification of ( c ). f Representative flow cytometry plots and Statistical analysis depicting the percentages of Tex subsets in liver metastases of veh or CLD liposome-treated TβRI-hWT and TβRI-hKO mice on days 28. g Representative bioluminescent image and liver macroscopic image of TβRI-hWT and TβRI-hKO mice inoculated with PAN02 + BMDM or PAN02 alone ( n = 5 mice/group). h Quantification of ( g ). i Statistical analysis depicting the percentages of Tex subsets about the indicated groups. Data are presented as the mean ± SD, and were analyzed with one-way ( h ) or two-way ( b , e , f , i ) ANOVA with Tukey’s multiple comparison test. Consistent results were observed across three biological replicates in ( b–i ). Source data are provided as a Source Data file.

Article Snippet: TβRI flox/flox (Cat JAX: 028701), Alb-Cre +/+ (Cat JAX: 003574) mice were obtained from the Jackson Laboratory (USA).

Techniques: Comparison, Flow Cytometry

Journal: Nature Communications

Article Title: Targeting hepatocytic TβRI ameliorates liver metastatic outcomes by revitalizing stem-like CD8 + Tex subsets

doi: 10.1038/s41467-025-65615-0

Figure Lengend Snippet: a Luminex showing the protein levels of inflammatory cytokines in tumor interstitial fluid of TβRI-hWT and TβRI-hKO mice ( n = 3 mice/group). Data are shown as a heatmap. b Experimental scheme for generating Gal-9 flox/flox Alb-Cre +/- (Gal9-hKO) mice. Gal-9 flox/flox Alb-Cre -/- and C57BL/6 J were used as Gal9-hWT mice. c Representative bioluminescent image of Gal9-hWT versus Gal9-hKO mice inoculated with TGFβ1 OE or TGFβ1 CTL -PAN02 cells in the portal vein on day 28 ( n = 6 mice/group). d Bioluminescence quantification of ( c ). e Statistical analysis depicting the percentages of CD39 + TOX + cells within CD8 + PD1 + CD44 + tumor-reactive T cells in the indicated groups. f Statistical analysis depicting the percentages of Tex subsets based on TCF1 and CX3CR1 within CD8 + PD1 + CD44 + tumor-reactive T cells. g statistical analysis depicting the TNF and IFNγ co-production by CD8 + CD44 + T subsets in the indicated groups upon ex vivo CD3/CD28 stimulation. h The diagram of experimental design. i Representative bioluminescent image of Gal9-hWT versus Gal9-hKO mice treated with IgG, αCD4, αCD8 or αPD-1( n = 5 mice/group). j Representative macroscopic image of ( i ). k Bioluminescence quantification of ( i ). l Statistical analysis depicting the percentages of Tex subsets based on Ly108 and CX3CR1 within CD8 + PD1 + CD44 + tumor-reactive T cells. Data are presented as the mean ± SD, and were analyzed with unpaired two-sided Student’s t test ( k ) or two-way ( d , e , f , g , l ) ANOVA with Tukey’s multiple comparison test. Consistent results were observed across three biological replicates in ( a , c–g , i–l ). Source data are provided as a Source Data file.

Article Snippet: TβRI flox/flox (Cat JAX: 028701), Alb-Cre +/+ (Cat JAX: 003574) mice were obtained from the Jackson Laboratory (USA).

Techniques: Luminex, Ex Vivo, Comparison

Journal: bioRxiv

Article Title: EPHB2 promotes diet-induced MASH liver fibrosis

doi: 10.1101/2025.11.19.688968

Figure Lengend Snippet: (A) Dot plot showing Ephb2 gene expression across different hepatic cell types in mouse snRNA-seq analyzed from published dataset GSE212837. (B) Representative immunofluorescence images of human liver sections of healthy and MASH patients stained with antibodies against phosphoEphB2 (Red) and αSMA (green), DAPI counterstain for nuclei is shown in blue. (C) Messenger RNA level of EPHB2 , COL1A2 , and ACTA2 in TGF-β1 stimulated primary human HSCs at various time point (n=3). (D) Western blot showing proteins level of EphB2, aSMA, pSmad2/3 Smad2/3 and Vinculin in TGF-β1 stimulated primary human HSCs at various time point. (E) Messenger RNA level of EPHB2 upon inhibition of canonical TGF-β/SMAD with SIS3 Smad3 inhibitor, SD208 TGFβR1 inhibitor, and SB525334 TGFβR2 inhibitor in TGF-β1-treated primary human HSCs (n=5-7). (F) Effects of silencing Smad2 and Smad3 with Mission®-esiRNA on TGF-β-induced EPHB2 mRNA expression in primary human HSCs (n=3). Non-targeting Mission®-esiRNA was used as control. (G) Accessibility peaks for the transcription factor SMAD3 on the promoter of human EPHB2 locus identified from a publicly available ChIP-seq dataset of TGF-β1 stimulated LX2 cells (GSE38103). (H) Enrichment of SMAD3 binding sites on the promoter of human EPHB2 validated by ChIP-qPCR in TGF-β1 stimulated primary human HSCs. All these data presented as mean ± SEM, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 by ANOVA ns = not significant.

Article Snippet: To assess the effect of TGF-β/SMAD canonical pathway on EphB2 expression, HSCs were incubated with the TGF-β type I receptor (TβRI) inhibitors SD208 (1μM), SB525334 (200nM), and the SMAD3 inhibitor SIS3 (5μM) (all from Tocris) for 4h before stimulation with recombinant human TGF-β1 (10ng/ml) for 48h.

Techniques: Gene Expression, Immunofluorescence, Staining, Western Blot, Inhibition, esiRNA, Expressing, Control, ChIP-sequencing, Binding Assay, ChIP-qPCR