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Farnesoid X receptor associates with β-catenin and inhibits its activity in hepatocellular carcinoma.

Liu X, Zhang X, Ji L, Gu J, Zhou M, Chen S - Oncotarget (2015)

Bottom Line: Activation of FXR attenuated the DNA-binding activity of β-Catenin/TCF4, and subsequently, its targeting gene-cyclin D1 expression.Importantly, FXR expression was markedly reduced in human HCC, an event which correlated with aberrant activation of β-Catenin.These data identified FXR as a negative regulator of HCC development through direct suppression of Wnt/β-catenin pathway.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Glycoconjugate Research Ministry of Public Health, Department of Biochemistry and Molecular Biology, Shanghai Medical college, Fudan University, Shanghai, China.

ABSTRACT
The association between the temporal activation of Wnt/β-catenin pathway and the spontaneous hepatocellular carcinoma (HCC) development in Farnesoid X receptor (FXR) knockout mice is not well understood. We found that Huh7 cells depleted with FXR by RNAi showed enhanced cell growth, migration and invasion in vitro and accelerated tumor xenografts formation in nude mice. And these phenotypes were attenuated by simultaneous knockdown of β-catenin with RNAi. Furthermore, we identified that FXR could bind with β-Catenin through AF1 domain, and disrupt the assembly of the core β-Catenin/TCF4 complex. Activation of FXR attenuated the DNA-binding activity of β-Catenin/TCF4, and subsequently, its targeting gene-cyclin D1 expression. Importantly, FXR expression was markedly reduced in human HCC, an event which correlated with aberrant activation of β-Catenin. These data identified FXR as a negative regulator of HCC development through direct suppression of Wnt/β-catenin pathway.

No MeSH data available.


Related in: MedlinePlus

Loss of FXR induced oncogenic behavior via Wnt/β-Catenin signaling in Huh7 cells(A) Expression of FXR, β-Catenin and active-β-Catenin in 9 hepatoma carcinoma cell lines. Western blot of FXR, β-Catenin and active-β-Catenin was performed using total cell lysates from 9 hepatocyte cell lines. β-Actin was used as a loading control. (B) Quantification of Western blot analysis (normalized by β-Actin). FXR expression level was negatively correlated with active-β-Catenin level in 9 hepatocyte cell lines. (C) Specific knockdown of FXR was confirmed by western blot in Huh7 cells. (D, E and F) Silencing FXR expression by FXR shRNA #2 in Huh7 cells promoted cell growth (D), migration (E) and invasion (F) as detected by MTT, Transwell migration and Matrigel invasion assays respectively. (G and H) Time course of xenograft growth in nude mice. Nude mice were subcutaneously injected with Huh7 cells infected with either control shRNA, or FXR shRNA#2, or FXR shRNA#2 in combination with β-Catenin shRNA. Tumor volume was measured and tumor mass was excised and imaged at indicated time after injection. Huh7/FXR shRNA cells displayed accelerated tumor growth. And this effect was attenuated by simultaneous knocking down β-Catenin expression. (I) Expression of FXR, Active-β-Catenin, CylinD1 and c-Myc in the excised tumors from nude mice was determined by Western blot. Error bars represent ± SEM from three independent samples. *, p < 0.01; **, p < 0.001.
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Figure 1: Loss of FXR induced oncogenic behavior via Wnt/β-Catenin signaling in Huh7 cells(A) Expression of FXR, β-Catenin and active-β-Catenin in 9 hepatoma carcinoma cell lines. Western blot of FXR, β-Catenin and active-β-Catenin was performed using total cell lysates from 9 hepatocyte cell lines. β-Actin was used as a loading control. (B) Quantification of Western blot analysis (normalized by β-Actin). FXR expression level was negatively correlated with active-β-Catenin level in 9 hepatocyte cell lines. (C) Specific knockdown of FXR was confirmed by western blot in Huh7 cells. (D, E and F) Silencing FXR expression by FXR shRNA #2 in Huh7 cells promoted cell growth (D), migration (E) and invasion (F) as detected by MTT, Transwell migration and Matrigel invasion assays respectively. (G and H) Time course of xenograft growth in nude mice. Nude mice were subcutaneously injected with Huh7 cells infected with either control shRNA, or FXR shRNA#2, or FXR shRNA#2 in combination with β-Catenin shRNA. Tumor volume was measured and tumor mass was excised and imaged at indicated time after injection. Huh7/FXR shRNA cells displayed accelerated tumor growth. And this effect was attenuated by simultaneous knocking down β-Catenin expression. (I) Expression of FXR, Active-β-Catenin, CylinD1 and c-Myc in the excised tumors from nude mice was determined by Western blot. Error bars represent ± SEM from three independent samples. *, p < 0.01; **, p < 0.001.

Mentions: Protein expression level of FXR, and β-Catenin was determined in nine different hepatocarcinoma cell lines. As shown in Figure 1A and 1B, FXR expression level is the highest in PLC-5, the lowest in MHCC-97L and median in Huh7 cell line. Interestingly, the expression profile of active-β-Catenin negatively correlated with FXR expression in these nine cell lines.


Farnesoid X receptor associates with β-catenin and inhibits its activity in hepatocellular carcinoma.

Liu X, Zhang X, Ji L, Gu J, Zhou M, Chen S - Oncotarget (2015)

Loss of FXR induced oncogenic behavior via Wnt/β-Catenin signaling in Huh7 cells(A) Expression of FXR, β-Catenin and active-β-Catenin in 9 hepatoma carcinoma cell lines. Western blot of FXR, β-Catenin and active-β-Catenin was performed using total cell lysates from 9 hepatocyte cell lines. β-Actin was used as a loading control. (B) Quantification of Western blot analysis (normalized by β-Actin). FXR expression level was negatively correlated with active-β-Catenin level in 9 hepatocyte cell lines. (C) Specific knockdown of FXR was confirmed by western blot in Huh7 cells. (D, E and F) Silencing FXR expression by FXR shRNA #2 in Huh7 cells promoted cell growth (D), migration (E) and invasion (F) as detected by MTT, Transwell migration and Matrigel invasion assays respectively. (G and H) Time course of xenograft growth in nude mice. Nude mice were subcutaneously injected with Huh7 cells infected with either control shRNA, or FXR shRNA#2, or FXR shRNA#2 in combination with β-Catenin shRNA. Tumor volume was measured and tumor mass was excised and imaged at indicated time after injection. Huh7/FXR shRNA cells displayed accelerated tumor growth. And this effect was attenuated by simultaneous knocking down β-Catenin expression. (I) Expression of FXR, Active-β-Catenin, CylinD1 and c-Myc in the excised tumors from nude mice was determined by Western blot. Error bars represent ± SEM from three independent samples. *, p < 0.01; **, p < 0.001.
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Related In: Results  -  Collection

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Figure 1: Loss of FXR induced oncogenic behavior via Wnt/β-Catenin signaling in Huh7 cells(A) Expression of FXR, β-Catenin and active-β-Catenin in 9 hepatoma carcinoma cell lines. Western blot of FXR, β-Catenin and active-β-Catenin was performed using total cell lysates from 9 hepatocyte cell lines. β-Actin was used as a loading control. (B) Quantification of Western blot analysis (normalized by β-Actin). FXR expression level was negatively correlated with active-β-Catenin level in 9 hepatocyte cell lines. (C) Specific knockdown of FXR was confirmed by western blot in Huh7 cells. (D, E and F) Silencing FXR expression by FXR shRNA #2 in Huh7 cells promoted cell growth (D), migration (E) and invasion (F) as detected by MTT, Transwell migration and Matrigel invasion assays respectively. (G and H) Time course of xenograft growth in nude mice. Nude mice were subcutaneously injected with Huh7 cells infected with either control shRNA, or FXR shRNA#2, or FXR shRNA#2 in combination with β-Catenin shRNA. Tumor volume was measured and tumor mass was excised and imaged at indicated time after injection. Huh7/FXR shRNA cells displayed accelerated tumor growth. And this effect was attenuated by simultaneous knocking down β-Catenin expression. (I) Expression of FXR, Active-β-Catenin, CylinD1 and c-Myc in the excised tumors from nude mice was determined by Western blot. Error bars represent ± SEM from three independent samples. *, p < 0.01; **, p < 0.001.
Mentions: Protein expression level of FXR, and β-Catenin was determined in nine different hepatocarcinoma cell lines. As shown in Figure 1A and 1B, FXR expression level is the highest in PLC-5, the lowest in MHCC-97L and median in Huh7 cell line. Interestingly, the expression profile of active-β-Catenin negatively correlated with FXR expression in these nine cell lines.

Bottom Line: Activation of FXR attenuated the DNA-binding activity of β-Catenin/TCF4, and subsequently, its targeting gene-cyclin D1 expression.Importantly, FXR expression was markedly reduced in human HCC, an event which correlated with aberrant activation of β-Catenin.These data identified FXR as a negative regulator of HCC development through direct suppression of Wnt/β-catenin pathway.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Glycoconjugate Research Ministry of Public Health, Department of Biochemistry and Molecular Biology, Shanghai Medical college, Fudan University, Shanghai, China.

ABSTRACT
The association between the temporal activation of Wnt/β-catenin pathway and the spontaneous hepatocellular carcinoma (HCC) development in Farnesoid X receptor (FXR) knockout mice is not well understood. We found that Huh7 cells depleted with FXR by RNAi showed enhanced cell growth, migration and invasion in vitro and accelerated tumor xenografts formation in nude mice. And these phenotypes were attenuated by simultaneous knockdown of β-catenin with RNAi. Furthermore, we identified that FXR could bind with β-Catenin through AF1 domain, and disrupt the assembly of the core β-Catenin/TCF4 complex. Activation of FXR attenuated the DNA-binding activity of β-Catenin/TCF4, and subsequently, its targeting gene-cyclin D1 expression. Importantly, FXR expression was markedly reduced in human HCC, an event which correlated with aberrant activation of β-Catenin. These data identified FXR as a negative regulator of HCC development through direct suppression of Wnt/β-catenin pathway.

No MeSH data available.


Related in: MedlinePlus