Transforming Growth Factor-β Promotes Liver Tumorigenesis in Mice via Up-regulation of Snail

Hyuk Moon, Hye Lim Ju, Sook In Chung, Kyung Joo Cho, Jung Woo Eun, Suk Woo Nam, Kwang Hyub Han, Diego F. Calvisi, Simon Weonsang Ro

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

Background & Aims Transforming growth factor beta (TGF-β) suppresses early stages of tumorigenesis, but also contributes to migration and metastasis of cancer cells. A large number of human tumors contain mutations that inactivate its receptors, or downstream proteins such as Smad transcription factors, indicating that the TGF-β signaling pathway prevents tumor growth. We investigated the effects of TGF-β inhibition on liver tumorigenesis in mice. Methods C57BL/6 mice received hydrodynamic tail-vein injections of transposons encoding HRASG12V and a short hairpin RNA (shRNA) to down-regulate p53, or those encoding HRASG12V and MYC, or those encoding HRASG12V and TAZS89A, to induce liver tumor formation; mice were also given injections of transposons encoding SMAD7 or shRNA against SMAD2, SMAD3, SMAD4, or SNAI1 (Snail), with or without ectopic expression of Snail. Survival times were compared, and livers were weighted and examined for tumors. Liver tumor tissues were analyzed by quantitative reverse-transcription PCR, RNA sequencing, immunoblots, and immunohistochemistry. We analyzed gene expression levels in human hepatocellular carcinoma samples deposited in The Cancer Genome Atlas. A cell proliferation assay was performed using human liver cancer cell lines (HepG2 and Huh7) stably expressing Snail or shRNA against Snail. Results TGF-β inhibition via overexpression of SMAD7 (or knockdown of SMAD2, SMAD3, or SMAD4) consistently reduced formation and growth of liver tumors in mice that expressed activated RAS plus shRNA against p53, or in mice that expressed activated RAS and TAZ. TGF-β signaling activated transcription of the Snail gene in liver tumors induced by HRASG12V and shRNA against p53, and by activated RAS and TAZ. Knockdown of Snail reduced liver tumor formation in both tumor models. Ectopic expression of Snail restored liver tumorigenesis suppressed by disruption of TGF-β signaling. In human hepatocellular carcinoma, Snail expression correlated with TGF-β activation. Ectopic expression of Snail increased cellular proliferation, whereas Snail knockdown led to reduced proliferation in human hepatocellular carcinoma cells. Conclusions In analyses of transgenic mice, we found TGF-β signaling to be required for formation of liver tumors upon expression of activated RAS and shRNA down-regulating p53, and upon expression of activated RAS and TAZ. Snail is the TGF-β target that is required for hepatic tumorigenesis in these models.

Original languageEnglish
Pages (from-to)1378-1391.e6
JournalGastroenterology
Volume153
Issue number5
DOIs
Publication statusPublished - 2017 Nov

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Transforming Growth Factors
Carcinogenesis
Up-Regulation
Transforming Growth Factor beta
Liver
Small Interfering RNA
Neoplasms
Snails
Hepatocellular Carcinoma
Cell Proliferation
RNA Sequence Analysis
Injections
Atlases
Hydrodynamics
Liver Neoplasms
Growth
Inbred C57BL Mouse
Transgenic Mice
Reverse Transcription
Tail

All Science Journal Classification (ASJC) codes

  • Hepatology
  • Gastroenterology

Cite this

Moon, H., Ju, H. L., Chung, S. I., Cho, K. J., Eun, J. W., Nam, S. W., ... Ro, S. W. (2017). Transforming Growth Factor-β Promotes Liver Tumorigenesis in Mice via Up-regulation of Snail. Gastroenterology, 153(5), 1378-1391.e6. https://doi.org/10.1053/j.gastro.2017.07.014
Moon, Hyuk ; Ju, Hye Lim ; Chung, Sook In ; Cho, Kyung Joo ; Eun, Jung Woo ; Nam, Suk Woo ; Han, Kwang Hyub ; Calvisi, Diego F. ; Ro, Simon Weonsang. / Transforming Growth Factor-β Promotes Liver Tumorigenesis in Mice via Up-regulation of Snail. In: Gastroenterology. 2017 ; Vol. 153, No. 5. pp. 1378-1391.e6.
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abstract = "Background & Aims Transforming growth factor beta (TGF-β) suppresses early stages of tumorigenesis, but also contributes to migration and metastasis of cancer cells. A large number of human tumors contain mutations that inactivate its receptors, or downstream proteins such as Smad transcription factors, indicating that the TGF-β signaling pathway prevents tumor growth. We investigated the effects of TGF-β inhibition on liver tumorigenesis in mice. Methods C57BL/6 mice received hydrodynamic tail-vein injections of transposons encoding HRASG12V and a short hairpin RNA (shRNA) to down-regulate p53, or those encoding HRASG12V and MYC, or those encoding HRASG12V and TAZS89A, to induce liver tumor formation; mice were also given injections of transposons encoding SMAD7 or shRNA against SMAD2, SMAD3, SMAD4, or SNAI1 (Snail), with or without ectopic expression of Snail. Survival times were compared, and livers were weighted and examined for tumors. Liver tumor tissues were analyzed by quantitative reverse-transcription PCR, RNA sequencing, immunoblots, and immunohistochemistry. We analyzed gene expression levels in human hepatocellular carcinoma samples deposited in The Cancer Genome Atlas. A cell proliferation assay was performed using human liver cancer cell lines (HepG2 and Huh7) stably expressing Snail or shRNA against Snail. Results TGF-β inhibition via overexpression of SMAD7 (or knockdown of SMAD2, SMAD3, or SMAD4) consistently reduced formation and growth of liver tumors in mice that expressed activated RAS plus shRNA against p53, or in mice that expressed activated RAS and TAZ. TGF-β signaling activated transcription of the Snail gene in liver tumors induced by HRASG12V and shRNA against p53, and by activated RAS and TAZ. Knockdown of Snail reduced liver tumor formation in both tumor models. Ectopic expression of Snail restored liver tumorigenesis suppressed by disruption of TGF-β signaling. In human hepatocellular carcinoma, Snail expression correlated with TGF-β activation. Ectopic expression of Snail increased cellular proliferation, whereas Snail knockdown led to reduced proliferation in human hepatocellular carcinoma cells. Conclusions In analyses of transgenic mice, we found TGF-β signaling to be required for formation of liver tumors upon expression of activated RAS and shRNA down-regulating p53, and upon expression of activated RAS and TAZ. Snail is the TGF-β target that is required for hepatic tumorigenesis in these models.",
author = "Hyuk Moon and Ju, {Hye Lim} and Chung, {Sook In} and Cho, {Kyung Joo} and Eun, {Jung Woo} and Nam, {Suk Woo} and Han, {Kwang Hyub} and Calvisi, {Diego F.} and Ro, {Simon Weonsang}",
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Moon, H, Ju, HL, Chung, SI, Cho, KJ, Eun, JW, Nam, SW, Han, KH, Calvisi, DF & Ro, SW 2017, 'Transforming Growth Factor-β Promotes Liver Tumorigenesis in Mice via Up-regulation of Snail', Gastroenterology, vol. 153, no. 5, pp. 1378-1391.e6. https://doi.org/10.1053/j.gastro.2017.07.014

Transforming Growth Factor-β Promotes Liver Tumorigenesis in Mice via Up-regulation of Snail. / Moon, Hyuk; Ju, Hye Lim; Chung, Sook In; Cho, Kyung Joo; Eun, Jung Woo; Nam, Suk Woo; Han, Kwang Hyub; Calvisi, Diego F.; Ro, Simon Weonsang.

In: Gastroenterology, Vol. 153, No. 5, 11.2017, p. 1378-1391.e6.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Transforming Growth Factor-β Promotes Liver Tumorigenesis in Mice via Up-regulation of Snail

AU - Moon, Hyuk

AU - Ju, Hye Lim

AU - Chung, Sook In

AU - Cho, Kyung Joo

AU - Eun, Jung Woo

AU - Nam, Suk Woo

AU - Han, Kwang Hyub

AU - Calvisi, Diego F.

AU - Ro, Simon Weonsang

PY - 2017/11

Y1 - 2017/11

N2 - Background & Aims Transforming growth factor beta (TGF-β) suppresses early stages of tumorigenesis, but also contributes to migration and metastasis of cancer cells. A large number of human tumors contain mutations that inactivate its receptors, or downstream proteins such as Smad transcription factors, indicating that the TGF-β signaling pathway prevents tumor growth. We investigated the effects of TGF-β inhibition on liver tumorigenesis in mice. Methods C57BL/6 mice received hydrodynamic tail-vein injections of transposons encoding HRASG12V and a short hairpin RNA (shRNA) to down-regulate p53, or those encoding HRASG12V and MYC, or those encoding HRASG12V and TAZS89A, to induce liver tumor formation; mice were also given injections of transposons encoding SMAD7 or shRNA against SMAD2, SMAD3, SMAD4, or SNAI1 (Snail), with or without ectopic expression of Snail. Survival times were compared, and livers were weighted and examined for tumors. Liver tumor tissues were analyzed by quantitative reverse-transcription PCR, RNA sequencing, immunoblots, and immunohistochemistry. We analyzed gene expression levels in human hepatocellular carcinoma samples deposited in The Cancer Genome Atlas. A cell proliferation assay was performed using human liver cancer cell lines (HepG2 and Huh7) stably expressing Snail or shRNA against Snail. Results TGF-β inhibition via overexpression of SMAD7 (or knockdown of SMAD2, SMAD3, or SMAD4) consistently reduced formation and growth of liver tumors in mice that expressed activated RAS plus shRNA against p53, or in mice that expressed activated RAS and TAZ. TGF-β signaling activated transcription of the Snail gene in liver tumors induced by HRASG12V and shRNA against p53, and by activated RAS and TAZ. Knockdown of Snail reduced liver tumor formation in both tumor models. Ectopic expression of Snail restored liver tumorigenesis suppressed by disruption of TGF-β signaling. In human hepatocellular carcinoma, Snail expression correlated with TGF-β activation. Ectopic expression of Snail increased cellular proliferation, whereas Snail knockdown led to reduced proliferation in human hepatocellular carcinoma cells. Conclusions In analyses of transgenic mice, we found TGF-β signaling to be required for formation of liver tumors upon expression of activated RAS and shRNA down-regulating p53, and upon expression of activated RAS and TAZ. Snail is the TGF-β target that is required for hepatic tumorigenesis in these models.

AB - Background & Aims Transforming growth factor beta (TGF-β) suppresses early stages of tumorigenesis, but also contributes to migration and metastasis of cancer cells. A large number of human tumors contain mutations that inactivate its receptors, or downstream proteins such as Smad transcription factors, indicating that the TGF-β signaling pathway prevents tumor growth. We investigated the effects of TGF-β inhibition on liver tumorigenesis in mice. Methods C57BL/6 mice received hydrodynamic tail-vein injections of transposons encoding HRASG12V and a short hairpin RNA (shRNA) to down-regulate p53, or those encoding HRASG12V and MYC, or those encoding HRASG12V and TAZS89A, to induce liver tumor formation; mice were also given injections of transposons encoding SMAD7 or shRNA against SMAD2, SMAD3, SMAD4, or SNAI1 (Snail), with or without ectopic expression of Snail. Survival times were compared, and livers were weighted and examined for tumors. Liver tumor tissues were analyzed by quantitative reverse-transcription PCR, RNA sequencing, immunoblots, and immunohistochemistry. We analyzed gene expression levels in human hepatocellular carcinoma samples deposited in The Cancer Genome Atlas. A cell proliferation assay was performed using human liver cancer cell lines (HepG2 and Huh7) stably expressing Snail or shRNA against Snail. Results TGF-β inhibition via overexpression of SMAD7 (or knockdown of SMAD2, SMAD3, or SMAD4) consistently reduced formation and growth of liver tumors in mice that expressed activated RAS plus shRNA against p53, or in mice that expressed activated RAS and TAZ. TGF-β signaling activated transcription of the Snail gene in liver tumors induced by HRASG12V and shRNA against p53, and by activated RAS and TAZ. Knockdown of Snail reduced liver tumor formation in both tumor models. Ectopic expression of Snail restored liver tumorigenesis suppressed by disruption of TGF-β signaling. In human hepatocellular carcinoma, Snail expression correlated with TGF-β activation. Ectopic expression of Snail increased cellular proliferation, whereas Snail knockdown led to reduced proliferation in human hepatocellular carcinoma cells. Conclusions In analyses of transgenic mice, we found TGF-β signaling to be required for formation of liver tumors upon expression of activated RAS and shRNA down-regulating p53, and upon expression of activated RAS and TAZ. Snail is the TGF-β target that is required for hepatic tumorigenesis in these models.

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