Hyuk Moon1, Hye-Lim Ju2, Sook In Chung2, Kyung Joo Cho1, Jung Woo Eun3, Suk Woo Nam3, Kwang-Hyub Han4, Diego F Calvisi5, Simon Weonsang Ro6. 1. Institute of Gastroenterology, Yonsei University College of Medicine, Seoul, South Korea; Brain Korea 21 Project for Medical Science College of Medicine, Yonsei University, Seoul, South Korea. 2. Institute of Gastroenterology, Yonsei University College of Medicine, Seoul, South Korea. 3. Department of Pathology, College of Medicine, The Catholic University of Korea, Seoul, South Korea. 4. Department of Internal Medicine, Yonsei University College of Medicine, Seoul, South Korea. 5. Institute of Pathology, University Medicine Greifswald, Greifswald, Germany. 6. Institute of Gastroenterology, Yonsei University College of Medicine, Seoul, South Korea. Electronic address: simonr@yuhs.ac.
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.
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 humantumors 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 humanhepatocellular carcinoma samples deposited in The Cancer Genome Atlas. A cell proliferation assay was performed using humanliver 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 humanhepatocellular carcinoma, Snail expression correlated with TGF-β activation. Ectopic expression of Snail increased cellular proliferation, whereas Snail knockdown led to reduced proliferation in humanhepatocellular 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.
Authors: Meng Xu; Jingxiao Wang; Zhong Xu; Rong Li; Pan Wang; Runze Shang; Antonio Cigliano; Silvia Ribback; Antonio Solinas; Giovanni Mario Pes; Katja Evert; Haichuan Wang; Xinhua Song; Shu Zhang; Li Che; Rosa Maria Pascale; Diego Francesco Calvisi; Qingguang Liu; Xin Chen Journal: Cancer Res Date: 2019-08-05 Impact factor: 12.701
Authors: Haichuan Wang; Xinhua Song; Haotian Liao; Pan Wang; Yi Zhang; Li Che; Jie Zhang; Yi Zhou; Antonio Cigliano; Cindy Ament; Daphne Superville; Silvia Ribback; Melissa Reeves; Giovanni M Pes; Binyong Liang; Hong Wu; Matthias Evert; Diego F Calvisi; Yong Zeng; Xin Chen Journal: Hepatology Date: 2021-06-15 Impact factor: 17.298