New insights into the crosstalk between the TGF-β and the EGF Receptor pathways during liver regeneration and hepatocarcinogenesis
[eng] Transforming growth factor-beta (TGF-β) is an important growth suppressor in hepatocytes, inhibiting proliferation and inducing cell death. However, it also plays a role in other processes such as epithelial-to-mesenchymal transition (EMT), which contributes to tumour invasion and progression....
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| Tipo de recurso: | tesis doctoral |
| Estado: | Versión publicada |
| Fecha de publicación: | 2016 |
| País: | España |
| Institución: | Universidad de Barcelona |
| Repositorio: | Dipòsit Digital de la UB |
| OAI Identifier: | oai:diposit.ub.edu:2445/123749 |
| Acceso en línea: | https://hdl.handle.net/2445/123749 http://hdl.handle.net/10803/586393 |
| Access Level: | acceso abierto |
| Palabra clave: | Carcinogènesi Hepatologia Regeneració (Biologia) Migració cel·lular Factors de creixement Carcinogenesis Hepatology Regeneration (Biology) Cell migration Growth factors |
| Sumario: | [eng] Transforming growth factor-beta (TGF-β) is an important growth suppressor in hepatocytes, inhibiting proliferation and inducing cell death. However, it also plays a role in other processes such as epithelial-to-mesenchymal transition (EMT), which contributes to tumour invasion and progression. Epidermal Growth Factor Receptor (EGFR) belongs to the kinase receptor family, and it is involved in cell proliferation, survival, differentiation and adhesion. Aiming to study the connection between both pathways, different animal and cellular models have been generated in our group. The in vivo model is based on transgenic mice that express specifically in hepatocytes a truncated form (in the kinase domain) of the human EGFR, acting as a dominant negative, decreasing the signalling of the pathway. On the other hand, it has been generated an in vitro model, decreasing the expression of EGFR by shRNA in different hepatocellular carcinoma (HCC) lines. This thesis focused, firstly, in the study of the crosstalk between both pathways during liver regeneration and HCC in the in vivo model. On the other hand, in the in vitro model, we analyzed this crosstalk related to the protumorigenic effects, such as adhesion and migration of tumour cells. Finally, we have evaluated the translational relevance of the results obtained, analyzing the gene expression in tissues from HCC patients. The results obtained in the in vivo model show that mice expressing the truncated form of EGFR in the hepatocytes present a delay in the process of liver regeneration after partial hepatectomy (PH) and in the appearance of tumours induced after injecting the carcinogen Diethylnitrosamine (DEN). Lack of EGFR catalytic activity promotes the overactivation of the TGF-β pathway during liver regeneration, correlating with the induction of the expression of cell cycle inhibitors, such as Cdkn1a (p21) and Cdkn2b (p15). EGFR pathway signalling is also required for the accumulation of lipid droplets and for the expression of a specific adipogenic profile during liver regeneration, suggesting a role for EGFR in the lipid metabolism in the liver. However, despite all these alterations, life of transgenic mice is not significantly compromised, and animals are able to fully regenerate the liver. Overactivation of the hepatocyte growth factor (HGF)/Met pathway and hyperplasia of the remaining cells could contribute to the success in the regenerative process. On the other hand, during DEN-induced tumorigenesis, delayed onset of tumours when EGFR catalytic activity is attenuated does not correlate with an increased Tgfb1 expression or overactivation of the Smads pathway. However, significant higher expression of Nox4, target of TGF-β and an inhibitor of cell proliferation in the liver, is observed. In the in vitro model, it is observed that the effects of TGF-β on the EMT phenotype and on the migratory capacities of HCC cells are heterogeneous. Thus, Hep3B cell line responds to it inducing a full EMT, whereas PLC/PRF/5 cell line undergoes partial EMT, increasing mesenchymal markers, but maintaining E-cadherin expression and cell-cell adhesion. EGFR silencing decreases cell-cell adhesions in PLC/PRF/5 and Hep3B. However, it is only able to decrease adhesion to matrix in the case of PLC/PRF/5, but not in Hep3B. All these results point that EGFR silencing induces a TGF-β-induced ameboid migration in PLC/ PRF/5 (but not in Hep3B), that correlates with higher actomyosin contractility and advantages in cell migration. Finally, analysis in tumour tissues of patients with HCC reveals that most of them present an overactivation of the TGF-β pathway and lower levels of EGFR. However, there is no correlation between the expression of TGFB1 and EGFR, suggesting that they could be regulated by independent mechanisms. Finally, the EMT-related genetic profile in HCC patients expressing high levels of TGFB1 is associated with EGFR expression, so that in tumours with low levels of EGFR, low expression of CDH1 and high expression of VIM is observed. Taken together, our results indicate that the EGFR pathway plays essential roles in the proliferation of hepatocytes under physiological conditions such as liver regeneration, regulating, among other processes, the cellular response to the suppressive effects of TGF-β. During carcinogenesis, at early stages it may contribute to the appearance of preneoplastic nodules, but once the tumour has formed, EGFR pathway may counteract some of the pro-migratory and invasive TGF-β signals. Therefore, it is necessary to deep into the mechanisms of crosstalk between both pathways, in order to target them therapeutically. The stratification of patients according to their gene expression may help in the application of the most appropriate therapies. This thesis provides information about some biomarkers that could be used for this purpose. |
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