Person: Sánchez Muñoz, Aranzazu
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First Name
Aranzazu
Last Name
Sánchez Muñoz
Affiliation
Universidad Complutense de Madrid
Faculty / Institute
Farmacia
Department
Bioquímica y Biología Molecular
Area
Bioquímica y Biología Molecular
Identifiers
14 results
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Item Lack of EGFR catalytic activity in hepatocytes improves liver regeneration following DDC‐induced cholestatic injury by promoting a pro‐restorative inflammatory response(Journal of Pathology, 2022) Lazcanoiturburu, Nerea; García‐Sáez, Juan; González‐Corralejo, Carlos; Roncero Romero, Cesáreo; Sanz Ortega, Julián; Martín‐Rodríguez, Carlos; Valdecantos, M Pilar; Martínez‐Palacián, Adoración; Almale Del Barrio, Laura; Bragado Domingo, Paloma; Calero‐Pérez, Silvia; Fernández, Almudena; García‐Bravo, María; Guerra, Carmen; Montoliu, Lluis; Segovia, José Carlos; Martínez Valverde, Ángela María; Fabregat Romero, María Isabel; Herrera González, Blanca María; Sánchez Muñoz, AranzazuDespite the well‐known hepatoprotective role of the epidermal growth factor receptor (EGFR) pathway upon acute damage, its specific actions during chronic liver disease, particularly cholestatic injury, remain ambiguous and unresolved. Here, we analyzed the consequences of inactivating EGFR signaling in the liver on the regenerative response following cholestatic injury. For that, transgenic mice overexpressing a dominant negative mutant human EGFR lacking tyrosine kinase activity (ΔEGFR) in albumin‐positive cells were submitted to liver damage induced by 3,5‐diethoxycarbonyl‐1,4‐dihydrocollidine (DDC), an experimental model resembling human primary sclerosing cholangitis. Our results show an early activation of EGFR after 1–2 days of a DDC‐supplemented diet, followed by a signaling switch‐off. Furthermore, ΔEGFR mice showed less liver damage and a more efficient regeneration following DDC injury. Analysis of the mechanisms driving this effect revealed an enhanced activation of mitogenic/survival signals, AKT and ERK1/2‐MAPKs, and changes in cell turnover consistent with a quicker resolution of damage in response to DDC. These changes were concomitant with profound differences in the profile of intrahepatic immune cells, consisting of a shift in the M1/M2 balance towards M2 polarity, and the Cd4/Cd8 ratio in favor of Cd4 lymphocytes, overall supporting an immune cell switch into a pro‐restorative phenotype. Interestingly, ΔEGFR livers also displayed an amplified ductular reaction, with increased expression of EPCAM and an increased number of CK19‐positive ductular structures in portal areas, demonstrating an overexpansion of ductular progenitor cells. In summary, our work supports the notion that hepatocyte‐specific EGFR activity acts as a key player in the crosstalk between parenchymal and non‐parenchymal hepatic cells, promoting the pro‐inflammatory response activated during cholestatic injury and therefore contributing to the pathogenesis of cholestatic liver disease. © 2022 The Pathological Society of Great Britain and Ireland.Item c-Met Signaling Is Essential for Mouse Adult Liver Progenitor Cells Expansion After Transforming Growth Factor-β-Induced Epithelial–Mesenchymal Transition and Regulates Cell Phenotypic Switch(Stem Cells, 2019) Almale Del Barrio, Laura; García-Álvaro, María; Martínez-Palacián, Adoración; García-Bravo, María; Lazcanoiturburu, Nerea; Addante, Annalisa; Roncero Romero, Cesáreo; Sanz Ortega, Julián; López, María de la O; Bragado Domingo, Paloma; Mikulits, Wolfgang; Factor, Valentina M.; Thorgeirsson, Snorri S.; Ignacio, Casal, J.; Segovia, José-Carlos; Rial, Eduardo; Fabregat Romero, María Isabel; Herrera González, Blanca María; Sánchez Muñoz, AranzazuAdult hepatic progenitor cells (HPCs)/oval cells are bipotential progenitors that participate in liver repair responses upon chronic injury. Recent findings highlight HPCs plasticity and importance of the HPCs niche signals to determine their fate during the regenerative process, favoring either fibrogenesis or damage resolution. Transforming growth factor-β (TGF-β) and hepatocyte growth factor (HGF) are among the key signals involved in liver regeneration and as component of HPCs niche regulates HPCs biology. Here, we characterize the TGF-β-triggered epithelial–mesenchymal transition (EMT) response in oval cells, its effects on cell fate in vivo, and the regulatory effect of the HGF/c-Met signaling. Our data show that chronic treatment with TGF-β triggers a partial EMT in oval cells based on coexpression of epithelial and mesenchymal markers. The phenotypic and functional profiling indicates that TGF-β-induced EMT is not associated with stemness but rather represents a step forward along hepatic lineage. This phenotypic transition confers advantageous traits to HPCs including survival, migratory/invasive and metabolic benefit, overall enhancing the regenerative potential of oval cells upon transplantation into a carbon tetrachloride-damaged liver. We further uncover a key contribution of the HGF/c-Met pathway to modulate the TGF-β-mediated EMT response. It allows oval cells expansion after EMT by controlling oxidative stress and apoptosis, likely via Twist regulation, and it counterbalances EMT by maintaining epithelial properties. Our work provides evidence that a coordinated and balanced action of TGF-β and HGF are critical for achievement of the optimal regenerative potential of HPCs, opening new therapeutic perspectives.Item Met signaling in cardiomyocytes is required for normal cardiac function in adult mice(Biochimica et Biophysica Acta (Molecular Basis of disease), 2013) Arechederra Calderón, María; Carmona Mejías, Rita; González-Nuñez, María; Gutiérrez Uzquiza, Álvaro; Bragado Domingo, Paloma; Cruz-González, Ignacio; Cano Rincón, Elena; Guerrero Arroyo, María Del Carmen; Sánchez Muñoz, Aranzazu; López-Novoa, José Miguel; Schneider, Michael D.; Maina, Flavio; Muñoz-Chápuli, Ramón; Porras Gallo, María AlmudenaHepatocyte growth factor (HGF) and its receptor, Met, are key determinants of distinct developmental processes. Although HGF exerts cardio-protective effects in a number of cardiac pathologies, it remains unknown whether HGF/Met signaling is essential for myocardial development and/or physiological function in adulthood. We therefore investigated the requirement of HGF/Met signaling in cardiomyocyte for embryonic and postnatal heart development and function by conditional inactivation of the Met receptor in cardiomyocytes using the Cre-α-MHC mouse line (referred to as α-MHCMet-KO). Although α-MHCMet-KO mice showed normal heart development and were viable and fertile, by 6 months of age, males developed cardiomyocyte hypertrophy, associated with interstitial fibrosis. A significant upregulation in markers of myocardial damage, such as β-MHC and ANF, was also observed. By the age of 9 months, α-MHCMet-KO males displayed systolic cardiac dysfunction. Mechanistically, we provide evidence of a severe imbalance in the antioxidant defenses in α-MHCMet-KO hearts involving a reduced expression and activity of catalase and superoxide dismutase, with consequent reactive oxygen species accumulation. Similar anomalies were observed in females, although with a slower kinetics. We also found that Met signaling down-regulation leads to an increase in TGF-β production and a decrease in p38MAPK activation, which may contribute to phenotypic alterations displayed in α-MHCMet-KO mice. Consistently, we show that HGF acts through p38α to upregulate antioxidant enzymes in cardiomyocytes. Our results highlight that HGF/Met signaling in cardiomyocytes plays a physiological cardio-protective role in adult mice by acting as an endogenous regulator of heart function through oxidative stress control.Item Dual role of protein tyrosine phosphatase 1B in the progression and reversion of non-alcoholic steatohepatitis(Molecular Metabolism, 2018) González-Rodríguez, Águeda; Valdecantos, M. Pilar; Rada, Patricia; Addante, Annalisa; Barahona, Inés; Rey, Esther; Pardo, Virginia; Ruiz, Laura; Laura M. Laiglesia; María J. Moreno-Aliaga; Carmelo García-Monzón; Sánchez Muñoz, Aranzazu; Martínez Valverde, Ángela MaríaObjectives: Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease in Western countries. Protein tyrosine phosphatase 1B (PTP1B), a negative modulator of insulin and cytokine signaling, is a therapeutic target for type 2 diabetes and obesity. We investigated the impact of PTP1B deficiency during NAFLD, particularly in non-alcoholic steatohepatitis (NASH). Methods: NASH features were evaluated in livers from wild-type (PTP1BWT) and PTP1B-deficient (PTP1BKO) mice fed methionine/cholinedeficient diet (MCD) for 8 weeks. A recovery model was established by replacing MCD to chow diet (CHD) for 2e7 days. Non-parenchymal liver cells (NPCs) were analyzed by flow cytometry. Oval cells markers were measured in human and mouse livers with NASH, and in oval cells from PTP1BWT and PTP1BKO mice. Results: PTP1BWT mice fed MCD for 8 weeks exhibited NASH, NPCs infiltration, and elevated Fgf21, Il6 and Il1b mRNAs. These parameters decreased after switching to CHD. PTP1B deficiency accelerated MCD-induced NASH. Conversely, after switching to CHD, PTP1BKO mice rapidly reverted NASH compared to PTP1BWT mice in parallel to the normalization of serum triglycerides (TG) levels. Among NPCs, a drop in cytotoxic natural killer T (NKT) subpopulation was detected in PTP1BKO livers during recovery, and in these conditions M2 macrophage markers were upregulated. Oval cells markers (EpCAM and cytokeratin 19) significantly increased during NASH only in PTP1B-deficient livers. HGF-mediated signaling and proliferative capacity were enhanced in PTP1BKO oval cells. In NASH patients, oval cells markers were also elevated. Conclusions: PTP1B elicits a dual role in NASH progression and reversion. Additionally, our results support a new role for PTP1B in oval cell proliferation during NAFLD.Item Bone morphogenetic protein 9 as a key regulator of liver progenitor cells in DDC‐induced cholestatic liver injury(Liver International, 2018) Addante, Annalisa; Roncero Romero, Cesáreo; Almale Del Barrio, Laura; Lazcanoiturburu, Nerea; García‐Álvaro, María; Fernández García De Castro, Margarita; Sanz Ortega, Julián; Hammad, Seddik; Nwosu, Zeribe C.; Lee, Se‐Jin; Fabregat Romero, María Isabel; Dooley, Steven; Dijke, Peter ten; Herrera González, Blanca María; Sánchez Muñoz, AranzazuBackground & Aims: Bone morphogenetic protein 9 (BMP9) interferes with liver regeneration upon acute injury, while promoting fibrosis upon carbon tetrachlorideinduced chronic injury. We have now addressed the role of BMP9 in 3,5 diethoxicarbonyl-1,4 dihydrocollidine (DDC)-induced cholestatic liver injury, a model of liver regeneration mediated by hepatic progenitor cell (known as oval cell), exemplified as ductular reaction and oval cell expansion. Methods: WT and BMP9KO mice were submitted to DDC diet. Livers were examined for liver injury, fibrosis, inflammation and oval cell expansion by serum biochemistry, histology, RT-qPCR and western blot. BMP9 signalling and effects in oval cells were studied in vitro using western blot and transcriptional assays, plus functional assays of DNA synthesis, cell viability and apoptosis. Crosslinking assays and short hairpin RNA approaches were used to identify the receptors mediating BMP9 effects. Results: Deletion of BMP9 reduces liver damage and fibrosis, but enhances inflammation upon DDC feeding. Molecularly, absence of BMP9 results in overactivation of PI3K/AKT, ERK-MAPKs and c-Met signalling pathways, which together with an enhanced ductular reaction and oval cell expansion evidence an improved regenerative response and decreased damage in response to DDC feeding. Importantly, BMP9 directly targets oval cells, it activates SMAD1,5,8, decreases cell growth and promotes apoptosis, effects that are mediated by Activin Receptor-Like Kinase 2 (ALK2) type I receptor. Conclusions: We identify BMP9 as a negative regulator of oval cell expansion in cholestatic injury, its deletion enhancing liver regeneration. Likewise, our work further supports BMP9 as an attractive therapeutic target for chronic liver diseases.Item Intestinal BMP-9 locally upregulates FGF19 and is down-regulated in obese patients with diabetes(Molecular and Cellular Endocrinology, 2023) Stephan Drexler; Cai, Chen; Hartmann, Anna-Lena; Moch, Denise; Gaitantzi, Haristi; Ney, Theresa; Kraemer, Malin; Chu, Yuan; Zheng, Yuwei; Rahbari, Mohammad; Treffs, Annalena; Reiser, Alena; Lenoir, Bénédicte; Valous, Nektarios A.; Jäger, Dirk; Birgin, Emrullah; Sawant, Tejas A.; Li, Qi; Xu, Keshu; Dong, Lingyue; Otto, Mirko; Itzel, Timo; Teufel, Andreas; Gretz, Norbert; Hawinkels, Lukas J.A.C.; Sánchez Muñoz, Aranzazu; Herrera González, Blanca María; Schubert, Rudolf; Moshage, Han; Reissfelder, Christoph; Ebert, Matthias P.A.; Rahbari, Nuh N.; Breitkopf-Heinlein, KatjaBone morphogenetic protein (BMP)-9, a member of the TGFβ-family of cytokines, is believed to be mainly produced in the liver. The serum levels of BMP-9 were reported to be reduced in newly diagnosed diabetic patients and BMP-9 overexpression ameliorated steatosis in the high fat diet-induced obesity mouse model. Furthermore, injection of BMP-9 in mice enhanced expression of fibroblast growth factor (FGF)21. However, whether BMP-9 also regulates the expression of the related FGF19 is not clear. Because both FGF21 and 19 were described to protect the liver from steatosis, we have further investigated the role of BMP-9 in this context. We first analyzed BMP-9 levels in the serum of streptozotocin (STZ)-induced diabetic rats (a model of type I diabetes) and confirmed that BMP-9 serum levels decrease during diabetes. Microarray analyses of RNA samples from hepatic and intestinal tissue from BMP-9 KO- and wild-type mice (C57/Bl6 background) pointed to basal expression of BMP-9 in both organs and revealed a down-regulation of hepatic Fgf21 and intestinal Fgf19 in the KO mice. Next, we analyzed BMP-9 levels in a cohort of obese patients with or without diabetes. Serum BMP-9 levels did not correlate with diabetes, but hepatic BMP-9 mRNA expression negatively correlated with steatosis in those patients that did not yet develop diabetes. Likewise, hepatic BMP-9 expression also negatively correlated with serum LPS levels. In situ hybridization analyses confirmed intestinal BMP-9 expression. Intestinal (but not hepatic) BMP-9 mRNA levels were decreased with diabetes and positively correlated with intestinal E-Cadherin expression. In vitro studies using organoids demonstrated that BMP-9 directly induces FGF19 in gut but not hepatocyte organoids, whereas no evidence of a direct induction of hepatic FGF21 by BMP-9 was found. Consistent with the in vitro data, a correlation between intestinal BMP-9 and FGF19 mRNA expression was seen in the patients’ samples. In summary, our data confirm that BMP-9 is involved in diabetes development in humans and in the control of the FGF-axis. More importantly, our data imply that not only hepatic but also intestinal BMP-9 associates with diabetes and steatosis development and controls FGF19 expression. The data support the conclusion that increased levels of BMP-9 would most likely be beneficial under pre-steatotic conditions, making supplementation of BMP-9 an interesting new approach for future therapies aiming at prevention of the development of a metabolic syndrome and liver steatosis.Item HGF/c-Met signaling promotes liver progenitor cell migration and invasion by an epithelial-mesenchymal transition-independent, phosphatidyl inositol-3 kinase-dependent pathway in an in vitro model(BBA - Molecular Cell Research, 2015) Suárez Causado, Amileth; Caballero Díaz, D.; Bertrán, E.; Roncero Romero, Cesáreo; Addante, Annalisa; García Álvaro, M.; Fernández García De Castro, Margarita; Herrera González, Blanca María; Porras Gallo, María Almudena; Fabregat, I.; Sánchez Muñoz, AranzazuOval cells constitute an interesting hepatic cell population. They contribute to sustain liver regeneration during chronic liver damage, but in doing this they can be target of malignant conversion and become tumor-initiating cells and drive hepatocarcinogenesis. The molecular mechanisms beneath either their pro-regenerative or pro-tumorigenic potential are still poorly understood. In this study, we have investigated the role of the HGF/c-Met pathway in regulation of oval cell migratory and invasive properties. Our results show that HGF induces c-Met-dependent oval cell migration both in normal culture conditions and after in vitro wounding. HGF-triggered migration involves Factin cytoskeleton reorganization, which is also evidenced by activation of Rac1. Furthermore, HGF causes ZO-1 translocation from cell-cell contact sites to cytoplasm and its concomitant activation by phosphorylation. However, no loss of expression of cell-cell adhesion proteins, including E-cadherin, ZO-1 and Occludin-1, is observed. Additionally, migration does not lead to cell dispersal but to a characteristic organized pattern in rows, in turn associated with Golgi compaction, providing strong evidence of a morphogenic collective migration. Besides migration, HGF increases oval cell invasion through extracellular matrix, process that requires PI3K activation and is at least partly mediated by expression and activation of metalloproteases. Altogether, our findings provide novel insights into the cellular and molecular mechanisms mediating the essential role of HGF/c-Met signaling during oval cell-mediated mouse liver regeneration.Item Case Report: An EGFR-Targeted 4-1BB-agonistic Trimerbody Does Not Induce Hepatotoxicity in Transgenic Mice With Liver Expression of Human EGFR(Frontiers in Immunology, 2021) Compte, Marta; Harwood, Seandean L.; Martínez-Torrecuadrada, Jorge; Pérez-Chacón, Gema; González-García, Patricia; Tapia-Galisteo, Antonio; Bergen en Henegouwen, Paul M. P. Van; Sánchez Muñoz, Aranzazu; Fabregat, Isabel; Sanz, Laura; Zapata, Juan M.; Álvarez-Vallina, Luis; Palazon, AsisAgonistic monoclonal antibodies (mAbs) targeting the co-stimulatory receptor 4-1BB are among the most effective immunotherapeutic agents across pre-clinical cancer models. However, clinical development of full-length 4-1BB agonistic mAbs, has been hampered by dose-limiting liver toxicity. We have previously developed an EGFR-targeted 4-1BBagonistic trimerbody (1D8N/CEGa1) that induces potent anti-tumor immunity without systemic toxicity, in immunocompetent mice bearing murine colorectal carcinoma cells expressing human EGFR. Here, we study the impact of human EGFR expression on mouse liver in the toxicity profile of 1D8N/CEGa1. Systemic administration of IgG-based anti-4-1BB agonist resulted in nonspecific immune stimulation and hepatotoxicity in a liver-specific human EGFR-transgenic immunocompetent mouse, whereas in 1D8N/CEGa1-treated mice no such immune-related adverse effects were observed. Collectively, these data support the role of FcgR interactions in the major offtumor toxicities associated with IgG-based 4-1BB agonists and further validate the safety profile of EGFR-targeted Fc-less 4-1BB-agonistic trimerbodies in systemic cancer immunotherapy protocols.Item Clathrin switches transforming growth factor-β role to pro-tumorigenic in liver cancer(Journal of Hepatology, 2020) Caballero-Díaz, Daniel; Bertran, Esther; Peñuelas-Haro, Irene; Moreno-Càceres, Joaquim; Malfettone, Andrea; López-Luque, Judit; Addante, Annalisa; Herrera González, Blanca María; Sánchez Muñoz, Aranzazu; Alay, Ania; Solé, Xavier; Serrano, Teresa; Ramos, Emilio; Fabregat Romero, María IsabelBackground & Aims: Upon ligand binding, tyrosine kinase receptors, such as epidermal growth factor receptor (EGFR), are recruited into clathrin-coated pits for internalization by endocytosis, which is relevant for signalling and/or receptor degradation. In liver cells, transforming growth factor-b (TGFb) induces both pro- and anti-apoptotic signals; the latter are mediated by the EGFR pathway. Since EGFR mainly traffics via clathrin-coated vesicles, we aimed to analyse the potential role of clathrin in TGF-b-induced signalling in liver cells and its relevance in liver cancer. Methods: Real-Time PCR and immunohistochemistry were used to analyse clathrin heavy-chain expression in human (CLTC) and mice (Cltc) liver tumours. Transient knockdown (siRNA) or overexpression of CLTC were used to analyse its role on TGF-b and EGFR signalling in vitro. Bioinformatic analysis was used to determine the effect of CLTC and TGFB1 expression on prognosis and overall survival in patients with hepatocellular carcinoma (HCC). Results: Clathrin expression increased during liver tumorigenesis in humans and mice. CLTC knockdown cells responded to TGF-b phosphorylating SMADs (canonical signalling) but showed impairment in the anti-apoptotic signals (EGFR transactivation). Experiments of loss or gain of function in HCC cells reveal an essential role for clathrin in inhibiting TGF-b-induced apoptosis and upregulation of its pro-apoptotic target NOX4. Autocrine TGF-b signalling in invasive HCC cells upregulates CLTC expression, switching its role to pro-tumorigenic. A positive correlation between TGFB1 and CLTC was found in HCC cells and patients. Patients expressing high levels of TGFB1 and CLTC had a worse prognosis and lower overall survival. Conclusions: This work describes a novel role for clathrin in liver tumorigenesis, favouring non-canonical pro-tumorigenic TGF-b pathways. CLTC expression in human HCC samples could help select patients that would benefit from TGF-b-targeted therapy. Lay summary: Clathrin heavy-chain expression increases during liver tumorigenesis in humans (CLTC) and mice (Cltc), altering the cellular response to TGF-b in favour of anti-apoptotic/ pro-tumorigenic signals. A positive correlation between TGFB1 and CLTC was found in HCC cells and patients. Patients expressing high levels of TGFB1 and CLTC had a worse prognosis and lower overall survival. CLTC expression in HCC human samples could help select patients that would benefit from therapies targeting TGF-b.Item New and Old Key Players in Liver Cancer(International Journal of Molecular Sciences, 2023) Cuesta Martínez, Ángel; Palao, Nerea; Bragado Domingo, Paloma; Gutiérrez Uzquiza, Álvaro; Herrera González, Blanca María; Sánchez Muñoz, Aranzazu; Porras Gallo, María Almudena; Arechederra, Maria; Tarantino, Giovanni; Berasain, CarmenLiver cancer represents a major health problem worldwide with growing incidence and high mortality, hepatocellular carcinoma (HCC) being the most frequent. Hepatocytes are likely the cellular origin of most HCCs through the accumulation of genetic alterations, although hepatic progenitor cells (HPCs) might also be candidates in specific cases, as discussed here. HCC usually develops in a context of chronic inflammation, fibrosis, and cirrhosis, although the role of fibrosis is controversial. The interplay between hepatocytes, immune cells and hepatic stellate cells is a key issue. This review summarizes critical aspects of the liver tumor microenvironment paying special attention to platelets as new key players, which exert both pro- and anti-tumor effects, determined by specific contexts and a tight regulation of platelet signaling. Additionally, the relevance of specific signaling pathways, mainly HGF/MET, EGFR and TGF-β is discussed. HGF and TGF-β are produced by different liver cells and platelets and regulate not only tumor cell fate but also HPCs, inflammation and fibrosis, these being key players in these processes. The role of C3G/RAPGEF1, required for the proper function of HGF/MET signaling in HCC and HPCs, is highlighted, due to its ability to promote HCC growth and, regulate HPC fate and platelet-mediated actions on liver cancer.