Antioxidant enzymes change in different non-metastatic stages in tumoral and peritumoral tissues of colorectal cancer
Graphical abstract
Introduction
Colorectal cancer (CRC) is the third most commonly diagnosed cancer in men and the second in women. Every year 1.1 million new CRC cases are detected, and 551,269 deaths occurred in 2018 (Bray et al., 2018). Mortality of this cancer has decreased due to an early detection through colonoscopy and screenings (Nishihara et al., 2013). However, there is a demanding need for specific biomarkers to detect CRC at early stages (Lee et al., 2018).
CRC can be classified into different stages attending to the extension of the tumour. Stage I is called cancer in situ, as the tumour has only invaded the submucosa; in stage II, the tumour has grown into other nearby tissues or organs, although it has not invaded any lymph node or blood vessels. Stage III tumours have grown through the wall of the colon and are attached to other nearby tissues or organs already spreading to lymph nodes and blood vessels. Finally, stage IV is known as metastatic (“Colorectal Cancer Stages”).
In the last decade, there is growing evidence that oxidative stress may influence CRC progression (Simone et al., 2011). Oxidative stress is a situation caused by an imbalance between Reactive Oxygen Species (ROS) production and antioxidant defences in cells. Both exogenous and endogenous factors can cause ROS production, but complex I and III of electron transport chain and NADPH oxidases are the main source of ROS (Barja de Quiroga, 1999; Guzy and Schumacker, 2006; Lambeth, 2004). These molecules are highly reactive and can cause DNA mutations, leading to genomic instability and epigenomic changes causing a downregulation of tumour suppressor genes and an upregulation of oncogenes (Ushijima, 2005; Wu and Ni, 2015). Furthermore, ROS can change redox status in cells and alter signalling pathways influencing the expression and/or the activity of different transcription factors. This way, ROS stimulate proliferation, invasion and protection from apoptosis of cancer cells (Galadari et al., 2017; Sainz et al., 2012). Nevertheless, too high levels of ROS can trigger apoptosis (Redza-Dutordoir and Averill-Bates, 2016; Ricci et al., 2008).
To counteract ROS effects, cells rely on antioxidant defences. SIRT3 (Sirtuin 3, SIRT3, SIR2L3, EC: 3.5.1.-) is a NAD+ dependent deacetylase involved in oxidative stress. SIRT3 induces a crucial antioxidant response, activating MnSOD (Manganese Superoxide Dismutase, SOD2, EC:1.15.1.1), and electron transport chain complexes by deacetylation (Ahn et al., 2008; Alhazzazi et al., 2013; Kincaid and Bossy-Wetzel, 2013; Ozden et al., 2011; Tao et al., 2010). MnSOD and CuZnSOD (Cupper Zinc Superoxide Dismutase, SOD1, EC: 1.15.1.1) detoxify superoxide anion into hydrogen peroxide. Superoxide anion is by far more reactive than hydrogen peroxide, but it has a very short half-life and it is not able to reach other spaces in the cell. Nevertheless, hydrogen peroxide has longer half-life and can reach other cell compartments. For these reasons, hydrogen peroxide can cause more damage in cells and its accumulation could cause serious consequences. GPx (Glutathione Peroxide, GPX EC: 1.11.1.9) and Catalase (Catalase, CAT, EC: 1.11.1.6) are antioxidant enzymes that detoxify hydrogen peroxide into molecular oxygen, decreasing oxidative stress (Winterbourn, 2013).
Recent evidence has determined that a tumour could present the behavior of a complete organ, which consist of stromal non-tumour cells apart from tumour cells which can communicate and influence each other promoting tumour progression and invasion (Egeblad et al., 2010). Furthermore, Sanz-Pamplona et al. and other authors have stablished that non-tumour adjacent tissue of tumour has an aberrant expression in comparison with normal tissues and may have some structural alterations (D’Alessio et al., 2019; Ganci et al., 2017; Huang et al., 2016; Sanz-Pamplona et al., 2014).
There are few studies that have compared redox status between the central part of tumour and the non-tumour adjacent tissue through different stages in some types of cancer, although the knowledge about this issue is limited (Cancer et al., 2001; Santandreu et al., 2008). Antioxidant proteins levels and their role in CRC in early stages have not been well stablished yet. For these reasons, our aim was to analyse antioxidant enzymes and proteins related to oxidative stress modulation in CRC biopsies both in tumour and non-tumour adjacent tissues of stages I, II, III, avoiding metastatic effects of stage IV, and determine whether these proteins could be potential biomarkers for non-metastatic stages in CRC.
Section snippets
Patients and tissue samples
This study was carried out on a group of 44 patients which were selected according to the following criteria: patients of both sexes without comorbidity in stage I, II and III of CRC were included. These patients had not been treated by radio- or chemotherapy before taking the samples: non-tumour adjacent and tumour tissue. Samples originating from Biobank of Hospital Son Llátzer were taken by surgery between the years 2005-2010. All patients signed an informed consent before surgery and this
Differences between tumour and non-tumour adjacent tissue of antioxidant enzymes levels
To determine differences between tumour and non-tumour adjacent tissue, the fold change ratio was calculated and is shown in Fig. 1. Fold change was calculated as log2 (tumour tissue/non-tumour adjacent tissue). Fold change representation showed that MnSOD and acMnSOD proteins were higher in tumour tissue in all stages compared to non-tumour adjacent tissue. CAT, GPx and GRd were higher expressed in non-tumour adjacent tissue compared to tumour tissue. CAT was higher in non-tumour adjacent
Discussion
We have shown that non-tumour adjacent tissue of CRC seemed to have higher levels of antioxidants enzymes that detoxify hydrogen peroxide in comparison to tumour tissue. In contrast, tumour tissue showed higher levels of MnSOD and acMnSOD than non-tumour adjacent tissue. Furthermore, changes in non-tumour adjacent tissue were found mainly between stage I and II in most of proteins analysed. Tumour tissue also showed a tendency to have higher levels of antioxidant enzymes in stage II compared to
Conclusions
Our results show that the role of antioxidant enzymes can not only take part in the advanced stages of CRC, but also, in the very early phases of the progression of cancer. Furthermore, non-tumour adjacent tissue also showed important changes in antioxidant proteins, suggesting an influence of the tumour tissue on it. For this reason, it may be important to analyse the expression of the antioxidant enzymes not only in tumours, but also in non-tumour adjacent tissue, with especial attention to
CRediT authorship contribution statement
Auba Gaya-Bover: Conceptualization, Methodology, Data curation, Writing - original draft. Reyniel Hernández-López: Conceptualization, Methodology. Marina Alorda-Clara: Conceptualization, Methodology. Javier M. Ibarra de la Rosa: Resources. Esther Falcó: Resources. Teresa Fernández: Resources. Maria Margarita Company: Resources. Margalida Torrens-Mas: Writing - review & editing. Pilar Roca: Data curation, Writing - original draft, Writing - review & editing. Jordi Oliver: Writing - review &
Declaration of Competing Interest
None declared.
Acknowledgments
This work was supported by grants from Fondo de Investigaciones Sanitarias of Instituto de Salud Carlos III [PI14/01434] of the Spanish Government confinanced by FEDER-Unión Europea (“Una manera de hacer Europa”). M. Torrens-Mas received an FPU grant [FPU14/07042] from Ministerio de Educación, Cultura y Deporte of Spanish Government.
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