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Differential response of hepatocellular carcinoma glycolytic metabolism and oxidative stress markers after exposure to human amniotic membrane proteins

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Abstract

Background

The human Amniotic Membrane (hAM) has been studied as a potential therapeutic option in cancer, namely in hepatocellular carcinoma. Previously, our research group evaluated the effect of human Amniotic Membrane Protein Extracts (hAMPE) in cancer therapy, demonstrating that hAMPE inhibit the metabolic activity of human hepatocellular carcinoma cell lines: Hep3B2.1-7, HepG2 and Huh7. Therefore, and considering the close relationship between metabolic activity and oxidative stress, the aim of this study was to evaluate the effect of hAMPE treatment in glucose metabolism and its role in oxidative stress of hepatocellular carcinoma.

Methods and Results

Glucose uptake and lactate production was assessed by 1 H-NMR, and the expression of several mediators of the glycolytic pathway was evaluated by Western blot or fluorescence. Total antioxidant capacity (TAC) and biomarkers of oxidative stress effects in proteins were detected. Our results showed that hAMPE treatment increased glucose consumption on Hep3B2.1-7, HepG2, and Huh7 through the increase of GLUT1 in Hep3B2.1-7 and Huh7, and GLUT3 in HepG2 cells. It was observed an increased expression of 6-phosphofrutokinase (PFK-1L) in all cell lines though glucose was not converted to lactate on HepG2 and Huh7 cells, suggesting that hAMPE treatment may counteract the Warburg effect observed in carcinogenesis. In Hep3B2.1-7, hAMPE treatment induced an increase in expression of lactate dehydrogenase (LDH) and monocarboxylate transporter isoform 4 (MCT4). We further detected that hAMPE enhances the TAC of culture media after 2 and 8 h. This was followed by a degree of protection against proteins nitration and carbonylation.

Conclusions

Overall, this work highlights the potential usefulness of hAMPE as anticancer therapy through the modulation of the glycolytic and oxidative profile in human hepatocellular carcinoma.

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Acknowledgements

Sandra Moreira Rocha (SFRH/BD/115693/2016) and Ana Catarina Mamede (SFRH/BD/73649/2010) wishes to thank to FCT for their PhD grants. The authors would also like to thank Obstetrics Service of Coimbra Hospital and University Centre for the collection of human tissues used in this work.

Funding

This work was support by the Infarmed (Health Research found 2015) and National Funds by FCT-Foundation for Science and Technology (Project UID/Multi/00709/2019).

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    Authors and Affiliations

    1. Health Science Research Center (CICS-UBI), University of Beira Interior, Av. Infante D. Henrique, 6200-506, Covilhã, Portugal

      Andreia P Alves, Sandra M Rocha, Ana C Mamede & Cláudio J Maia

    2. Biophysics Unit, Faculty of Medicine, University of Coimbra, Coimbra, Portugal

      Andreia P Alves, Ana C Mamede & Filomena M Botelho

    3. Faculty of Medicine, CIMAGO, University of Coimbra, Coimbra, Portugal

      Ana C Mamede & Filomena M Botelho

    4. CNC.IBILI, University of Coimbra, Coimbra, Portugal

      Ana C Mamede & Filomena M Botelho

    5. Department of Anatomy, Unit for Multidisciplinary Research in Biomedicine (UMIB), Institute of Biomedical Sciences Abel Salazar (ICBAS), University of Porto, 4050-523, Porto, Portugal

      Patrícia C Braga & Marco G Alves

    6. Chemistry Department, University of Aveiro, Aveiro, Portugal

      Pedro F Oliveira

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    Contributions

    All authors participated in the conception and design of study. APA, SMR, ACM and CJM interpret and analyzed of the data; APA, SMR, ACM, PCB and MGA conducted the experiments; ACM, MGA, PFO, MFB and CJM critically read the article; and APA and SMR wrote the manuscript. APA and SMR contributed equally to this study.

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    Correspondence to Cláudio J Maia.

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    Andreia P Alves and Sandra M Rocha contributed equally to this work.

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    Alves, A.P., Rocha, S.M., Mamede, A.C. et al. Differential response of hepatocellular carcinoma glycolytic metabolism and oxidative stress markers after exposure to human amniotic membrane proteins. Mol Biol Rep 49, 7731–7741 (2022). https://doi.org/10.1007/s11033-022-07598-5

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