DOI: https://doi.org/10.7705/biomedica.v37i3.3239

Efecto mutagénico y genotóxico, y expresión de los genes Rad51C, Xiap, P53 y Nrf2 inducidos por extractos antipalúdicos de plantas recolectadas en el Vaupés medio, Colombia

Claudia Viviana Barbosa, Carlos Enrique Muskus, Luz Yaneth Orozco, Adriana Pabón

Resumen


Introducción. Dada la resistencia de Plasmodium a los medicamentos antipalúdicos, es necesario encontrar nuevas alternativas terapéuticas para su tratamiento y control. Con base en el saber indígena colombiano, se recopilaron extractos de plantas del Vaupés medio con potencial efecto antipalúdico.
Objetivo. Evaluar el efecto mutagénico y genotóxico, y la expresión de los genes Rad51C, Xiap, P53 y Nrf2, inducidos por cuatro extractos etanólicos con actividad anti-Plasmodium (R001, T002, T015 y T028).
Materiales y métodos. Se evaluó el potencial mutagénico de cuatro extractos etanólicos con efecto antiplasmódico utilizando el test de Ames y el efecto genotóxico, con un ensayo del cometa; asimismo, se analizó la expresión de los genes Rad51C, Xiap, P53 y Nrf2 en células HepG2.
Resultados. Los extractos no fueron mutágenos en la cepa TA98 de Salmonella typhimurium en presencia y ausencia de actividad metabólica de la fracción S9. En la cepa TA100, los extractos R001, T015 y T028 se comportaron como mutágenos débiles en presencia de S9, con índices mutagénicos de 1,58; 1,38; 1,53 y 1,61, respectivamente; T015 tuvo el mismo comportamiento en ausencia de S9, con un índice mutagénico de 1,36. En el ensayo del cometa, todos los extractos provocaron daño de categorías 1 o 2, con colas de cometas entre 36,7 y 51,48 μm de longitud; sin embargo, el índice de daño genético sugirió que los tratamientos afectaron la mayoría de las células. En los genes en estudio, los extractos R001 y T028 indujeron una sobreexpresión de 1,84 a 3,99 frente a las células sin tratar de los genes Xiap y P53.
Conclusiones. Los resultados evidenciaron que el extracto T002 fue el más seguro, ya que presentó actividad anti-Plasmodium, no fue citotóxico en las células HepG2, no fue mutágeno, causó daño de categoría 1 en el ADN y no modificó la expresión de los genes evaluados.


Palabras clave


malaria; resistencia a medicamentos; mutación; daño del ADN; apoptosis; estrés oxidativo

Texto completo:

PDF HTML

Referencias


1. World Health Organisation. World malaria report, 2014. Fecha de consulta: 14 de junio de 2016. Disponible en:
http://www.who.int/malaria/publications/world_malaria_report_2014/report/en/
2. Gligorijevic B, Purdy K, Elliott DA, Cooper RA, Roepe PD. Stage independent chloroquine resistance and chloroquine toxicity revealed via spinning disk confocal microscopy. Mol Biochem Parasitol. 2008;159:7-23. https://doi.org/10.1016/j.molbiopara.2007.12.014
3. Ramírez MO, Cardona NF, Pabón VA, Blair TS. Etnobotánica de las plantas antimaláricas del Vaupés Medio: recuperación del saber médico tradicional. Actu Biol. 2012;34:154.
4. Dike IP, Obembe OO, Adebiyi FE. Ethnobotanical survey for potential anti-malarial plants in south-western Nigeria. J Ethnopharmacol. 2012;144:618-26. https://doi.org/10.1016/j.jep.2012.10.002
5. Murambiwa P, Masola B, Govender T, Mukaratirwa S, Musabayane CT. Anti-malarial drug formulations and novel delivery systems: A review. Acta Trop. 2011;118:71-9. https://doi.org/10.1016/j.actatropica.2011.03.005
6. Oksman-Caldentey K-M, Inzé D. Plant cell factories in the post-genomic era: New ways to produce designer secondary metabolites. Trends Plant Sci. 2004;9:433-40. https://doi.org/10.1016/j.tplants.2004.07.006
7. Pabón A, Ramírez O, Ríos A, López E, de Las Salas B, Cardona F, et al. Antiplasmodial and cytotoxic activity of raw plant extracts as reported by knowledgeable indigenous people of the Amazon region (Vaupés Medio in Colombia). Planta Med. 2016;82:717-22. https://doi.org/10.1055/s-0042-104283
8. S. Flückiger-Isler MK. The Ames MPFTM 98/100 assay: Novel mutagenicity testing in liquid microplate format using S. typhimurium TA98 and TA100. Gewerbestrasse: Xenometrix; 2006.
9. Westerink WM, Stevenson JC, Horbach GJ, Schoonen WG. The development of RAD51C, Cystatin A, p53 and Nrf2 luciferase-reporter assays in metabolically competent HepG2 cells for the assessment of mechanism-based genotoxicity and of oxidative stress in the early research phase of drug development. Mutat Res. 2010;696:21-40. https://doi.org/10.1016/j.mrgentox.2009.12.007
10. García-Huertas P, Pabón A, Arias C, Blair S. Evaluación del efecto citotóxico y del daño genético de extractos estandarizados de Solanum nudum con actividad antiplasmodial. Biomédica. 2013;33:78-87. https://doi.org/10.7705/biomedica.v33i1.838
11. Mayence A, Vanden-Eynde JJ, Kaiser M, Brun R, Yarlett N, Huang TL. Bis(oxyphenylene)benzimidazoles: A novel class of anti-Plasmodium falciparum agents. Bioorg Med Chem. 2011;19:7493-500. https://doi.org/10.1016/j.bmc.2011.10.039
12. Maron DM, Ames BN. Revised methods for the Salmonella mutagenicity test. Mutat Res. 1983;113:173-215. https://doi.org/10.1016/0165-1161(83)90010-9
13. Flückiger-Isler S, Kamber M. Direct comparison of the Ames microplate format (MPF) test in liquid medium with the standard Ames pre-incubation assay on agar plates by use of equivocal to weakly positive test compounds. Mutat Res. 2012;747:36-45. https://doi.org/10.1016/j.mrgentox.2012.03.014
14. Platel A, Gervais V, Sajot N, Nesslany F, Marzin D, Claude N. Study of gene expression profiles in TK6 human cells exposed to DNA-oxidizing agents. Mutat Res. 2010;689:21-49. https://doi.org/10.1016/j.mrfmmm.2010.04.004
15. Singh NP, McCoy MT, Tice RR, Schneider EL. A simple technique for quantitation of low levels of DNA damage in individual cells. Exp Cell Res. 1988;175:184-91. https://doi.org/10.1016/0014-4827(88)90265-0
16. Zúñiga-Venegas LA, Creus-Capdevila A, Marcos-Dauder R. Optimizaciones metodológicas del ensayo del cometa y su aplicación en biomonitorización humana. Barccelona: Universitat Autònoma de Barcelona; 2009.
17. Guilherme S, Gaivão I, Santos MA, Pacheco M. European eel (Anguilla anguilla) genotoxic and pro-oxidant responses following short-term exposure to Roundup--a glyphosatebased herbicide. Mutagenesis. 2010;25:523-30. https://doi.org/10.1093/mutage/geq038
18. Rodrigues CD, Hannus M, Prudêncio M, Martin C, Gonçalves LA, Portugal S, et al. Host scavenger receptor SR-BI plays a dual role in the establishment of malaria parasite liver infection. Cell Host Microbe. 2008;4:271-82. https://doi.org/10.1016/j.chom.2008.07.012
19. Verma N, Pink M, Rettenmeier AW, Schmitz-Spanke S. Benzo[a]pyrene-mediated toxicity in primary pig bladder epithelial cells: A proteomic approach. J Proteomics. 2013;85:53-64. https://doi.org/10.1016/j.jprot.2013.04.016
20. Chen M, Gu H, Ye Y, Lin B, Sun L, Deng W, et al. Protective effects of hesperidin against oxidative stress of tert-butyl hydroperoxide in human hepatocytes. Food Chem Toxicol. 2010;48:2980-7. https://doi.org/10.1016/j.fct.2010.07.037
21. Minicucci EM, Ribeiro DA, da Silva GN, Pardini MI, Montovani JC, Salvadori DM.The role of the TP53 gene during rat tongue carcinogenesis induced by 4-nitroquinoline 1-oxide. Exp Toxicol Pathol. 2011;63:483-9. https://doi.org/10.1016/j.etp.2010.03.009
22. Amiri F, Zarnani A-H, Zand H, Koohdani F, Jeddi-Tehrani M, Vafa M. Synergistic anti-proliferative effect of resveratrol and etoposide on human hepatocellular and colon cancer cell lines. Eur J Pharmacol. 2013;718:34-40. https://doi.org/10.1016/j.ejphar.2013.09.020
23. Mukherjee A, Misra S, Howlett NG, Karmakar P. Multinucleation regulated by the Akt/PTEN signaling pathway is a survival strategy for HepG2 cells. Mutat Res. 2013;755:135-40. https://doi.org/10.1016/j.mrgentox.2013.06.009
24. Murugaiyan J, Rockstroh M, Wagner J, Baumann S, Schorsch K, Trump S, et al. Benzo[a]pyrene affects Jurkat T cells in the activated state via the antioxidant response element dependent Nrf2 pathway leading to decreased IL-2 secretion and redirecting glutamine metabolism. Toxicol Appl Pharmacol. 2013;269:307-16. https://doi.org/10.1016/j.taap.2013.03.032
25. Das PJ, Paria N, Gustafson-Seabury A, Vishnoi M, Chaki SP, Love CC, et al. Total RNA isolation from stallion sperm and testis biopsies. Theriogenology. 2010;74:1099-106. https://doi.org/10.1016/j.theriogenology.2010.04.023
26. Viegas O, Žegura B, Pezdric M, Novak M, Ferreira IM, Pinho O, et al. Protective effects of xanthohumol against the genotoxicity of heterocyclic aromatic amines MeIQx and PhIP in bacteria and in human hepatoma (HepG2) cells. Food Chem Toxicol. 2012;50:949-55. https://doi.org/10.1016/j.fct.2011.11.031
27. Stephenson FH. Calculations for molecular biology and biotechnology. A guide to mathematics in the laboratory.
Second edition. London: Academic Press; 2010. p. 458.
28. Koch A, Tamez P, Pezzuto J, Soejarto D. Evaluation of plants used for antimalarial treatment by the Maasai of Kenya. J Ethnopharmacol. 2005;101:95-9. https://doi.org/10.1016/j.jep.2005.03.011
29. Ahir BK, Pratten MK. Developmental cardiotoxicity effects of four commonly used antiepileptic drugs in embryonic chick heart micromass culture and embryonic stem cell culture systems. Toxicol In Vitro. 2014;28:948-60. https://doi.org/10.1016/j.tiv.2014.04.001
30. Darwish W, Ikenaka Y, Eldaly E, Ishizuka M. Mutagenic activation and detoxification of benzo[a]pyrene in vitro by hepatic cytochrome P450 1A1 and phase II enzymes in three meat-producing animals. Food Chem Toxicol. 2010;48:2526-31. https://doi.org/10.1016/j.fct.2010.06.026
31. Zeiger E. Historical perspective on the development of the genetic toxicity test battery in the United States. Environ Mol Mutagen. 2010;51:781-91. https://doi.org/10.1002/em.20602
32. Rodrigues S, Antunes SC, Correia AT, Nunes B. Acute and chronic effects of erythromycin exposure on oxidative stress and genotoxicity parameters of Oncorhynchus mykiss. Sci Total Environ. 2016;545-546:591-600. https://doi.org/10.1016/j.scitotenv.2015.10.138
33. Madikizela B, Ndhlala AR, Finnie JF, van Staden J. Antimycobacterial, anti-inflammatory and genotoxicity evaluation of plants used for the treatment of tuberculosis and related symptoms in South Africa. J Ethnopharmacol. 2014;153:386-91. https://doi.org/10.1016/j.jep.2014.02.034
34. Renglin-Lindh A, Schultz N, Saleh-Gohari N, Helleday T. RAD51C (RAD51L2) is involved in maintaining centrosome number in mitosis. Cytogenet Genome Res. 2007;116:38-45. https://doi.org/10.1159/000097416
35. de Almagro MC, Vucic D. The inhibitor of apoptosis (IAP) proteins are critical regulators of signaling pathways and targets for anti-cancer therapy. Exp Oncol. 2012;34:200-11.
36. Sun M, Meares G, Song L, Jope RS. XIAP associates with GSK3 and inhibits the promotion of intrinsic apoptotic signaling by GSK3. Cell Signal. 2009;21:1857-65. https://doi.org/10.1016/j.cellsig.2009.08.002
37. Boehme K, Dietz Y, Hewitt P, Mueller SO. Activation of P53 in HepG2 cells as surrogate to detect mutagens and promutagens in vitro. Toxicol Lett. 2010;198:272-81. https://doi.org/10.1016/j.toxlet.2010.07.007
38. Bryan HK, Olayanju A, Goldring CE, Park BK. The Nrf2 cell defence pathway: Keap1-dependent and -independent mechanisms of regulation. Biochem Pharmacol. 2013;85:705-17. https://doi.org/10.1016/j.bcp.2012.11.016
39. Klaassen CD, Reisman SA. Nrf2 the rescue: Effects of the antioxidative/electrophilic response on the liver. Toxicol Appl Pharmacol. 2010;244:57-65. https://doi.org/10.1016/j.taap.2010.01.013
40. Stępkowski TM, Kruszewski MK. Molecular cross-talk between the NRF2/KEAP1 signaling pathway, autophagy, and apoptosis. Free Radic Biol Med. 2011;50:1186-95. https://doi.org/10.1016/j.freeradbiomed.2011.01.033
41. Roy A, Sil PC. Taurine protects murine hepatocytes against oxidative stress-induced apoptosis by tert-butyl hydroperoxide via PI3K/Akt and mitochondrial-dependent pathways. Food Chem. 2012;131:1086-96. https://doi.org/10.1016/j.foodchem.2011.09.057
42. Jayakumar S, Pal D, Sandur SK. Nrf2 facilitates repair of radiation induced DNA damage through homologous recombination repair pathway in a ROS independent manner in cancer cells. Mutat Res. 2015;779:33-45. https://doi.org/10.1016/j.mrfmmm.2015.06.007
43. Slamenova D, Kozics K, Hunakova L, Melusova M, Navarova J, Horvathova E. Comparison of biological processes induced in HepG2 cells by tert-butyl hydroperoxide (t-BHP) and hydroperoxide (H2O2): The influence of carvacrol. Mutat Res. 2013;757:15-22. https://doi.org/10.1016/j.mrgentox.2013.03.014
44. International Agency for Research on Cancer -IARC. Some non-heterocyclic polycyclic aromatic hydrocarbons and some related exposures. IARC Monogr. 2010;92:35-773.
45. Cheng T, Scadden DT. Cell cycle regulators in stem cells. In: Lanza R, Atala A, editors. Essentials of Stem Cell Biology. Third edition. Boston: Academic Press; 2014. p. 95-106. https://doi.org/10.1016/B978-0-12-409503-8.00008-1
46. Tyson JJ, Novák B. Irreversible transitions, bistability and checkpoint controls in the eukaryotic cell cycle: A systemslevel understanding. In: Walhout AJM, Vidal M, Dekker J, editors. Handbook of Systems Biology. San Diego: Elsevier; 2013. p. 265-85. https://doi.org/10.1016/B978-0-12-385944-0.00014-9


Métricas de artículo

Cargando métricas ...

Metrics powered by PLOS ALM




Revista Biomédica -  https://doi.org/10.7705/issn.0120-4157
ISSN 0120-4157

Instituto Nacional de Salud
INSTITUTO NACIONAL DE SALUD
Avenida Calle 26 No. 51-20
Apartado aéreo 80334 y 80080
Bogotá, D.C., Colombia, S.A.
Teléfono: 05712207700 Ext. 1386
Correo electrónico: biomedica@ins.gov.co