1887

Abstract

Naegleria fowleri and Naegleria gruberi belong to the free-living amoebae group. It is widely known that the non-pathogenic species N. gruberi is usually employed as a model to describe molecular pathways in this genus, mainly because its genome has been recently described. However, N. fowleri is an aetiological agent of primary amoebic meningoencephalitis, an acute and fatal disease. Currently, the most widely used drug for its treatment is amphotericin B (AmB). It was previously reported that AmB has an amoebicidal effect in both N. fowleri and N. gruberi trophozoites by inducing morphological changes that resemble programmed cell death (PCD). PCD is a mechanism that activates morphological, biochemical and genetic changes. However, PCD has not yet been characterized in the genus Naegleria. The aim of the present work was to evaluate the typical markers to describe PCD in both amoebae. These results showed that treated trophozoites displayed several parameters of apoptosis-like PCD in both species. We observed ultrastructural changes, an increase in reactive oxygen species, phosphatidylserine externalization and a decrease in intracellular potassium, while DNA degradation was evaluated using the TUNEL assay and agarose gels, and all of these parameters are related to PCD. Finally, we analysed the expression of apoptosis-related genes, such as sir2 and atg8, in N. gruberi. Taken together, our results showed that AmB induces the morphological, biochemical and genetic changes of apoptosis-like PCD in the genus Naegleria.

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2017-07-19
2019-10-17
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References

  1. Visvesvara GS, Moura H, Schuster FL. Pathogenic and opportunistic free-living amoebae: Acanthamoeba spp., Balamuthia mandrillaris, Naegleria fowleri, and Sappinia diploidea. FEMS Immunol Med Microbiol 2007;50:1–26 [CrossRef][PubMed]
    [Google Scholar]
  2. Trabelsi H, Dendana F, Sellami A, Sellami H, Cheikhrouhou F et al. Pathogenic free-living amoebae: epidemiology and clinical review. Pathol Biol 2012;60:399–405 [CrossRef][PubMed]
    [Google Scholar]
  3. Martínez-Castillo M, Cárdenas-Zúñiga R, Coronado-Velázquez D, Debnath A, Serrano-Luna J et al. Naegleria fowleri after 50 years: is it a neglected pathogen?. J Med Microbiol 2016;65:885–896 [CrossRef][PubMed]
    [Google Scholar]
  4. Jain R, Prabhakar S, Modi M, Bhatia R, Sehgal R. Naegleria meningitis: a rare survival. Neurol India 2002;50:470–472[PubMed]
    [Google Scholar]
  5. Vargas-Zepeda J, Gómez-Alcalá AV, Vásquez-Morales JA, Licea-Amaya L, de Jonckheere JF et al. Successful treatment of Naegleria fowleri meningoencephalitis by using intravenous amphotericin B, fluconazole and rifampicin. Arch Med Res 2005;36:83–86 [CrossRef][PubMed]
    [Google Scholar]
  6. Sharma A, Sharma A, Guleria S. Successful treatment of a case of primary amoebic meningoencephalitis: how important is history taking. Indian J Crit Care Med 2015;19:126–127 [CrossRef][PubMed]
    [Google Scholar]
  7. Yadav D, Aneja S, Dutta R, Maheshwari A, Seth A. Youngest survivor of Naegleria meningitis. Indian J Pediatr 2013;80:253–254 [CrossRef][PubMed]
    [Google Scholar]
  8. Brajtburg J, Bolard J. Carrier effects on biological activity of amphotericin B. Clin Microbiol Rev 1996;9:512–531[PubMed]
    [Google Scholar]
  9. Fritz-Laylin LK, Prochnik SE, Ginger ML, Dacks JB, Carpenter ML et al. The genome of Naegleria gruberi illuminates early eukaryotic versatility. Cell 2010;140:631–642 [CrossRef][PubMed]
    [Google Scholar]
  10. Fritz-Laylin LK, Ginger ML, Walsh C, Dawson SC, Fulton C. The Naegleria genome: a free-living microbial eukaryote lends unique insights into core eukaryotic cell biology. Res Microbiol 2011;162:607–618 [CrossRef][PubMed]
    [Google Scholar]
  11. Opperdoes FR, de Jonckheere JF, Tielens AG. Naegleria gruberi metabolism. Int J Parasitol 2011;41:915–924 [CrossRef][PubMed]
    [Google Scholar]
  12. Schuster FL, Rechthand E. In vitro effects of amphotericin B on growth and ultrastructure of the amoeboflagellates Naegleria gruberi and Naegleria fowleri. Antimicrob Agents Chemother 1975;8:591–605 [CrossRef][PubMed]
    [Google Scholar]
  13. Collins RJ, Harmon BV, Gobé GC, Kerr JF. Internucleosomal DNA cleavage should not be the sole criterion for identifying apoptosis. Int J Radiat Biol 1992;61:451–453 [CrossRef][PubMed]
    [Google Scholar]
  14. Edinger AL, Thompson CB. Death by design: apoptosis, necrosis and autophagy. Curr Opin Cell Biol 2004;16:663–669 [CrossRef][PubMed]
    [Google Scholar]
  15. Bruchhaus I, Roeder T, Rennenberg A, Heussler VT. Protozoan parasites: programmed cell death as a mechanism of parasitism. Trends Parasitol 2007;23:376–383 [CrossRef][PubMed]
    [Google Scholar]
  16. Kerr JF, Wyllie AH, Currie AR. Apoptosis: a basic biological phenomenon with wide-ranging implications in tissue kinetics. Br J Cancer 1972;26:239–257 [CrossRef][PubMed]
    [Google Scholar]
  17. Dhurga DB, Suresh K, Tan TC. Granular formation during apoptosis in Blastocystis sp. exposed to metronidazole (MTZ). PLoS One 2016;11:e0155390 [CrossRef][PubMed]
    [Google Scholar]
  18. Yuan J. Transducing signals of life and death. Curr Opin Cell Biol 1997;9:247–251 [CrossRef][PubMed]
    [Google Scholar]
  19. Nasirudeen AM, Hian YE, Singh M, Tan KS. Metronidazole induces programmed cell death in the protozoan parasite Blastocystis hominis. Microbiology 2004;150:33–43 [CrossRef][PubMed]
    [Google Scholar]
  20. Nasirudeen AM, Tan KS. Caspase-3-like protease influences but is not essential for DNA fragmentation in Blastocystis undergoing apoptosis. Eur J Cell Biol 2004;83:477–482 [CrossRef][PubMed]
    [Google Scholar]
  21. Villalba JD, Gómez C, Medel O, Sánchez V, Carrero JC et al. Programmed cell death in Entamoeba histolytica induced by the aminoglycoside G418. Microbiology 2007;153:3852–3863 [CrossRef][PubMed]
    [Google Scholar]
  22. Jiménez-Ruiz A, Alzate JF, Macleod ET, Lüder CG, Fasel N et al. Apoptotic markers in protozoan parasites. Parasit Vectors 2010;3:104 [CrossRef][PubMed]
    [Google Scholar]
  23. Monroy VS, Flores MO, Villalba-Magdaleno JD, Garcia CG, Ishiwara DG. Entamoeba histolytica: differential gene expression during programmed cell death and identification of early pro- and anti-apoptotic signals. Exp Parasitol 2010;126:497–505 [CrossRef][PubMed]
    [Google Scholar]
  24. Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods 2001;25:402–408 [CrossRef][PubMed]
    [Google Scholar]
  25. Nguewa PA, Fuertes MA, Valladares B, Alonso C, Pérez JM. Programmed cell death in trypanosomatids: a way to maximize their biological fitness?. Trends Parasitol 2004;20:375–380 [CrossRef][PubMed]
    [Google Scholar]
  26. Al-Olayan EM, Williams GT, Hurd H. Apoptosis in the malaria protozoan, Plasmodium berghei: a possible mechanism for limiting intensity of infection in the mosquito. Int J Parasitol 2002;32:1133–1143 [CrossRef][PubMed]
    [Google Scholar]
  27. Lee N, Bertholet S, Debrabant A, Muller J, Duncan R et al. Programmed cell death in the unicellular protozoan parasite Leishmania. Cell Death Differ 2002;9:53–64 [CrossRef][PubMed]
    [Google Scholar]
  28. Pais-Morales J, Betanzos A, García-Rivera G, Chávez-Munguía B, Shibayama M. Resveratrol induces apoptosis-like death and prevents in vitro and in vivo virulence of Entamoeba histolytica. PLoS One 2016;11:e0146287[CrossRef]
    [Google Scholar]
  29. Cornillon S, Foa C, Davoust J, Buonavista N, Gross JD et al. Programmed cell death in Dictyostelium. J Cell Sci 1994;107:2691–2704[PubMed]
    [Google Scholar]
  30. Figarella K, Rawer M, Uzcategui NL, Kubata BK, Lauber K et al. Prostaglandin D2 induces programmed cell death in Trypanosoma brucei bloodstream form. Cell Death Differ 2005;12:335–346 [CrossRef][PubMed]
    [Google Scholar]
  31. Wanderley JL, Pinto da Silva LH, Deolindo P, Soong L, Borges VM et al. Cooperation between apoptotic and viable metacyclics enhances the pathogenesis of leishmaniasis. PLoS One 2009;4:e5733 [CrossRef][PubMed]
    [Google Scholar]
  32. Kim JH, Kim D, Shin HJ. Contact-independent cell death of human microglial cells due to pathogenic Naegleria fowleri trophozoites. Korean J Parasitol 2008;46:217–221 [CrossRef][PubMed]
    [Google Scholar]
  33. Mukherjee SB, Das M, Sudhandiran G, Shaha C. Increase in cytosolic Ca2+ levels through the activation of non-selective cation channels induced by oxidative stress causes mitochondrial depolarization leading to apoptosis-like death in Leishmania donovani promastigotes. J Biol Chem 2002;277:24717–24727 [CrossRef][PubMed]
    [Google Scholar]
  34. Bortner CD, Cidlowski JA. Caspase independent/dependent regulation of K+, cell shrinkage, and mitochondrial membrane potential during lymphocyte apoptosis. J Biol Chem 1999;274:21953–21962 [CrossRef][PubMed]
    [Google Scholar]
  35. Lang F, Föller M, Lang K, Lang P, Ritter M et al. Cell volume regulatory ion channels in cell proliferation and cell death. Methods Enzymol 2007;428:209–225 [CrossRef][PubMed]
    [Google Scholar]
  36. Tiphine M, Letscher-Bru V, Herbrecht R. Amphotericin B and its new formulations: pharmacologic characteristics, clinical efficacy, and tolerability. Transpl Infect Dis 1999;1:273–283 [CrossRef][PubMed]
    [Google Scholar]
  37. Almonte-Becerril M, Navarro-Garcia F, Gonzalez-Robles A, Vega-Lopez MA, Lavalle C et al. Cell death of chondrocytes is a combination between apoptosis and autophagy during the pathogenesis of osteoarthritis within an experimental model. Apoptosis 2010;15:631–638 [CrossRef][PubMed]
    [Google Scholar]
  38. Levine B, Yuan J. Autophagy in cell death: an innocent convict?. J Clin Invest 2005;115:2679–2688 [CrossRef][PubMed]
    [Google Scholar]
  39. Xue L, Fletcher GC, Tolkovsky AM. Autophagy is activated by apoptotic signalling in sympathetic neurons: an alternative mechanism of death execution. Mol Cell Neurosci 1999;14:180–198 [CrossRef][PubMed]
    [Google Scholar]
  40. Brennand A, Gualdrón-López M, Coppens I, Rigden DJ, Ginger ML et al. Autophagy in parasitic protists: unique features and drug targets. Mol Biochem Parasitol 2011;177:83–99 [CrossRef][PubMed]
    [Google Scholar]
  41. Cárdenas-Zúñiga R, Sánchez-Monroy V, Bermúdez-Cruz RM, Rodríguez MA, Serrano-Luna J et al. Ubiquitin-like Atg8 protein is expressed during autophagy and the encystation process in Naegleria gruberi. Parasitol Res 2017;116:303–312 [CrossRef][PubMed]
    [Google Scholar]
  42. Wyllie AH. Glucocorticoid-induced thymocyte apoptosis is associated with endogenous endonuclease activation. Nature 1980;284:555–556 [CrossRef][PubMed]
    [Google Scholar]
  43. Kaczanowski S, Sajid M, Reece SE. Evolution of apoptosis-like programmed cell death in unicellular protozoan parasites. Parasit Vectors 2011;4:44 [CrossRef][PubMed]
    [Google Scholar]
  44. Verhoven B, Schlegel RA, Williamson P. Mechanisms of phosphatidylserine exposure, a phagocyte recognition signal, on apoptotic T lymphocytes. J Exp Med 1995;182:1597–1601 [CrossRef][PubMed]
    [Google Scholar]
  45. Bortner CD, Hughes FM, Cidlowski JA. A primary role for K+ and Na+ efflux in the activation of apoptosis. J Biol Chem 1997;272:32436–32442 [CrossRef][PubMed]
    [Google Scholar]
  46. Hughes FM, Bortner CD, Purdy GD, Cidlowski JA. Intracellular K+ suppresses the activation of apoptosis in lymphocytes. J Biol Chem 1997;272:30567–30576 [CrossRef][PubMed]
    [Google Scholar]
  47. Madeo F, Fröhlich E, Ligr M, Grey M, Sigrist SJ et al. Oxygen stress: a regulator of apoptosis in yeast. J Cell Biol 1999;145:757–767 [CrossRef][PubMed]
    [Google Scholar]
  48. Vergnes B, Sereno D, Madjidian-Sereno N, Lemesre JL, Ouaissi A. Cytoplasmic SIR2 homologue overexpression promotes survival of Leishmania parasites by preventing programmed cell death. Gene 2002;296:139–150 [CrossRef][PubMed]
    [Google Scholar]
  49. Geng J, Klionsky DJ. The Atg8 and Atg12 ubiquitin-like conjugation systems in macroautophagy. EMBO Rep 2008;9:859–864 [CrossRef][PubMed]
    [Google Scholar]
  50. Xie Z, Nair U, Klionsky DJ. Atg8 controls phagophore expansion during autophagosome formation. Mol Biol Cell 2008;19:3290–3298 [CrossRef][PubMed]
    [Google Scholar]
  51. Reed JC. Mechanisms of apoptosis. Am J Pathol 2000;157:1415–1430 [CrossRef][PubMed]
    [Google Scholar]
  52. Mcluskey K, Moss CX, Mottram JC. Purification, characterization, and crystallization of Trypanosoma metacaspases. Methods Mol Biol 2014;1133:203–221 [CrossRef][PubMed]
    [Google Scholar]
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