, Volume 17, Issue 6, pp 707–713 | Cite as

Zinc protects against arsenic-induced apoptosis in a neuronal cell line, measured by DEVD-caspase activity

  • Austin Graham Milton
  • Peter David Zalewski
  • Ranjit Nihal Ratnaike


Acute and chronic arsenic exposure results in toxicity in humans and causes many neurological and other manifestations. For the first time the present study reports that zinc decreases arsenic-induced apoptosis and also confirms a single report of apoptosis induced by arsenic in a neuronal cell line. Apoptosis measured by DEVD-caspase activity peaked between 10 μM and 20 μM of arsenic trioxide. Higher concentrations of arsenic up to 40 μM caused increasing cell death with diminishing DEVD-caspase activity. The beneficial effect of zinc was proportional to its concentration with a significant decrease in arsenic-induced DEVD-caspase activity at 50 μM and 75 μM zinc (P<0.05). This finding may be of therapeutic benefit in people suffering from chronic exposure to arsenic from natural sources, a global problem especially relevant to millions of people on the Indian subcontinent.

arsenic trioxide 2.3D mouse neuroepithelial cells DEVD-caspase zinc sulfate 


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  1. Abernathy CO, Liu YP, Longfellow D, Aposhian HV, Beck B, Fowler B, Goyer R, Menzer R, Rossman T, Thompson C, Waalkes M. 1999 Arsenic: health effects, mechanisms of actions, and research issues. Environ Health Perspect 107, 593–597.Google Scholar
  2. Akao Y, Nakagawa Y, Akiyama K. 1999 Arsenic trioxide induces apoptosis in neuroblastoma cell lines through the activation of caspase3 in vitro.FEBS Letters 455, 59–62.Google Scholar
  3. Bartlett PF, Reid HH, Bailey KA, Bernard O. 1988 Immortalisation of mouse neural precursor cells by the ic-myconcogene. Proc Nat Acad Sci 85, 3255–3259.Google Scholar
  4. Campbell JP, Alvarez JA. 1989 Acute arsenic intoxication. Am Family Physician 40, 93–97.Google Scholar
  5. Chinnaiyan AM, Dixit VM. 1996 The cell-death machine. Curr Biol 6(5), 555–562.Google Scholar
  6. Cohen GM. 1997 Caspases: the executioners of apoptosis. Biochem J 326, 1–16.Google Scholar
  7. Cousins RJ. 1986 Toward a molecular understanding of zinc metabolism. Clin Physiol Biochem4(1), 20–30.Google Scholar
  8. Durnam DM, Palmiter RD. 1981 Transcriptional regulation of the mouse metallothionein-I gene by heavy metals. J Biol Chem 256(11), 5712–5716.Google Scholar
  9. Feng Y, LeBlanc MH. 2003 Treatment of hypoxic-ischemic brain injury in newborn rats with TPCK 3 h after hypoxia decreases caspase-9 activation and improves neuropathological outcome. Dev Neurosci 25, 34–40.Google Scholar
  10. Fraker PJ, Telford WG. 1997 A reappraisal of the role of zinc in life and death decisions of cells. Proc Soc Exp Biol Med 215, 229–236.Google Scholar
  11. Hall AH. 2002 Chronic arsenic poisoning. Toxicol Lett 128(1–3), 69–72.Google Scholar
  12. Haridas V, Higuchi M, Jayatilake GS, Bailey D, Mujoo K, Blake ME, Arntzen CJ, Gutterman JU. 2001 Avicins: Triterpenoid saponins from Acacia victoriae (Bentham) induce apoptosis by mitochondrial perturbation. Proc Nat Acad Sci 98, 5821–5826.Google Scholar
  13. Ho LH, Ratnaike RN, Zalewski PD. 2000 Involvement of intra-cellular labile zinc in suppression of DEVD-Caspase activity in human neuroblastoma cells. Biochem Biophys Res Comm 268, 148–154.Google Scholar
  14. Huang RN, Lee TC. 1996 Cellular uptake of trivalent arsenite and pentavalent arsenate in KB cells cultured in phosphate-free medium. Toxicol Appl Pharmacol 136(2), 243–249.Google Scholar
  15. Johanning GL, O'Dell BL. 1989 Effect of zinc deficiency and food restriction in rats on erythrocyte membrane zinc, phospholipid and protein content. J Nutr 119(11), 1654–1660.Google Scholar
  16. Kambe T, Yamaguchi-Iwai Y, Sasaki R, Nagao M. 2004 Overview of mammalian zinc transporters. Cell Mol Life Sci 61, 49–68.Google Scholar
  17. Krzyzowska M, Schollenberger A, Skierski J, Niemialtowski M. 2002 Apoptosis during ectromelia orthopoxvirus infection is DEVDase dependent: in vitro and in vivo studies. Microbes Infect 4, 599–611.Google Scholar
  18. Kwon KB, Yang JY, Ryu DG, Rho HW, Kim JS, Park JW, Kim HR, Park BH. 2001 Vibrio vulnificus cytolysin induces superoxide anion-initiated apoptotic signalling pathway in human ECV304 cells. J Biol Chem 276, 47518–47523.Google Scholar
  19. Larochette N, Decaudin D, Jacotot E, Brenner C, Marzo I, Susin SA, Zamzami N, Xie Z, Reed J, Kroemer G. 1999 Arsenite induces apoptosis via a direct effect on the mitochondrial permeability transition pore. Exp Cell Res 249, 413–421.Google Scholar
  20. Le Quesne PM. 1982 Metal-induced diseases of the nervous system. Br J Hosp Med 28, 534–538.Google Scholar
  21. Li W, Chou IN. 1992 Effects of sodium arsenite on the cytoskel-eton and cellular glutathione levels in cultured cells. Toxicol Appl Pharmacol 114(1), 132–139.Google Scholar
  22. Lorenzo HK, Susin SA, Penninger J, Kroemer G. 1999 Apop-tosis inducing factor (AIF): a phylogenetically old, caspase-independent effector of cell death. Cell Death Differ 6(6), 516–524.Google Scholar
  23. Menzel DB, Hamadeh HK, Lee E, Meacher DM, Said V, Rasmussen RE, Greene H, Roth RN. 1999 Arsenic binding proteins from human lymphoblastoid cells. Toxicol Lett 105, 89–101.Google Scholar
  24. Mundle SD, Reza S, Ali A, Mativi Y, Shetty V, Venugopal P, Gregory SA, Raza A. 1999 Correlation of tumor necrosis factor alpha (TNF alpha) with high caspase 3-like activity in myelodys-plastic syndromes. Cancer Lett 140, 201–207.Google Scholar
  25. Pelicano H, Feng L, Zhou Y, Carew JS, Hileman EO, Plunkett W, Keating MJ, Huang P. 2002 Inhibition of mitochondrial respir-ation: a novel strategy to enhance drug-induced apoptosis in human leukemia cells by a reactive oxygen species-mediated mechanism. J Biol Chem 278, 37832–37839.Google Scholar
  26. Pfeiffer CJ, Cho CH. 1980 Modulating effect by zinc on hep-atic lysosomal fragility induced by surface-active agents. Res Commun Chem Pathol Pharmacol 27(3), 587–598.Google Scholar
  27. Ratnaike RN. 2003 Acute and chronic arsenic toxicity. Postgrad Med J 79(933), 391–396.Google Scholar
  28. Shen ZX, Chen GQ, Ni JH, Li XS, Xiong SM, Qiu QY, Zhu J, Tang W, Sun GL, Yang KQ, Chen Y, Zhou L, Fang ZW, Wang YT, Ma J, Zhang P, Zhang TD, Chen SJ, Chen Z, Wang ZY. 1997 Use of arsenic trioxide (As 2 O 3 )in the treatment of acute promyelocytic leukemia (APL): II. Clinical efficacy and pharmacokinetics in relapsed patients. Blood 89, 3354–3360.Google Scholar
  29. Shen ZY, Shen WY, Chen MH, Shen J, Zeng Y. 2003 Reactive oxy-gen species and antioxidants in apoptosis of esophageal cancer cells induced by As2O3. Int J Mol Med 11, 479–484.Google Scholar
  30. Soignet SL, Maslak P, Wang ZG, Jhanwar S, Calleja E, Dardashti LJ, Corso D, DeBlasio A, Gabrilove J, Scheinberg DA, Pandolfi PP, Warrell RP Jr. 1998 Complete remission after treatment of acute promyelocytic leukemia with arsenic trioxide. N Engl J Med 339, 1341–1348.Google Scholar
  31. Srinivasula SM, Fernandes-Alnemri T, Zangrilli J, Robertson N, Armstrong RC, Wang L, Trapani JA, Tomaselli KJ, Litwack. G, Alnemri ES. 1996 The ced-3/interleukin 1beta converting enzyme-like homolog Mch6 and the lamin-cleaving enzyme Mch2alpha are substrates for the apoptotic mediator CPP32. J Biol Chem 271, 27099–27106.Google Scholar
  32. Stennicke HR, Salvesen GS. 1997 Biochemical characteristics of Caspases-3,-6,-7, and-8. J Biol Chem 272, 25719–25723.Google Scholar
  33. Sullivan JF, Jetton MM, Burch RE. 1979 A zinc tolerance test. J Lab Clin Med 93, 485–492.Google Scholar
  34. Sun HD, Ma L, Hu XC, Zhang TD. 1992 Ai-Lin 1 treated 32 cases of acute promyelocytic leukemia. Chin J Integrat of Chinese and Western Med 12, 170.Google Scholar
  35. Sunderman FW Jr. 1995 The influence of zinc on apoptosis. Ann Clin Lab Sci 25, 134–142.Google Scholar
  36. Susin SA, Daugas E, Ravagnan L, Samejima K, Zamzami K, Loeffler M, Costantini P, Ferri KF, Irinopoulou T, Prevost MC, Brothers G, Mak TW, Penninger J, Earnshaw WC, Kroemer G. 2000 Two distinct pathways leading to nuclear apoptosis. J Exp Med 192, 571–580.Google Scholar
  37. Sweetman SC. ed. Martindale-the complete drug reference, 33rd edition 2002. London, UK: Pharmaceutical Press: p. 1398.Google Scholar
  38. Takahashi A, Alnemri ES, Lazebnik YA, Fernandes-Alnemri T, Litwack G, Moir RD, Goldman RD, Poirier GG, Kaufmann SH, Earnshaw WC. 1996 Cleavage of lamin A by Mch2 alpha but not CPP32: multiple interleukin 1 beta-converting enzyme-related proteases with distinct substrate recognition properties are active in apoptosis. Proc Natl Acad Sci 93(16), 8395–8400.Google Scholar
  39. Toyama M, Yamashita M, Hirayama N, Murooka Y. 2002 Interactions of arsenic with human metallothionein-2. J Bioche m132,217–221.Google Scholar
  40. Truong-Tran AQ, Ho LH, Chai F, Zalewski PD. 2000 Cellular zinc fluxes and the regulation of apoptosis/gene-directed cell death. J Nutr 130(5S Suppl), 1459S-1466S.Google Scholar
  41. Vallee BL, Falchuk KH. 1993 The biochemical basis of zinc physiology. Physiol Rev 73, 79–105.Google Scholar
  42. Zalewski PD, Forbes IJ, Betts WH. 1993 Correlation of apoptosis with change in intracellular labile Zn(II) using Zinquin ((2-methyl-8-p-toluenesulphonamido-6-quinolyloxy)acetic acid), a new specific fluorescent probe for Zn(II). Biochem J 296, 403–408.Google Scholar
  43. Zhang TL, Gao YX, Lu JF, Wang K. 2000 Arsenite, arsenate and vanadate affect human erythrocyte membranes. J Inorg Biochem 79, 195–203.Google Scholar

Copyright information

© Kluwer Academic Publishers 2004

Authors and Affiliations

  • Austin Graham Milton
    • 1
  • Peter David Zalewski
    • 1
  • Ranjit Nihal Ratnaike
    • 1
  1. 1.Department of MedicineThe Queen Elizabeth HospitalWoodvilleSouth Australia

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