Skip to main content
NCBI home page
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1998 Dec 1;102(11):1961–1968. doi: 10.1172/JCI3437

Role of oxygen radicals generated by NADPH oxidase in apoptosis induced in human leukemia cells.

W Hiraoka 1, N Vazquez 1, W Nieves-Neira 1, S J Chanock 1, Y Pommier 1
PMCID: PMC509148  PMID: 9835621

Abstract

We have used a human leukemia cell line that, after homologous recombination knockout of the gp91-phox subunit of the phagocyte respiratory-burst oxidase cytochrome b-558, mimics chronic granulomatous disease (X-CGD) to study the role of oxygen radicals in apoptosis. Camptothecin (CPT), a topoisomerase I inhibitor, induced significantly more apoptosis in PLB-985 cells than in X-CGD cells. Sensitivity to CPT was enhanced after neutrophilic differentiation, but was lost after monocytic differentiation. No difference between the two cell lines was observed after treatment with other apoptosis inducers, including etoposide, ultraviolet radiation, ionizing radiation, hydrogen peroxide, or 7-hydroxystaurosporine. After granulocytic differentiation of both cell lines, CPT still induced apoptosis, suggesting independence from replication in fully differentiated and growth-arrested cells. Pyrrolidine dithiocarbamate (an antioxidant inhibitor of NF-kappaB) and catalase partially inhibited CPT-induced DNA fragmentation in granulocytic-differentiated PLB-985 cells, but had no effect in X-CGD cells. Flow cytometry analysis revealed that reactive oxygen intermediates were generated in CPT-treated PLB-985 cells. These data indicate that oxygen radicals generated by NADPH oxidase may contribute directly or indirectly to CPT-induced apoptosis in human leukemia and in neutrophilic-differentiated cells.

Full Text

The Full Text of this article is available as a PDF (373.2 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Akinaga S., Gomi K., Morimoto M., Tamaoki T., Okabe M. Antitumor activity of UCN-01, a selective inhibitor of protein kinase C, in murine and human tumor models. Cancer Res. 1991 Sep 15;51(18):4888–4892. [PubMed] [Google Scholar]
  2. Bertrand R., Sarang M., Jenkin J., Kerrigan D., Pommier Y. Differential induction of secondary DNA fragmentation by topoisomerase II inhibitors in human tumor cell lines with amplified c-myc expression. Cancer Res. 1991 Dec 1;51(23 Pt 1):6280–6285. [PubMed] [Google Scholar]
  3. Bogdanov K. V., Chukhlovin A. B., Zaritskey A. Y., Frolova O. I., Afanasiev B. V. Ultraviolet irradiation induces multiple DNA double-strand breaks and apoptosis in normal granulocytes and chronic myeloid leukaemia blasts. Br J Haematol. 1997 Sep;98(4):869–872. doi: 10.1046/j.1365-2141.1997.2913109.x. [DOI] [PubMed] [Google Scholar]
  4. Briehl M. M., Baker A. F., Siemankowski L. M., Morreale J. Modulation of antioxidant defenses during apoptosis. Oncol Res. 1997;9(6-7):281–285. [PubMed] [Google Scholar]
  5. Bustamante J., Slater A. F., Orrenius S. Antioxidant inhibition of thymocyte apoptosis by dihydrolipoic acid. Free Radic Biol Med. 1995 Sep;19(3):339–347. doi: 10.1016/0891-5849(95)00045-y. [DOI] [PubMed] [Google Scholar]
  6. Chanock S. J., el Benna J., Smith R. M., Babior B. M. The respiratory burst oxidase. J Biol Chem. 1994 Oct 7;269(40):24519–24522. [PubMed] [Google Scholar]
  7. Dancey J., Eisenhauer E. A. Current perspectives on camptothecins in cancer treatment. Br J Cancer. 1996 Aug;74(3):327–338. doi: 10.1038/bjc.1996.362. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Dinauer M. C. The respiratory burst oxidase and the molecular genetics of chronic granulomatous disease. Crit Rev Clin Lab Sci. 1993;30(4):329–369. doi: 10.3109/10408369309082591. [DOI] [PubMed] [Google Scholar]
  9. Feng L., Xia Y., Garcia G. E., Hwang D., Wilson C. B. Involvement of reactive oxygen intermediates in cyclooxygenase-2 expression induced by interleukin-1, tumor necrosis factor-alpha, and lipopolysaccharide. J Clin Invest. 1995 Apr;95(4):1669–1675. doi: 10.1172/JCI117842. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Fernandes R. S., McGowan A. J., Cotter T. G. Mutant H-ras overexpression inhibits drug and U.V. induced apoptosis. Anticancer Res. 1996 Jul-Aug;16(4A):1691–1705. [PubMed] [Google Scholar]
  11. Fukui T., Ishizaka N., Rajagopalan S., Laursen J. B., Capers Q., 4th, Taylor W. R., Harrison D. G., de Leon H., Wilcox J. N., Griendling K. K. p22phox mRNA expression and NADPH oxidase activity are increased in aortas from hypertensive rats. Circ Res. 1997 Jan;80(1):45–51. doi: 10.1161/01.res.80.1.45. [DOI] [PubMed] [Google Scholar]
  12. Goldwasser F., Bae I., Valenti M., Torres K., Pommier Y. Topoisomerase I-related parameters and camptothecin activity in the colon carcinoma cell lines from the National Cancer Institute anticancer screen. Cancer Res. 1995 May 15;55(10):2116–2121. [PubMed] [Google Scholar]
  13. Goldwasser F., Shimizu T., Jackman J., Hoki Y., O'Connor P. M., Kohn K. W., Pommier Y. Correlations between S and G2 arrest and the cytotoxicity of camptothecin in human colon carcinoma cells. Cancer Res. 1996 Oct 1;56(19):4430–4437. [PubMed] [Google Scholar]
  14. Gorman A., McGowan A., Cotter T. G. Role of peroxide and superoxide anion during tumour cell apoptosis. FEBS Lett. 1997 Mar 3;404(1):27–33. doi: 10.1016/s0014-5793(97)00069-0. [DOI] [PubMed] [Google Scholar]
  15. Gupta M., Fujimori A., Pommier Y. Eukaryotic DNA topoisomerases I. Biochim Biophys Acta. 1995 May 17;1262(1):1–14. doi: 10.1016/0167-4781(95)00029-g. [DOI] [PubMed] [Google Scholar]
  16. Huang R. P., Wu J. X., Fan Y., Adamson E. D. UV activates growth factor receptors via reactive oxygen intermediates. J Cell Biol. 1996 Apr;133(1):211–220. doi: 10.1083/jcb.133.1.211. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kanatate T., Nagao N., Sugimoto M., Kageyama K., Fujimoto T., Miwa N. Differential susceptibility of epidermal keratinocytes and neuroblastoma cells to cytotoxicity of ultraviolet-B light irradiation prevented by the oxygen radical-scavenger ascorbate-2-phosphate but not by ascorbate. Cell Mol Biol Res. 1995;41(6):561–567. [PubMed] [Google Scholar]
  18. Kasahara Y., Iwai K., Yachie A., Ohta K., Konno A., Seki H., Miyawaki T., Taniguchi N. Involvement of reactive oxygen intermediates in spontaneous and CD95 (Fas/APO-1)-mediated apoptosis of neutrophils. Blood. 1997 Mar 1;89(5):1748–1753. [PubMed] [Google Scholar]
  19. Lee R., Beauparlant P., Elford H., Ponka P., Hiscott J. Selective inhibition of l kappaB alpha phosphorylation and HIV-1 LTR-directed gene expression by novel antioxidant compounds. Virology. 1997 Aug 4;234(2):277–290. doi: 10.1006/viro.1997.8642. [DOI] [PubMed] [Google Scholar]
  20. Lin K. I., Lee S. H., Narayanan R., Baraban J. M., Hardwick J. M., Ratan R. R. Thiol agents and Bcl-2 identify an alphavirus-induced apoptotic pathway that requires activation of the transcription factor NF-kappa B. J Cell Biol. 1995 Dec;131(5):1149–1161. doi: 10.1083/jcb.131.5.1149. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Lorès P., Morin L., Luna R., Gacon G. Enhanced apoptosis in the thymus of transgenic mice expressing constitutively activated forms of human Rac2GTPase. Oncogene. 1997 Jul 31;15(5):601–605. doi: 10.1038/sj.onc.1201378. [DOI] [PubMed] [Google Scholar]
  22. Los M., Schenk H., Hexel K., Baeuerle P. A., Dröge W., Schulze-Osthoff K. IL-2 gene expression and NF-kappa B activation through CD28 requires reactive oxygen production by 5-lipoxygenase. EMBO J. 1995 Aug 1;14(15):3731–3740. doi: 10.1002/j.1460-2075.1995.tb00043.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Morel F., Doussiere J., Vignais P. V. The superoxide-generating oxidase of phagocytic cells. Physiological, molecular and pathological aspects. Eur J Biochem. 1991 Nov 1;201(3):523–546. doi: 10.1111/j.1432-1033.1991.tb16312.x. [DOI] [PubMed] [Google Scholar]
  24. Morris E. J., Geller H. M. Induction of neuronal apoptosis by camptothecin, an inhibitor of DNA topoisomerase-I: evidence for cell cycle-independent toxicity. J Cell Biol. 1996 Aug;134(3):757–770. doi: 10.1083/jcb.134.3.757. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Peter M. E., Heufelder A. E., Hengartner M. O. Advances in apoptosis research. Proc Natl Acad Sci U S A. 1997 Nov 25;94(24):12736–12737. doi: 10.1073/pnas.94.24.12736. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Pommier Y. Eukaryotic DNA topoisomerase I: genome gatekeeper and its intruders, camptothecins. Semin Oncol. 1996 Feb;23(1 Suppl 3):3–10. [PubMed] [Google Scholar]
  27. Potmesil M. Camptothecins: from bench research to hospital wards. Cancer Res. 1994 Mar 15;54(6):1431–1439. [PubMed] [Google Scholar]
  28. Pourzand C., Rossier G., Reelfs O., Borner C., Tyrrell R. M. Overxpression of Bcl-2 inhibits UVA-mediated immediate apoptosiinrat 6 fibroblasts: evidence for the involvement of Bcl-2 as an antioxidant. Cancer Res. 1997 Apr 15;57(8):1405–1411. [PubMed] [Google Scholar]
  29. Quillet-Mary A., Jaffrézou J. P., Mansat V., Bordier C., Naval J., Laurent G. Implication of mitochondrial hydrogen peroxide generation in ceramide-induced apoptosis. J Biol Chem. 1997 Aug 22;272(34):21388–21395. doi: 10.1074/jbc.272.34.21388. [DOI] [PubMed] [Google Scholar]
  30. Quinn M. T., Evans T., Loetterle L. R., Jesaitis A. J., Bokoch G. M. Translocation of Rac correlates with NADPH oxidase activation. Evidence for equimolar translocation of oxidase components. J Biol Chem. 1993 Oct 5;268(28):20983–20987. [PubMed] [Google Scholar]
  31. Roos D., de Boer M., Kuribayashi F., Meischl C., Weening R. S., Segal A. W., Ahlin A., Nemet K., Hossle J. P., Bernatowska-Matuszkiewicz E. Mutations in the X-linked and autosomal recessive forms of chronic granulomatous disease. Blood. 1996 Mar 1;87(5):1663–1681. [PubMed] [Google Scholar]
  32. Schulze-Osthoff K., Bakker A. C., Vanhaesebroeck B., Beyaert R., Jacob W. A., Fiers W. Cytotoxic activity of tumor necrosis factor is mediated by early damage of mitochondrial functions. Evidence for the involvement of mitochondrial radical generation. J Biol Chem. 1992 Mar 15;267(8):5317–5323. [PubMed] [Google Scholar]
  33. Shao R. G., Cao C. X., Pommier Y. Activation of PKCalpha downstream from caspases during apoptosis induced by 7-hydroxystaurosporine or the topoisomerase inhibitors, camptothecin and etoposide, in human myeloid leukemia HL60 cells. J Biol Chem. 1997 Dec 12;272(50):31321–31325. doi: 10.1074/jbc.272.50.31321. [DOI] [PubMed] [Google Scholar]
  34. Shao R. G., Shimizu T., Pommier Y. 7-Hydroxystaurosporine (UCN-01) induces apoptosis in human colon carcinoma and leukemia cells independently of p53. Exp Cell Res. 1997 Aug 1;234(2):388–397. doi: 10.1006/excr.1997.3650. [DOI] [PubMed] [Google Scholar]
  35. Shimizu T., O'Connor P. M., Kohn K. W., Pommier Y. Unscheduled activation of cyclin B1/Cdc2 kinase in human promyelocytic leukemia cell line HL60 cells undergoing apoptosis induced by DNA damage. Cancer Res. 1995 Jan 15;55(2):228–231. [PubMed] [Google Scholar]
  36. Slichenmyer W. J., Rowinsky E. K., Donehower R. C., Kaufmann S. H. The current status of camptothecin analogues as antitumor agents. J Natl Cancer Inst. 1993 Feb 17;85(4):271–291. doi: 10.1093/jnci/85.4.271. [DOI] [PubMed] [Google Scholar]
  37. Wang J. C., Caron P. R., Kim R. A. The role of DNA topoisomerases in recombination and genome stability: a double-edged sword? Cell. 1990 Aug 10;62(3):403–406. doi: 10.1016/0092-8674(90)90002-v. [DOI] [PubMed] [Google Scholar]
  38. Wang J. F., Jerrells T. R., Spitzer J. J. Decreased production of reactive oxygen intermediates is an early event during in vitro apoptosis of rat thymocytes. Free Radic Biol Med. 1996;20(4):533–542. doi: 10.1016/0891-5849(95)02085-3. [DOI] [PubMed] [Google Scholar]
  39. Wenger R. H., Marti H. H., Schuerer-Maly C. C., Kvietikova I., Bauer C., Gassmann M., Maly F. E. Hypoxic induction of gene expression in chronic granulomatous disease-derived B-cell lines: oxygen sensing is independent of the cytochrome b558-containing nicotinamide adenine dinucleotide phosphate oxidase. Blood. 1996 Jan 15;87(2):756–761. [PubMed] [Google Scholar]
  40. Wu J., Liu L. F. Processing of topoisomerase I cleavable complexes into DNA damage by transcription. Nucleic Acids Res. 1997 Nov 1;25(21):4181–4186. doi: 10.1093/nar/25.21.4181. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Yan S. R., Berton G. Regulation of Src family tyrosine kinase activities in adherent human neutrophils. Evidence that reactive oxygen intermediates produced by adherent neutrophils increase the activity of the p58c-fgr and p53/56lyn tyrosine kinases. J Biol Chem. 1996 Sep 20;271(38):23464–23471. doi: 10.1074/jbc.271.38.23464. [DOI] [PubMed] [Google Scholar]
  42. Yu L., Zhen L., Dinauer M. C. Biosynthesis of the phagocyte NADPH oxidase cytochrome b558. Role of heme incorporation and heterodimer formation in maturation and stability of gp91phox and p22phox subunits. J Biol Chem. 1997 Oct 24;272(43):27288–27294. doi: 10.1074/jbc.272.43.27288. [DOI] [PubMed] [Google Scholar]
  43. Zhen L., King A. A., Xiao Y., Chanock S. J., Orkin S. H., Dinauer M. C. Gene targeting of X chromosome-linked chronic granulomatous disease locus in a human myeloid leukemia cell line and rescue by expression of recombinant gp91phox. Proc Natl Acad Sci U S A. 1993 Nov 1;90(21):9832–9836. doi: 10.1073/pnas.90.21.9832. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. el Benna J., Ruedi J. M., Babior B. M. Cytosolic guanine nucleotide-binding protein Rac2 operates in vivo as a component of the neutrophil respiratory burst oxidase. Transfer of Rac2 and the cytosolic oxidase components p47phox and p67phox to the submembranous actin cytoskeleton during oxidase activation. J Biol Chem. 1994 Mar 4;269(9):6729–6734. [PubMed] [Google Scholar]

Articles from Journal of Clinical Investigation are provided here courtesy of American Society for Clinical Investigation

RESOURCES