Abstract
Cellular imbalance in the levels of antioxidants and reactive oxygen species (ROS) is directly associated with a number of pathological states and results in programmed cell death or apoptosis. We demonstrate the use ofin vitro culturedSpodoptera frugiperda (sf9) insect cells as a model to study oxidative stress induced programmed cell death. Apoptosis ofin vitro cultured sf9 cells was induced by the exogenous treatment of H2O2 to cells growing in culture. The AD50 (concentration of H2O2 inducing about 50% apoptotic response) varied with the duration of treatment, batch to batch variation of H2O2 and the physiological state of cells. At 24 h post-treatment with H2O2 AD50 was about 475 Μm. Apoptosis could also be induced byin situ generation of H2O2 by the inhibition of catalase activity upon hydroxylamine treatment. Hydroxylamine acted synergistically with H2O2 with an AD50 of 2.2 mM. DMSO, a free radical scavenger, inhibited H2O2-induced apoptosis thereby confirming the involvement of reactive oxygen species. Exposure of cells to UV radiation (312 nm) resulted in a dose-dependent induction of apoptosis. These results provide evidence on the novel use of insect cells as a model for oxidative stress-induced apoptosis.
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Abate C, Patel L, Rauscher III F J and Curran T 1990 Redox regulation of Fos and Jun DNA-binding activityin vitro;Science 249 1157–1161
Ahmad M, Srinivasula S M, Wang L, Litwack G, Fernandes-Alnemri T and Alnemri E S 1997Spodoptera frugiperda caspase-1, a novel insect death protease that cleaves the nuclear immunophilin FKBP46, is the target of the baculovirus antiapoptotic protein p35;J. Biol. Chem. 272 1421–1424
Begum R, Taneja T K, Mohan M, Pathak N, Sah N K, Athar M and Hasnain S E 1999 The control and execution of programmed cell death; inRecent aspects of fundamental and applied radiobiology (eds) F H ASchneeweiss and R NSharan (Julisch, GmBH: Foschungszentrum) vol. 31 (in press)
Bump N J, Hackett M, Hugunin M, Seshagiri S, Brady K, Chen P, Ferenz S, Ghayur T, Li P, Licari P, Mankovich J, Shi L, Greenberg A H, Miller L K and Wong W W 1995 Inhibition of ICE family proteases by baculovirus antiapoptotic protein p35;Science 269 1885–1887
Clem R J and Miller L K 1994 Control of programmed cell death by baculovirus genesp35 andiap;Mol. Cell. Biol. 14 5212–5222
Cohen J J and Al-Rubeai M 1995 Apoptosis-targeted therapies: the next big thing in biotechnology;Trends Biotechnol. 13 281–283
Cotton J and Spandau D F 1997 Ultraviolet B-radiation dose influences the induction of apoptosis and p53 in human keratinocytes;Radiat. Res. 147 148–155
Crook N E, Clem R J and Miller L K 1993 An apoptosis-inhibiting baculovirus gene with a zinc finger-like motif;J. Virol. 67 2168–2174
Evan G and Littlewood T 1998 A matter of life and cell death;Science,281 1317–1322
Gorman A, McGowan A and Cotter T G 1997 Role of peroxide and Superoxide anion during tumour cell apoptosis;FEBS Lett. 404 27–33
Hershberger P A, Lacount D J and Friesen P D 1994 The apoptotic suppressor p35 is required early during baculovirus replication and is targeted to the cytosol of infected cells;J. Virol. 68 3467–3477
Hockenbery D M, Oltvai Z N, Yin X-M, Milliman C L and Korsmeyer S J 1993 bcl2 functions in an anti-oxidant pathway to prevent apoptosis;Cell 75 241–251
Imlay J A and Linn S 1988 DNA damage and oxygen radical toxicity;Science 240 1302–1309
Ishida Y, Agata Y, Shibahara K and Honjo T 1992 Induced expression of PD-1, a novel member of the immunoglobulin gene superfamily upon programmed cell death;EMBO J. 11 3887–3895
Jain A, Ranjan A, Chatterji U, Das P, Ghosh S, Habib S, Pandey S, Ramachandran A, Venkaiah B and Hasnain S E 1997 High level of expression of heterologous genes in insect cells is not dependent on promoters alone; inInvertebrate cell culture: Novel directions and biotechnology applications (eds) KMarmorosch and JMitsuhashi (New Hampshire: Science Publishers) pp 221–227
Jacobson M D 1996 Reactive oxygen species and programmed cell death;Trends Biochem. Sci. 21 83–86
Kidd M and Emery V C 1993 The use of baculoviruses as expression vectors;Appl. Biochem. Biotech. 42 137–159
Kono Y and Fridovich I 1983 Inhibition and reactivation of Mn-catalse;J Biol. Chem. 258 13646–13648
Laochumroonvorapong P, Paul S, Elkon K B and Kaplan G 1996 Hydrogen peroxide induces monocyte apoptosis and reduces viability ofMycobacterium avium-M. intracellulare within cultured human monocyte;Infect. Immun. 64 452–459
Luckow V A and Summers M D 1988 Trends in the development of baculovirus expression vectors;Biotechnology 6 470–475
Marmorosch K and Mitsuhashi J 1997Invertebrate cell culture: Novel directions and biotechnology applications (New Hampshire: Science Publishers)
Masaki H, Atsumi T and Sakurai H 1995 Detection of hydrogen peroxide and hydroxyl radicals in murine skin fibroblasts under UV-B irradiation;Biochem. Biophys. Res. Commun. 206 474–479
McCarthy M J, Rubin L L and Philpott K L 1997 Involvement of caspases in sympathetic neuron apoptosis;J. Cell. Sci. 110 2165–2173
Nicholson D W 1996 ICE/CED3-like proteases as therapeutic targets for the control of inappropriate apoptosis;Nature Biotechnol. 14 297–301
Nicholson D W and Thornberry N A 1997 Caspases: killer proteases;TIBS 22 299–306
O’Reilly D R, Miller L K and Luckow V A 1992Baculovirus expression vectors: A laboratory manual (New York: WH Freeman)
Robertson N M, Zangrilli J, Fernandes-Alnemri T, Friesen P D, Litwack G and Alnemri E S 1997 Baculovirus p35 inhibits glucocorticoid-mediated pathway of cell death;Cancer Res. 57 43–47
Sambrook J, Fritsch E F and Maniatis T 1989Molecular cloning—A laboratory manual, (New York: Cold Spring Harbor Laboratory)
Sen C K and Packer L, 1996 Antioxidant and redox regulation of gene transcription;FASEB J. 10 709–720
Sridhar P, Awasthi A K, Azim C A, Burma S, Habib S, Jain A, Mukherjee B, Ranjan A and Hasnain S E 1994 Baculovirus vector-mediated expression of heterologous genes in insect cells;J. Biosci. 19 603–614
Steller H 1995 Mechanisms and genes of cellular suicide;Science 267 1445–1449
Thompson C B 1995 Apoptosis in the pathogenesis and treatment of disease;Science 267 1456–1462
Thomberry N A and Lazebnik Y 1998 Caspases: enemies within;Science 281 1312–1316
Villa P, Kaufmann S H and Eamshaw W C 1997 Caspases and caspase inhibitors;TIBS 22 388–393
Wertz I E and Hanley M R 1996 Diverse molecular provocation of programmed cell death;TIBS 21 359–364
White E 1996 Life, death, and the pursuit of apoptosis;Genes Dev. 10 1–15
Yan L-J, Levine R L and Sohal R S 1997 Oxidative damage during aging targets mitochondrial aconitase;Proc. Natl. Acad. Sci. USA 94 11168–11172
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Hasnain, S.E., Taneja, T.K., Sah, N.K. et al. Invitro culturedSpodoptera frugiperda insect cells: Model for oxidative stress-induced apoptosis. J. Biosci. 24, 13–19 (1999). https://doi.org/10.1007/BF02941101
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DOI: https://doi.org/10.1007/BF02941101