title: A calcium-insensitive attenuated nitrosative stress response contributes significantly in the radioresistance of Sf9 insect cells.
abstract: Lepidopteran insects/insect cells display 50-100 times higher radioresistance than humans, and are evolutionarily closest to mammals amongst all radioresistant organisms known. Compared to mammalian cells, Lepidopteran cells (TN-368, Sf9) display more efficient antioxidant system and DNA repair and suffer considerably less radiation-induced DNA/cytogenetic damage and apoptosis. Recent studies indicate that a considerably lower radiation-induced oxidative stress may significantly reduce macromolecular damage in Lepidopteran cells. Since nitrosative stress contributes in radiation-induced cellular damage, we investigated its nature in the γ-irradiated Sf9 cells (derived from Spodoptera frugiperda; order Lepidoptera; family Noctuidae) and compared with BMG-1 human cell line having significant NOS expression. Radiation induced considerably less ROS/RNS in Sf9 cells, which remained unchanged on treatment with NOS inhibitor l-NMMA. Surprisingly, growth of Sf9 cultures or irradiation could not induce NO or its metabolites, indicating negligible basal/radiation-induced NOS activity that remained unchanged even after supplementation with arginine. Cytosolic calcium release following high-dose (1000-2000Gy at 61.1cGys(-1)) γ-irradiation or H(2)O(2) (250μM) treatment also failed to generate NO in Sf9 cells having high constitutive levels of calmodulin, whereas BMG-1 cells displayed considerable calcium-dependent NO generation even following 10Gy dose. These results strongly imply the lack of calcium-mediated NOS activity in Sf9 cells. Addition of exogenous NO from GSH-NO caused considerable increase in radiation-induced apoptosis, indicating significant contribution of constitutively attenuated nitrosative stress response into the radioresistance of Lepidopteran cells. Our study demonstrates for the first time that a calcium-insensitive, attenuated nitrosative stress response may contribute significantly in the unusual radioresistance displayed by Lepidopteran insect cells.
authors.affiliation: Natural Radiation Response Mechanisms Group, Division of Radiation Biosciences, Institute of Nuclear Medicine & Allied Sciences (INMAS), Brig. S.K. Mazumdar Road, Delhi 110054, India.
authors.forename: Shubhankar
authors.initials: S
authors.lastname: Suman
authors.forename: Rakesh Kumar
authors.initials: RK
authors.lastname: Seth
authors.forename: Sudhir
authors.initials: S
authors.lastname: Chandna
mesh_terms: D000818:Animals; D001120:Arginine / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000528:radiation effects; D002147:Calmodulin / Q000378:metabolism; D002460:Cell Line; D006427:Hemin / Q000494:pharmacology; D006801:Humans; D019476:Insect Proteins / Q000378:metabolism; D007915:Lepidoptera / Q000166:cytology* / Q000201:enzymology / Q000528:radiation effects; D053078:Membrane Potential, Mitochondrial; D009566:Nitrates / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism; D009573:Nitrites / Q000378:metabolism; D018384:Oxidative Stress / Q000528:radiation effects; D011836:Radiation Tolerance; D026361:Reactive Nitrogen Species / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism; D019323:omega-N-Methylarginine / Q000494:pharmacology
chemical_list: D002147:Calmodulin; D019476:Insect Proteins; D009566:Nitrates; D009573:Nitrites; D026361:Reactive Nitrogen Species; D017382:Reactive Oxygen Species; D019323:omega-N-Methylarginine; D009569:Nitric Oxide; D006427:Hemin; D001120:Arginine; D019001:Nitric Oxide Synthase; D002118:Calcium