A calcium-insensitive attenuated nitrosative stress response contributes significantly in the radioresistance of Sf9 insect cells

作者：Suman Shubhankar; Seth Rakesh Kumar; Chandna Sudhir^*

来源：International Journal of Biochemistry & Cell Biology, 2011, 43(9): 1340-1353.

DOI：10.1016/j.biocel.2011.05.012

• Ionizing radiation
• Insect cell radioresistance
• Apoptosis
• Reactive oxygen species (ROS)
• Nitric oxide synthase (NOS)
• Lepidopteran cell
• Calcium

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 gamma-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 (1000-2000 Gy at 61.1 cGy s(-1)) gamma-irradiation or H2O2 (250 mu M) 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 10 Gy 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.

• 出版日期2011-9