title: ROS dependent copper toxicity in Hydra-biochemical and molecular study.
abstract: Copper, an essential microelement, is known to be toxic to aquatic life at concentrations higher than that could be tolerated. Copper-induced oxidative stress has been documented in vitro, yet the in vivo effects of metal-induced oxidative stress have not been extensively studied in the lower invertebrates. The objective of the present study has been to find the effect of ROS-mediated toxicity of environmentally relevant concentrations of copper at organismal and cellular levels in Hydra magnipapillata. Exposure to copper at sublethal concentrations (0.06 and 0.1mg/L) for 24 or 48h resulted in generation of significant levels of intracellular reactive oxygen species (ROS). We infer that the free radicals here originate predominantly at the lysosomes but partly at the mitochondria also as visualized by H2-DHCFDA staining. Quantitative real-time PCR of RNA extracted from copper-exposed polyps revealed dose-dependent up-regulation of all antioxidant response genes (CAT, SOD, GPx, GST, GR, G6PD). Concurrent increase of Hsp70 and FoxO genes suggests the ability of polyps to respond to stress, which at 48h was not the same as at 24h. Interestingly, the transcript levels of all genes were down-regulated at 48h as compared to 24h incubation period. Comet assay indicated copper as a powerful genotoxicant, and the DNA damage was dose- as well as duration-dependent. Western blotting of proteins (Bax, Bcl-2 and caspase-3) confirmed ROS-mediated mitochondrial cell death in copper-exposed animals. These changes correlated well with changes in morphology, regeneration and aspects of reproduction. Taken together, the results indicate increased production of intracellular ROS in Hydra on copper exposure.
authors.affiliation: Mahatma Gandhi-Doerenkamp Center, Bharathidasan University, Tiruchirappalli 620024, India; Department of Environmental Biotechnology, Bharathidasan University, Tiruchirappalli 620024, India.
authors.forename: Mohammed
authors.initials: M
authors.lastname: Zeeshan
authors.affiliation: Mahatma Gandhi-Doerenkamp Center, Bharathidasan University, Tiruchirappalli 620024, India; Department of Environmental Biotechnology, Bharathidasan University, Tiruchirappalli 620024, India.
authors.forename: Anbazhagan
authors.initials: A
authors.lastname: Murugadas
authors.affiliation: Developmental Biology Group, MACS-Agharkar Research Institute, Pune 411004, India.
authors.forename: Surendra
authors.initials: S
authors.lastname: Ghaskadbi
authors.affiliation: Department of Environmental Biotechnology, Bharathidasan University, Tiruchirappalli 620024, India.
authors.forename: Ramasamy Babu
authors.initials: RB
authors.lastname: Rajendran
authors.affiliation: Mahatma Gandhi-Doerenkamp Center, Bharathidasan University, Tiruchirappalli 620024, India; Department of Food Science and Nutrition, College of Food and Agriculture, King Saud University, Riyadh, Saudi Arabia. Electronic address: mgdcaua@yahoo.in.
authors.forename: Mohammad Abdulkader
authors.initials: MA
authors.lastname: Akbarsha
mesh_terms: D000818:Animals; D000975:Antioxidants / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D015153:Blotting, Western; D020552:Comet Assay; D019327:Copper Sulfate / Q000633:toxicity*; D004249:DNA Damage; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation; D006829:Hydra / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D007928:Lethal Dose 50; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D012038:Regeneration / Q000187:drug effects; D012098:Reproduction / Q000187:drug effects; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors; D014874:Water Pollutants, Chemical / Q000633:toxicity*
chemical_list: D000975:Antioxidants; D051017:Apoptosis Regulatory Proteins; D017382:Reactive Oxygen Species; D014874:Water Pollutants, Chemical; D019327:Copper Sulfate
keywords: Apoptosis; Copper; Genotoxicity; Hydra; Oxidative stress; ROS; Regeneration