Metabolic Toxins: Reactive Oxygen Species - In-Depth Notes

Page 2: Learning Objectives

• At the end of this lecture, you will be able to:
  • Describe reactive oxygen species and oxidative damage
  • List the importance of antioxidants within the body
  • Describe how antioxidants prevent cellular damage

Page 3: Case Study on Hyperlipidemia

• Overview:
  Hyperlipidemia can lead to atherosclerosis, especially when lipoproteins become oxidized. This case study will serve as a contextual backdrop as we discuss reactive oxygen species (ROS).

Page 4: Understanding Reactive Oxygen Species

• Key Characteristics of Oxygen:

  • Oxygen is essential for life but can also be toxic.
  • Oxygen contains two single electrons in different orbitals, both with the same spin—this configuration makes it a “biradical.”

• Reactive Nature:

  • These two electrons do not easily oxidize organic compound bonds due to “spin restriction” which typically requires an enzyme to assist.
  • Upon receiving an extra electron, oxygen becomes a radical, significantly increasing its reactivity.

• Definition of Radicals:

  • Radicals are atoms with an unpaired electron in their outer valence shell.

Page 5: Generation of Radicals

• Enzymatic Reactions:
  • Radicals often form during enzymatic reactions as enzymes facilitate the transfer of electrons, creating “free radicals” which can exist independently.
  • When free radicals interact with other compounds, they can initiate chain reactions, causing extensive cellular damage.
  • Proportion of ROS Generated:
  • 3-5% of consumed oxygen is converted into ROS.

Page 6: Sources of ROS: Electron Transport Chain (ETC)

• Role in ATP Production:

  • The mitochondrial electron transport chain (ETC) produces ATP by transferring electrons between carriers and complexes until they reach oxygen.

• Superoxide Formation:

  • Occasionally, an electron can escape the ETC (usually from Coenzyme Q), forming superoxide, a type of ROS.

Page 7: Sources of ROS: Ionizing Radiation

• Effects of Radiation:
  • Ionizing radiation (like X-rays) can create ROS from water.
  • Ultraviolet (UV) radiation leads to ROS formation in skin cells.

Page 8: Sources of ROS: Drug Metabolism

• Role of Cytochrome P450 Enzymes:
  • These enzymes metabolize drugs (including alcohol), oxidizing substrates to enhance excretion.
  • Unlike some enzymes, these can be “leaky,” allowing radical intermediates to escape and form free radicals, leading to cell damage.

Page 9: Sources of ROS: Inflammation

• Inflammatory Response:
  • ROS play a role in eliminating invading pathogens and cleaning damaged tissues during inflammation.
  • In activated neutrophils, the “respiratory burst” consumes oxygen to generate reactive substances that kill bacteria, but this may also damage nearby tissues.

Page 10: Reactive Nitrogen-Oxygen Species (RNOS)

• Understanding RNOS:
  • RNOS, which are also free radicals, can damage cellular components, including DNA and cell membranes.
  • Sources of RNOS may include dietary elements and environmental pollution, highlighting their external origins in addition to internal generation.

Page 11: ROS Damage

• Mechanism of Damage:
  • ROS can inflict damage on nearly every cellular component, triggering chain reactions that perpetuate oxidative damage.
  • For example, if a free radical pulls an electron from a stable molecule, that molecule becomes a radical and can further damage others, effectively creating a cascade of harmful effects.

Page 12: Example of DNA Damage

• DNA Vulnerability:

  • ROS can cause strand breaks or various alterations in DNA that may result in mutations.

• Specific Alteration:

  • One example is the oxidation of guanine to 8-hydroxyguanine, which may lead to faulty base pairing if not repaired.

Page 13: Consequences of 8-Hydroxyguanine

• Base Pairing Issue:
  • 8-hydroxyguanine can base pair incorrectly with adenine instead of cytosine, resulting in a G-C pair replicating as a T-A pair, leading to mutations.
  • DNA repair mechanisms can correct some alterations, but missed repairs may lead to an accumulation of mutations over time.

Page 14: Preventing Mutation from 8-Hydroxyguanine

• Question: Which mechanism could prevent a permanent mutation when guanine is converted to 8-hydroxyguanine?
  • a) 3’-5’ exonuclease in DNA polymerase
  • b) Base excision repair system
  • c) Telomerase
  • d) DNA ligase

Page 15: Cellular Defense Mechanisms

• Defense Against Free Radicals:
  • Cells harness various mechanisms to combat the effects of free radicals via:
  • Enzymes that neutralize radicals
  • Antioxidants that donate electrons but do not turn into radicals themselves

Page 16: Protective Enzymes and Proteins

• Enzyme Functions:
  • Enzymes like superoxide dismutases (SOD) and catalases exist in various cellular compartments.
  • Glutathione, a peptide made from glycine, cysteine, and glutamate, serves to remove hydrogen peroxide generated outside peroxisomes, using minerals like Cu, Zn, Mn, Fe, and Se.

Page 17: Antioxidants

• Role of Vitamins:
  • Vitamins can function as antioxidants:
  • Vitamin E (fat-soluble) is present in plasma membranes and can donate electrons to radicals.
  • Vitamin C (water-soluble) is located in blood and cytoplasm, regenerating Vitamin E’s antioxidant capacity by donating electrons.
  • Carotenoids from the diet also exhibit antioxidant properties.

Page 18: Overview of Cellular Defense Mechanisms

• Summary:
  • The body has multiple defenses against ROS/RNOS, and excessive levels are associated with various diseases.

Page 19: Connection to Cancer

• Cancer and ROS:
  • Various carcinogens, including DMBA and NNK, lead to DNA damage, contributing to mutations in oncogenes and tumor suppressor genes, exacerbated by factors like oxidative stress.
  • The progression from normal cells to initiated cells involves single-strand and double-strand breaks leading to increased genetic instability and potential for transformation into cancerous cells.

Page 20: Sources of ROS - Quiz Question

• Question: Which of the following is a source of reactive oxygen species?
  • a) Neutrophils recruited during inflammation
  • b) Superoxide dismutase
  • c) DNA repair enzymes
  • d) Urea from protein breakdown

Page 21: Preventing Mitochondrial DNA Damage - Quiz Question

• Question: Which enzyme could mitigate damage to mitochondrial DNA, where oxidative damage levels are significantly higher compared to nuclear DNA?
  • a) Cytochrome P450 reductase
  • b) Superoxide dismutase (SOD)
  • c) Xanthine oxidase
  • d) Lipid peroxidase

Page 22: Case Study Recap

• Hyperlipidemia Impact:
  • Atherosclerosis is associated with the oxidation of specific lipoproteins.
  • Understanding the oxidation processes is crucial in this case study.

Page 23: Lipoprotein and Atherosclerosis Question

• Question: Which lipoprotein is deposited in artery intimal layers and contributes to atherosclerosis upon oxidation?
  • a) Chylomicrons
  • b) VLDL
  • c) LDL
  • d) HDL

Page 24: Oxidation Process Question

• Question: Which process contributes to the oxidation of lipoproteins in atherosclerosis?
  • a) The electron transport chain (ETC)
  • b) Inflammation
  • c) Pollution
  • d) UV radiation

Page 25: References

• Chikara, S. et al. (2018). Oxidative stress and dietary phytochemicals: Role in cancer chemoprevention and treatment. Cancer Letters, 413, 122–134. https://doi.org/10.1016/J.CANLET.2017.11.002
• D’Augustin, O. et al. (2020). Lost in the Crowd: How Does Human 8-Oxoguanine DNA Glycosylase 1 (OGG1) Find 8-Oxoguanine in the Genome? International Journal of Molecular Sciences, 21, 8360. https://doi.org/10.3390/IJMS21218360
• Lieberman, M. & Peet, A. (2023). MARKS’ Basic Medical Biochemistry: A Clinical Approach (6th Edition). Wolters Kluwer Health.
• Smolin, L. A. et al. (2020). Nutrition: Science and Application (3rd Canadian Edition). John Wiley & Sons Canada.