BIOCHEM 2.3 - REACTIVE OXYGEN SPECIES

Reactive Oxygen Species (ROS)

• ROS are oxygen-containing free radicals and reactive molecules generated during normal metabolic processes.
• These molecules can cause damage to cells and contribute to various diseases.
• Excessive ROS can lead to oxidative stress, an imbalance between ROS production and the body's antioxidant defense mechanisms.

Objectives

• Evaluate oxygen free radicals and reactive oxygen species (ROS) formation in human metabolism and disease.
• Identify ROS generated by normal metabolic processes.
• List the steps in ROS generation, including Haber-Weiss and Fenton reactions.
• List reactive oxygen species (ROS) and reactive nitrogen-oxygen species (RNOS) produced by metabolic processes.
• Explain lipid peroxidation steps, cellular localization, consequences, and the role of mitochondria in ROS production.
• Summarize RNOS generation and their role in human systems like inflammation, diabetes, and vasculature.
• Summarize cellular defense mechanisms against oxidative damage, including the roles of various important enzymes and vitamins.

Terminology

• Radical: A molecule with a single unpaired electron in an orbital, capable of independent existence.
• Free radical: A molecule with an unpaired electron.
• Adduct: Two components united via covalent bonds, affecting their form and function and potentially leading to inappropriate accumulation (e.g., vascular accumulation contributing to atherosclerosis).

Reduction of O2

• Oxygen is a biradical molecule with two unpaired electrons, making it highly reactive. In normal metabolic processes, oxygen can accept single electrons typically from reduced electron carriers in the electron transport chain.
• The process generates oxygen radical species.
• Roughly 3–5% of all oxygen consumed is converted to free radicals.
• These reactions generate superoxide, hydrogen peroxide, and hydroxyl radicals.

Oxygen Free Radicals and Reactive Oxygen Species

• Free radicals can be oxygen alone or in compounds.
• They may originate from enzymatic or non-enzymatic processes; they are both accidental byproducts and major products.
• Free radicals initiate chain reactions by stealing electrons to complete orbitals, causing damage to cells.
• ROS damage proteins, DNA, and cellular components, exacerbate cellular damage from other processes, and can result in disease states, as seen in ionizing radiation exposure.
• ROS can result from normal cellular processes, accidents, and exposure to harmful factors.

Oxidative Stress

• Maintaining a balance between ROS production and the body's antioxidant defense mechanisms is crucial.
• Factors that increase ROS: metals such as iron and copper, decompartmentalization, inflammation, certain drugs and xenobiotics, alcohol intake, smoking, and environmental exposures.
• Factors that decrease ROS: antioxidant enzymes, vitamins, small molecules (e.g., glutathione, carnosine), metal sequestration, cellular compartmentalization.

Lipid Peroxidation

• Chain reactions in membranes generate lipid free radicals and lipid peroxides.
• Lipid peroxidation is a major contributor to ROS injury.
• It leads to cellular damage, increased permeability, mitochondrial damage, and oxidation, often targeting sulfhydryl groups and other amino acid residues, thus disrupting cellular processes.
• Oxidizing DNA can damage the cell's genetic material.
• Oxidized DNA fragments, aldehyde production, and reaction with proteins cause extensive damage.

Mitochondria and ROS

• Mitochondria play a major role in generating ATP through electron transport chain processes.
• A by-product of electron transport reactions are semiquinone radicals that lead to superoxide radical generation.

Additional Free Radicals and ROS Including Hypochlorous Acid

• Hypochlorous acid (HOCl) produced by phagocytes is a strong oxidizing agent.
• Nitric oxide (NO) and other reactive nitrogen-oxygen species (RNOS) are involved in several neurodegenerative and chronic inflammatory diseases.
• Nitric oxide (NO) can combine with oxygen (O2) or superoxide (O2−) to form additional reactive nitrogen-oxygen species.
• Other RNOS such as peroxynitrite are also notable for their strong oxidizing properties.

Free Radicals and Inflammation

• Free radicals are deliberately generated during the inflammatory response, often involving rapid oxygen consumption (respiratory burst) by phagocytes.
• NO production along with NADPH oxidase activation damages surrounding tissues, and inflammatory processes lead to damage, particularly in ischemic areas following infarction or similar tissue damage.

ROS in Phagocytosis

• Superoxide is a critical component of phagocytosis, a cellular process involving recognition, engulfment, and destruction of foreign or damaged components.
• Defects in NADPH oxidase can lead to chronic granulomatous disease (CGD), a life-threatening condition.

Reactive Oxygen Species in Disease

• ROS can exacerbate pre-existing conditions or cause new problems related to diseases.
• Diabetes mellitus is associated with increased ROS, leading to vascular endothelial dysfunction.
• Increased xanthine oxidase activity during ischemia-reperfusion leads to further ROS production and damage.

Cellular Defense Against Oxygen Toxicity

• Protective mechanisms include antioxidant enzymes (e.g., superoxide dismutase, catalase, glutathione peroxidase, and reductase), dietary antioxidants (vitamins E, C, carotenoids), endogenous antioxidants (uric acid, protein thiols, and melatonin), and cellular compartmentalization to isolate ROS generation and effects.
• Repair mechanisms are also vital for restoring cellular function after damage.

Ascorbate (Vitamin C)

• Ascorbate (vitamin C) is an antioxidant which can interact with free radicals containing oxygen, and through reactions, regenerate itself. The enzyme dehydroascorbate reductase facilitates this recycling, using reduced glutathione.

Glutathione (GSH)

• Glutathione (GSH) is a major non-protein thiol in cells and a critical cofactor for many antioxidant enzymes.
• It's found in both the mitochondria and cytosol and acts in transport processes within and between cells.

Copper Activity against ROS

• Copper is a key component of superoxide dismutase, requiring covalent incorporation for function.
• Copper/Zinc superoxide dismutase plays a role in increasing the ability of cells to counteract ROS after exposure to reactive molecules.
• In addition to its role in counteracting ROS, Copper/Zinc superoxide dismutase may also function as a transcriptional factor, regulating gene expression involved in oxidative stress repair.

Superoxide Dismutase and Catalase

• Enzymes like superoxide dismutase convert superoxide radicals to less reactive molecules like hydrogen peroxide.
• Catalase further breaks down hydrogen peroxide into water and oxygen, completing the detoxification process.