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Reactive oxygen species & signal transduction 2

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Reactive oxygen species & signal transduction 2

  1. 1. PRESENTED BY SHANOO SUROOWAN Reactive oxygen species & signal transduction
  2. 2. <ul><li>A radical is an atom/ group of atoms having one/more unpaired electrons. </li></ul><ul><li>There are many types of radicals, but the most prominent in biological systems are derived from oxygen collectively known as Reactive Oxygen Species (ROS), </li></ul><ul><li>Oxygen in its ground state has 2 unpaired electrons........Remember? O 8 : 1s 2 2s 2 2p 4 </li></ul><ul><li>So it is easy for Oxygen to accept electrons to form free radicals (Reactive Oxygen Species in this case!) </li></ul>
  3. 3. ROS formation in cells <ul><li>These are formed as normal aerobic processes that occur in the body; some as necessary intermediates of enzymatic reactions. </li></ul><ul><li>Most are produced in the ETC when oxygen is reduced to water in the mitochondria. </li></ul><ul><li> O 2 reduction H 2 O </li></ul><ul><li>During this conversion various reactive oxygen species are formed! </li></ul><ul><li>ē ē ē ē </li></ul><ul><li>O 2 . O 2 – H2O2 . OH - H2O </li></ul><ul><li>(oxygen) (superoxide) (hydrogen peroxide) (hydroxyl ion) (water) </li></ul>
  4. 4. Overproduction of ROS <ul><li>Neutrophils are specialized in ROS formation which destroy pathogens as part of host defence. </li></ul><ul><li>When cells are exposed to abnormal environments such as hypoxia and hyperoxia. </li></ul><ul><li>From ionizing radiation in biological systems. Ionizing radiation will ionize molecular oxygen by pushing an electron in its outer orbit. </li></ul><ul><li>If oxygen species are overproduced it will be difficult for the cell to detoxify them and repair the damages they make! Result: Oxidative stress! </li></ul><ul><li>So Oxidative stress is an imbalance btw reactive oxygen species and antioxidants! </li></ul>
  5. 5. Harmful activities of ROS <ul><li>Damage to a number of macromolecules such as lipids, proteins and DNA caused by ROS implicate them in many disease processes, ranging from arthritis, atherosclerosis, pulmonary fibrosis, cancer, neurodegenerative diseases, and aging </li></ul><ul><li>Toxic effects such as damage to cell membranes initiated by lipid peroxidation. </li></ul><ul><li>A common target for peroxidation is unsaturated fatty acids present in membrane phospholipids </li></ul>
  6. 6. Lipid peroxidation.... <ul><li>Consequences of lipid peroxidation: </li></ul><ul><ul><li>increased membrane rigidity </li></ul></ul><ul><ul><li>decreased activity of membrane-bound enzymes (e.g. sodium pumps ) </li></ul></ul><ul><ul><li>altered activity of membrane receptors. </li></ul></ul><ul><ul><li>altered permiability </li></ul></ul>
  7. 7. Antioxidants of the body <ul><li>Intracellular antioxidants: </li></ul><ul><ul><li>vitamin E </li></ul></ul><ul><ul><li>ascorbate(vitamin C) </li></ul></ul><ul><ul><li>glutathione (glutamate-cysteine-glycine) [GSH] </li></ul></ul><ul><li>Enzymatic antioxidants: </li></ul><ul><ul><li>Superoxide dismutases(SOD’s) </li></ul></ul><ul><ul><li>Catalase </li></ul></ul><ul><ul><li>Gluthione peroxidase </li></ul></ul>
  8. 8. Intracellular antioxidants <ul><li>Vitamin E; a lipid soluble antioxidant that traps peroxy radicals while doing so it itself becomes a radical </li></ul><ul><li>Vitamin C regenerates back vitamin E from its radical form </li></ul>
  9. 9. Enzymatic antioxidants <ul><li>Superoxide dismutases (SOD’s) </li></ul><ul><ul><li>Mostly found in the mitochondria </li></ul></ul><ul><ul><li>They depend on cofactors such as manganese, copper or zinc for their antioxidant activity. </li></ul></ul><ul><ul><li>They convert 2 superoxide ions into oxygen and hydrogen peroxide </li></ul></ul><ul><li>Catalase hydrolyses hydrogen peroxide into water and oxygen! </li></ul><ul><ul><li>Site of location; peroxisomes </li></ul></ul><ul><li>Glutathione peroxidase also hydrolyze hydrogen peroxide and can convert organic peroxides to alcohol </li></ul>
  10. 10. <ul><li>SIGNAL TRANSDUCTION </li></ul>
  11. 11. <ul><li> Extracellular Signal </li></ul><ul><li>(growth factor/cytokines/neurotransmitter/hormone) </li></ul><ul><li> Binds to specific Receptor </li></ul><ul><li>Interaction of receptor-ligand complex </li></ul><ul><li>Generates a wide variety of intracellular signals: </li></ul><ul><li>1. Changes in ion concentration </li></ul><ul><li>2. Activation of trimeric GTP binding regulatory proteins </li></ul><ul><li>3. Activation of receptor kinases </li></ul><ul><li>Downstream signaling by secondary messengers </li></ul><ul><li>(cAMP, Ca2+, phospholipid metabolites) </li></ul><ul><li>Activation of transcription factors for transcription of </li></ul><ul><li> specific genes for diverse cellular functions </li></ul>
  12. 12. Oxidants and signal transduction <ul><li>Oxidants modulate cell signaling events by modifying cell surface receptors, phosphatases and protein phosphorylation, etc . </li></ul><ul><li>These phenomena are important in transactivation of transcription factors </li></ul><ul><li>activation/inactivation of gene transcription that may regulate steps in the development of disease. </li></ul>
  13. 13. ROS and Signal Transduction <ul><li>ROS are important mediators in signal transduction </li></ul><ul><li> Receptor + hormone </li></ul><ul><li> (upstream signaling) </li></ul><ul><li> Receptor-ligand complex </li></ul><ul><li> Intracellular ROS production </li></ul><ul><li> (downstream signaling) </li></ul><ul><li>activates </li></ul><ul><li> Other pathways </li></ul><ul><li>promote regions of intermediate response genes governing cell proliferation, differentiation, etc . </li></ul><ul><ul><li>ROS are involved in both upregulation and downregulation pathways! </li></ul></ul>
  14. 14. ROS and Signal Transduction <ul><li>ROS are not only injurious by-products of cellular metabolism but also essential participants in cell signaling and regulation </li></ul><ul><li>The cellular functions/toxic properties of ROS is dependent on their concentration. </li></ul><ul><li>For e.g. when produced in low concentrations by nitric oxide synthase(NOS) NO· functions as a signaling molecule mediating vasodilation </li></ul><ul><li>While when produced in high concentrations in macrophages, it is a toxic oxidant for microbicidal killing </li></ul>
  15. 15. Cellular sources and regulation of ROS <ul><li>Cellular production of ROS occurs from both enzymatic and nonenzymatic sources </li></ul><ul><li>any electron-transferring protein or enzymatic system can result in the formation of ROS as “by-products” </li></ul><ul><li>ROS are mostly produced in the mitochondria , H2O2 can diffuse out in the cytoplasm while O2.- remains trapped in. </li></ul><ul><li>O 2 − ·-generating microsomal NADH oxidoreductase may function as a potential pulmonary artery O 2 sensor in pulmonary artery smooth muscle cells </li></ul>
  16. 16. Cellular sources and regulation of ROS <ul><li>Nuclear membranes contain cytochrome oxidases and ETC. Electron “leaks” from these enzymatic systems gives rise to ROS that can damage cellular DNA </li></ul><ul><li>Peroxisomes are an important source of total cellular H 2 O 2 production. They contain a number of H 2 O 2 -generating enzymes including glycolate oxidase, d-amino acid oxidase. </li></ul><ul><li>Peroxisomal catalase utilizes H 2 O 2 produced by these oxidases to oxidize a variety of other substrates in “peroxidative” reactions e.g. detoxification of alcohol in the liver </li></ul><ul><li>Intracellular soluble enzymes such as xanthine oxidase , aldehyde oxidase , can generate ROS during catalytic cycling </li></ul>
  17. 17. Cellular sources and regulation of ROS <ul><li>Autooxidation of small molecules such as dopamine, epinephrine can be an important source of intracellular ROS production. </li></ul><ul><li>Prooxidant effects of dopamine autooxidation is implicated in the pathogenesis of neurodegenerative diseases such as Parkinson's disease </li></ul><ul><li>Plasma membrane-associated oxidases have been implicated as the sources of most growth factor- and/or cytokine-stimulated oxidant production </li></ul><ul><li>The phagocytic NADPH oxidase, which serves a specialized function in host defense against invading microorganisms </li></ul>
  18. 18. Cellular sources and regulation of ROS <ul><li>functional components of the phagocytic NADPH are present in nonphagocytic cells. </li></ul><ul><li>P22 phox is a component of NADPH oxidase and plays a key role in its activation </li></ul><ul><li>Expression of p22 phox has been demonstrated in the adventitial smooth muscle cells of coronary arteries and the aorta </li></ul><ul><li>Increased aortic adventitial O 2 − · production contributes to hypertension by blocking the vasodilatory effects of NO </li></ul><ul><li>ROS production in coronary arteries is related to hypertension </li></ul>
  19. 19. ROS in cell signaling
  20. 20. ROS signaling by receptors <ul><li>The following receptors are involved in ROS signaling :- </li></ul><ul><ul><li>Cytokine receptors </li></ul></ul><ul><ul><li>Receptor tyrosine kinases (RTKs) </li></ul></ul><ul><ul><li>Receptor serine/threonine kinases </li></ul></ul><ul><ul><li>G-protein coupled receptor </li></ul></ul><ul><ul><li>These receptors will generate intracellular signals for ROS production </li></ul></ul>
  21. 21. ROS signaling by cytokine receptors <ul><li>1.TNF- α </li></ul><ul><li>Mediate ROS formation in mitochondria </li></ul><ul><li>Activates the TF nuclear factor (NF)-kB </li></ul><ul><li>nuclear factor (NF)-kB dependent transcription </li></ul><ul><li>Makes cancer cells resistant to apoptosis </li></ul><ul><li>Activates apoptosis signal-regulating kinase-1 (ASK 1) </li></ul><ul><li> How? </li></ul><ul><li>Oxidant dependent dimerization of ASK 1 </li></ul><ul><li>ASK1 has been found to be involved in cancer, diabetes, cardiovascular and neurodegenerative diseases </li></ul>
  22. 22. <ul><li>TNF- α is an autocrine cytokine involved in ROS signaling by cytokine receptors (upstream transduction) </li></ul><ul><li>The ROS produced have various implications in downstream signaling such as:- </li></ul><ul><ul><li>the expression of cell adhesion molecules from genes </li></ul></ul><ul><ul><li>production of chemokines </li></ul></ul><ul><ul><li>In pathophysiological conditions such as induction of cardiac myocyte hypertrophy </li></ul></ul><ul><li>2.IFN- γ </li></ul><ul><li>(activator of the phagocytic NADPH oxidase) </li></ul><ul><li>Stimulates cyclooygenase-dependent peroxide production in human hepatocyte </li></ul><ul><li>Resistance to bacteria </li></ul>
  23. 23. ROS signaling by RTKs <ul><li>A number of growth factors that bind to RTKs generate intracellular ROS essential for mitogenic signaling :- </li></ul><ul><ul><li>Mitogens will be produced </li></ul></ul><ul><ul><li>Produced mitogens activate MAPK ( Mitogen activated protein kinase) </li></ul></ul><ul><ul><li>that trigger mitosis </li></ul></ul><ul><li>These growth factors that act on receptor tyrosine kinases (RTKs) include :- </li></ul><ul><ul><li>PDGF (plasma dependent growth factor) </li></ul></ul><ul><ul><li>EGF (epidermal growth factor) </li></ul></ul><ul><ul><li>FGF (fibroblast growth factor) </li></ul></ul>
  24. 24. PDGF <ul><li>Increases intracellular concentrations of hydrogen peroxides </li></ul><ul><li>Induce tyrosine phosphorylation </li></ul><ul><li>MAPK activation </li></ul><ul><li>DNA synthesis & chemotaxis </li></ul><ul><li>Regulates gene expression by . O 2 – dependent pathways </li></ul><ul><li>. O 2 – produced </li></ul><ul><li> involved in the upregulation of inducible NOS & NO dependent release of PGE2 in fibroblasts </li></ul><ul><li> induces fever </li></ul>
  25. 25. Receptor serine/threonine kinases <ul><li>These are receptors of the transforming growth factor β (TGF- β ) superfamily </li></ul><ul><li>TGF- β </li></ul><ul><li>Stimulates extracellular production of ROS </li></ul><ul><li>Regulates a number of physiological actions </li></ul><ul><li>Apoptosis, collagen synthesis, growth inhibitory effects </li></ul>
  26. 26. G protein-coupled receptors <ul><li>The ligands for these receptors include:- </li></ul><ul><ul><li>ANG II (Angiotensin II) </li></ul></ul><ul><ul><li>Serotonin (5-hydroxytryptamine) </li></ul></ul><ul><ul><li>Bradykinin </li></ul></ul><ul><ul><li>Thrombin </li></ul></ul>
  27. 27. <ul><li>Angiotensin II </li></ul><ul><li>activates both NADH- and NADPH-dependent O 2 − · production in vascular smooth muscle cells </li></ul><ul><li>A variety of physiological actions of ANG II are mediated by ROS </li></ul><ul><li>its vasopressor activity, smooth muscle cell hypertrophy, activation of cell survival PK Akt/PKB, induction of insulin-like growth factor-1 receptor </li></ul>
  28. 28. Mechanisms of ROS action <ul><li>ROS act via two mechanisms:- </li></ul><ul><li>alterations in intracellular redox state </li></ul><ul><li>2 ) oxidative modifications of protein s </li></ul>
  29. 29. Alterations in intracellular redox states <ul><li>The cytosol is maintained under strong reducing conditions </li></ul><ul><li>This is accomplished by the “redox-buffering” capacity of intracellular thiols, primarily glutathione (GSH)and thioredoxin (TRX). </li></ul><ul><li>They reduce both H 2 O 2 and lipid peroxides, reactions that are catalyzed by peroxidases </li></ul><ul><li>e.g. GSH peroxidase catalyzes the reaction H 2 O 2 + 2GSH -> 2H 2 O + GSSG </li></ul><ul><li>GSH and TRX are antioxidants that play important roles in cell signaling </li></ul>
  30. 30. Alterations in intracellular redox states <ul><li>During oxidative stress , the concentration of the oxidized form of GSH increases </li></ul><ul><li>[H 2 O 2 + 2GSH -> 2H 2 O + GSSG] </li></ul><ul><li>Decreased cell proliferation in vascular endothelial cells </li></ul><ul><li> increased proliferation of fibroblasts </li></ul><ul><li> induces the binding of some TFs to DNA. </li></ul>
  31. 32. Conclusion <ul><li>ROS are mediators of cell signaling </li></ul><ul><li>They cause a series of changes during cell signaling </li></ul>
  32. 33. References <ul><li>http://www.jleukbio.org/content/65/3/337.full.pdf </li></ul><ul><li>http://www.sciencedirect.com/science/article/pii/S0014579300016690 </li></ul><ul><li>http://jcb.rupress.org/content/194/1/7 </li></ul><ul><li>http://www.heribert-hirt.info/pdf/prr85_ros.pdf </li></ul><ul><li>http://ajplung.physiology.org/content/279/6/L1005.full </li></ul>

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