1. Oxidants and Aging Rolf J. Mehlhorn Lawrence Berkeley Laboratory Rolf J. Mehlhorn Lawrence Berkeley Laboratory

2. Free Radicals  Free radicals are unstable  React quickly with other compounds, doing cell and body damage  Once produced, they multiply unless neutralized by anti- oxidants or other free-radical scavengers.  Free radicals are unstable  React quickly with other compounds, doing cell and body damage  Once produced, they multiply unless neutralized by anti- oxidants or other free-radical scavengers.

3. The Free Radical Theory of Aging “Aging results from the deleterious effects of free radicals produced in the course of cellular metabolism” Harman D., Aging: A theory based on free radical and radiation chemistry, J. Gerontol. 11: 298, 1956 Harman D., Aging: A theory based on free radical and radiation chemistry, J. Gerontol. 11: 298, 1956 “Aging results from the deleterious effects of free radicals produced in the course of cellular metabolism” Harman D., Aging: A theory based on free radical and radiation chemistry, J. Gerontol. 11: 298, 1956 Harman D., Aging: A theory based on free radical and radiation chemistry, J. Gerontol. 11: 298, 1956

4. What is a free radical? FIt is a molecule having unpaired electrons FTherefore, free radicals are unstable FA single molecule can have several unpaired electrons, notably oxygen which can be thought of as a bi-radical FROS = Reactive Oxygen Species FFree radicals rarely occur in nature FMagnetic properties allow for detection

5. Free Radical Chemistry FReactive radicals attack indiscriminately FCan add to unsaturated bonds FCan abstract electrons or hydrogen atoms FPropagate chain reactions FCan cause bond scission FCan cause crosslinking FCrosslinking - Formation of bonds among polymeric chains FProduce secondary toxic agents FReactive radicals attack indiscriminately FCan add to unsaturated bonds FCan abstract electrons or hydrogen atoms FPropagate chain reactions FCan cause bond scission FCan cause crosslinking FCrosslinking - Formation of bonds among polymeric chains FProduce secondary toxic agents

6. Implications for Aging FSome free radical-induced chemical modifications may have unique impacts FCrosslinked products may not be degradable FScission of bonds in DNA, particularly multiple events may erase vital information FSome free radical-induced chemical modifications may have unique impacts FCrosslinked products may not be degradable FScission of bonds in DNA, particularly multiple events may erase vital information

8. Oxygen Toxicity FAt high pressure oxygen is always lethal FConvulsions precede death -- the onset time is a direct function of the oxygen pressure FAnimals have a limited capacity to adapt to oxygen toxicity FMicrobes from anaerobic environments are killed by traces of oxygen FAt high pressure oxygen is always lethal FConvulsions precede death -- the onset time is a direct function of the oxygen pressure FAnimals have a limited capacity to adapt to oxygen toxicity FMicrobes from anaerobic environments are killed by traces of oxygen

9. How can oxygen be toxic? FIt is chemically much more reactive than most other small molecules FIt can be activated by sequential one- electron reductions to superoxide, hydrogen peroxide or hydroxyl radical (ROS) FIt can be converted to reactive singlet oxygen FFor example: Reactive oxygen molecule produced by respiratory burst in immune cells, phagocytes, are toxic to microbial cells FIt is chemically much more reactive than most other small molecules FIt can be activated by sequential one- electron reductions to superoxide, hydrogen peroxide or hydroxyl radical (ROS) FIt can be converted to reactive singlet oxygen FFor example: Reactive oxygen molecule produced by respiratory burst in immune cells, phagocytes, are toxic to microbial cells

10. Immune System Oxidants Activated T-Cell Antigen

11. What are the Major Oxidants? FHydroxyl radical (OH. ) FHypochlorite (HOCl) FSinglet oxygen 1 O 2 FPeroxynitrite (OONO - ) FHydrogen peroxide (H 2 O 2 ) FFree or loosely-bound iron, copper or heme FSuperoxide radical (O 2. - ) FNitric oxide (NO. ) FHydroxyl radical (OH. ) FHypochlorite (HOCl) FSinglet oxygen 1 O 2 FPeroxynitrite (OONO - ) FHydrogen peroxide (H 2 O 2 ) FFree or loosely-bound iron, copper or heme FSuperoxide radical (O 2. - ) FNitric oxide (NO. )

12. Lipid Peroxidation FPUFAs* contain weakly bonded hydrogen atoms between double bonds FChain reactions are probable because of high local concentrations of double bonds FPUFAs* contain weakly bonded hydrogen atoms between double bonds FChain reactions are probable because of high local concentrations of double bonds *PUFAs means polyunsaturated fatty acids

13. Oxidant Sources Table 5.1 FEnzymes involved in cell signaling FImmune cells FEnzymes involved in cell signaling FImmune cells F“Leaky” electron transport FDamaged proteins and lipids FToxins (food, water) FSmoke FIrradiation (UV) F“Leaky” electron transport FDamaged proteins and lipids FToxins (food, water) FSmoke FIrradiation (UV) Regulated Unregulated

14. Major Antioxidants Table 5.2 FVitamins E and C FThiols, particularly glutathione FUric acid FSuperoxide dismutases (Cu/Zn or Mn SOD) FCatalase and glutathione peroxidase FHeme oxygenases FProtein surface groups (Msr) FVitamins E and C FThiols, particularly glutathione FUric acid FSuperoxide dismutases (Cu/Zn or Mn SOD) FCatalase and glutathione peroxidase FHeme oxygenases FProtein surface groups (Msr)

15. Antioxidants FVitamin E FVitamin C FVitamin E FVitamin C

16. Free Radical Damage Markers FFluorescent age pigments FDNA products like hydroxylated bases FProtein products like carbonyls & bityrosine FLipid products like isoprostanes FFluorescent age pigments FDNA products like hydroxylated bases FProtein products like carbonyls & bityrosine FLipid products like isoprostanes

17. Mitochondria FHave been regarded as the “pacemakers of aging” by some investigators FSuperoxide is released as a direct function of the energy state (high ATP/ADP, strongly reducing conditions) FUncoupling proteins dissipate excess energy as heat, are induced by superoxide; some require free fatty acids for activity FHave been regarded as the “pacemakers of aging” by some investigators FSuperoxide is released as a direct function of the energy state (high ATP/ADP, strongly reducing conditions) FUncoupling proteins dissipate excess energy as heat, are induced by superoxide; some require free fatty acids for activity

18. Glucose and Oxidants FIn cell culture models high glucose correlates with oxidant production FThree diabetes-linked effects can be correlated with superoxide production FInsulin pathway and life-span extension FIn cell culture models high glucose correlates with oxidant production FThree diabetes-linked effects can be correlated with superoxide production FInsulin pathway and life-span extension

19. Are Oxidants the Cause of Aging? (Table 5.8) ProCon FCaloric restriction may reduce oxidative stress FLife span extension in mutants may be associated with stress resistance FKnockout mice lacking MnSOD have restricted survival FEnzyme mimetics extend life span in some aging models. FSome drugs, probably acting as antioxidants, have been claimed to extend lifespan. FVitamin C, a superb free radical scavenger, is not synthesized by long-lived primates. FChronic radiation in low doses does not shorten life span (may increase it). FDietary supplementation with Vitamin E and C does not extend life span. FTissue comparison (brain vs. muscle) seems incompatible with oxidant/antioxidant models of aging. FExercise, that increases oxidant stress, improves life span.