H2O2 in physiological and pathological states

1. Reactive oxygen species especially on H2O2
in physiological and pathological conditions
mainly in vascular diseases
Mazen SAEED
M.A student at Pharmacology Department, Ege University 2015

2. REACTIVE OXYGEN SPECIES
• Radicals are species containing one or more
unpaired electrons.
Two types:
1. Free radical such as the oxygen radical
superoxide (O 2
-)
2. The non-radical oxidants such as hydrogen
peroxide (H2O2) and hypochlorous acid
(HOCl) are produced during normal
metabolism and perform several useful
functions.

3. Types of ROS

4. ROS in physiological state
• Intracellular ROS is primarily produced by NADPH
oxidase enzymes (NOXs), the mitochondria, the
endoplasmic reticulum, and the peroxisome.

5. ROS in physiological state
• Cytosolic superoxide (O2 _) is rapidly converted into
hydrogen peroxide (H2O2) by superoxide dismutase 1
(SOD1).
• H2O2 can either act as a signaling molecule by oxidizing
critical thiols within proteins to regulate numerous
biological processes, including metabolic adaptation,
differentiation, and proliferation or be detoxified to
water (H2O) by the scavenging enzymes peroxiredoxin
(PRX), glutathione peroxidase (GPX), and catalase (CAT).

6. ROS in physiological state
• In addition, H2O2 can react with metal cations (Fe2+
or Cu+) to generate the hydroxyl radical (OH _),
which causes irreversible oxidative damage to lipids,
proteins, and DNA

8. HYDEROGEN PEROXIDE IN PHYSIOLOGIC STATE

9. HYDEROGEN PEROXIDE
• H2O2 is produced as a by-product of protein
oxidation in the endoplasmic reticulum (ER), as an
end product in numerous peroxisomal oxidation
pathways such as in the beta-oxidation of very long-
chain fatty acids, and by a wide range of enzymes
including cytochrome P450.

10. HYDEROGEN PEROXIDE
• H2O2 act as signaling molecules is through the
oxidation of critical cysteine residues within redox-
sensitive proteins.

11. H2O2-mediated cysteine oxidation of
redox-sensitive proteins mechanism
• Susceptible cysteine residues have a low pKa and
exist as a thiolate anion (S _) at physiological pH,
making them more reactive than the protonated
cysteine thiol group (SH) and providing a level of
selectivity and specificity .

12. H2O2-mediated cysteine oxidation of redox-
sensitive proteins mechanism
• H2O2 oxidization of the thiolate anion to the sulfenic
form (SO _) impinges on cellular signaling by altering
protein conformation and activity. In the presence of
high concentrations of H2O2, SO _ is further oxidized
to form sulfinic (SO2 _) and sulfonic (SO3 _) acids
(i.e. hyperoxidation), where SO3 _ generally
represents an irreversible oxidative modification.

13. H2O2-mediated cysteine oxidation of redox-
sensitive proteins mechanism
• To prevent irreversible cysteine oxidation, the SO _
intermediate is com-monly incorporated into a
disulfide (S–S) or sulfenic- amide (S–N) bond. These
modifications are reversible by the actions of
glutaredoxin (GRX) and thioredoxin (TRX), which
restore protein function; the oxidized protein is
returned to its reduced state.

14. H2O2-mediated cysteine oxidation of redox-
sensitive proteins

15. Possible mechanisms for H2O2-dependent
signal transduction
• H2O2 can oxidize critical cysteine thiol groups of
phosphatases including PTEN, PTP1B, and MAPK
opened up the possibility that H2O2 serve as
signaling molecules
• There are many mechanisms explined that including
redox relay, floodgate model and others.

16. Redox relay
• The redox relay mechanism uses a scavenging
enzyme such as glutathione peroxidase (GPX) or
peroxiredoxin (PRX) to transduce the H2O2 signal
and oxidize the target protein

17. Floodgate model
• H2O2 inactivates the scavenger, perhaps through
hyperoxidation to sulfinic (SO2 _) acid or through a
posttranslational modification (PTM), to allow for
H2O2-mediated oxidation of the target protein.

18. Other mechanisms
• The scavenging enzyme accepts H2O2 oxidation and
transfers the oxidation to an intermediate redox
protein such as thioredoxin (TRX), which
subsequently oxidizes the target protein.

19. Other mechanisms
• Dissociation of the target protein from the oxidized
scavenging enzyme results in target protein
activation.

20. Oxidative stress
• A condition of imbalance between pro-
oxidant (ROS) and anti-oxidant as defined
by Sies in 1985

21. Hydrogen peroxide-induced oxidative
stress
• Hydrogen peroxide (H2O2) is a cell-permeant and highly
stable reactive oxygen species (ROS)
• H2O2 produced mainly by the dismutation of
superoxide anion (O2 .-) by superoxide dismutase and
also directly by the NOX-4 isoform of the NAD(P)H
oxidase .
• H2O2 levels are regulated by intracellular and
extracellular enzymes including catalase, glutathione
peroxidase, thioredoxin and other peroxyredoxins,
which convert H2O2 to water and O2(1)

22. The vascular functional effects of H2O2
• Are very complex and depend among other factors, of the
specific vascular bed studied and the nature of the
precontractile agent.(1)
• H2O2 can either contract or relax arteries from different
species (in resting state or after preconstriction with
stimuli(1)
• According to an other studies that indicated the same
things “H2O2 can act as a vasodilator and/or
vasoconstrictor depending on the vascular bed, species,
and experimental conditions”.(2)

23. Mechanisms whereby H2O2 get its
effects on vessels
H2O2 induces vasoconstriction by :-
• Stimulation of cyclooxygenase and release of TXA2 or
TXA2 analogue U46619
• Activation of c-Src(c-Src mediates H2O2 contractile
responses by increase of TP activation, H2O2-induced
TXA2 production)

24. Mechanisms whereby H2O2 get its
effects on vessels
• Activation of ERK1/2 mitogen activated protein
kinases (MAPK)
• Activation of Rho kinase(1)
• Activation of phospholipase A2 and phospholipase
C ,pathways (2).

25. Mechanisms whereby H2O2 get its
effects on vessels

26. Vasorelaxation Effects of H202
• The vasoactive role of H2O2 in endothelium-
dependent vasorelaxation was recently recognized
to involve a multiplicity of responses.
• H2O2 is considered as a primar EDHF, because it is
produced by endothelial cells and causes vascular
smooth muscle relaxation through activation of
Ca2+-dependent K+ (KCa) channels

27. Mechanisms whereby H2O2 get its effects on vessels
1. Activation of potassium channels
pathway
H2O2 can activates different potassium
channels in vascular smooth muscle cells by
produce membrane hyperpolarization . (3)

28. Activation of potassium channels pathway
• There are also conflicting results about the types of
potassium channels involved in relaxant effect of
H2O2 in different vascular tissues

29. Type of vessels References Subtypes of Ca+2 channel involved
in H2O2 vessel relaxation
Cat cerebral arteries Wei EP, Kontos HA, Beckman JS. Mechanisms
of cerebral
vasodilation by superoxide, hydrogen
peroxide, and peroxynitrite.
Am J Physiol 1996;271:H1262–6.
ATPdependent
potassium channels
Rat cerebral arteries Sobey CG, Heistad DD, Faraci FM. Mechanisms
of bradykinininduced
cerebral vasodilatation in rats: evidence that
reactive
oxygen species activate K+ channels. Stroke
1997;28:2290–5.
Ca2+-activated potassium channels
Canine cerebral arteries Iida Y, Katusic ZS. Mechanisms of cerebral
arterial relaxations to
hydrogen peroxide. Stroke 2000;31:2224–
3220.
Ca2+-activated potassium channels
and voltage-dependent potassium
Channel
Mouse mesenteric Matoba T, Shimokawa H, Nakashima M,
Hirakawa Y, Mukai Y,
Hirano K, et al. Hydrogen peroxide is an
endothelium-derived
hyperpolarizing factor in mice. J Clin Invest
2000;106:1521–30.
Ca2+-activated potassium channels
Porcine coronary arteries Barlow RS, White RE. Hydrogen peroxide
relaxes porcine
coronary arteries by stimulating BKCa channel
activity. Am J
Physiol 1998;275:H1283–9.
Ca2+-activated potassium channels

30. Human atrial Miura H, Bosnjak JJ, Ning G, Saito T, Miura M.
Gutterman DD.
Role of hydrogen peroxide in flow-induced
dilation of human
coronary arterioles. Circ Res 2003;92:e31–40.
Ca2+-activated potassium
channels
Pig coronary arterioles. Thengchaisri N, Kuo L. Hydrogen peroxide
induces endotheliumdependent
and -independent coronary arteriolar dilation:
role of
cyclooxygenase and potassium channels. Am J
Physiol 2003;285:
H2255–63.
Ca2+-activated potassium
channels
Human ITA voltage-dependent potassium
channel
Canine coronary arteries Rogers PA, Dick GM, Knudson JD, Focardi M,
Bratz IN,
Swafford AN, et al. H2O2-induced redox-
sensitive coronary
vasodilation is mediated by 4-aminopyridine-
sensitive K+ channels.
Am J Physiol 2006;291:H2473–82.
voltage-dependent potassium
channel
Rat coronary arteries Rogers PA, Dick GM, Knudson JD, Focardi M,
Bratz IN,
Swafford AN, et al. H2O2-induced redox-
sensitive coronary
vasodilation is mediated by 4-aminopyridine-
sensitive K+ channels.
Am J Physiol 2006;291:H2473–82.
voltage-dependent potassium
channel

31. 2. Activation of COX pathway
H2O2 activate COX pathway starting with the activates
phospholipase A2 to release arachidonic acid from
vascular smooth muscle cells and produces COX
products .This products may have role in the relaxation
effect of H2O2.(3)

32. 3. Producation of nitric oxide (NO):
• It have been reported that H2O2 improve release of many
reporte endothelial nitric oxide (NO) and accumulate of
cyclic GMP in smooth muscle resulting in relaxation effect
of H2O2.(4)

33. 1. GARCÍA-REDONDO, Ana B., et al. c-Src, ERK1/2 and Rho kinase mediate hydrogen
peroxide-induced vascular contraction in hypertension: role of TXA2, NAD (P) H
oxidase and mitochondria. Journal of hypertension, 2015, 33.1: 77-87.
2. HATOUM, Ossama A., et al. Role of hydrogen peroxide in ACh-induced dilation of
human submucosal intestinal microvessels. American Journal of Physiology-Heart and
Circulatory Physiology, 2005, 288.1: H48-H54.
3. NACITARHAN, Cahit, et al. The effect of hydrogen peroxide in human internal thoracic
arteries: role of potassium channels, nitric oxide and cyclooxygenase
products. Cardiovascular Drugs and Therapy, 2007, 21.4: 257-262.
4. YANG, Zhi-wei, et al. Endothelium-dependent relaxation to hydrogen peroxide in
canine basilar artery: a potential new cerebral dilator mechanism. Brain research
bulletin, 1998, 47.3: 257-263.
5. Reczek, Colleen R., and Navdeep S. Chandel. "ROS-dependent signal
transduction." Current opinion in cell biology 33 (2015): 8-13.
REFERENECES