Antioxidant for subfertility their role, highlights on generic, trade name and studies relevant

1. www.themegallery.com

2. Presentation By
M.A.Wadood Aref

3.  Infertility or Subfertility is defined as
the failure to conceive after 1 year of
regular, unprotected intercourse with
the same partner.
 10–17% of all couples experience
primary or secondary infertility at some
time during their reproductive life.
M.A.Wadood Aref

4.  Male factor is considered a major contributory
factor to infertility in 50% of cases, with no
identifiable cause in 25%.
 Along with the conventional causes for male
infertility such as varicocele, cryptorchidism,
infections, obstructive lesions, cystic fibrosis,
trauma, and tumors, a new, yet important
cause has been identified: oxidative stress.
M.A.Wadood Aref

5.  male factor
Idiopathic
 Idiopathic
25%  tubal conditions
male factor
40%
 ovulation disorders
tubal  age related
conditions  Endometriosis
15%
 Coital Disorders
Coital disorders
ovulation age
5%
disorders related Endometriosis
15% 10% 10%
Total % exceeds 100% as 15% of couples have more
than one cause of Infertility.

6. Minority (of cases) Majority
Known cause Idiopathic
Specific Empirical
medical treatment medical treatment

7. 1. Gonadotrophins
2. Dopamine Agonists
3. Anti-estrogens
4. Aromatase Inhibitors
5. Androgens
M.A.Wadood Aref

8.  Includes Anti-oxidants or Combination therapy
 Indications
1. Idiopathic infertility: in an attempt to improve
semen parameters and subsequent fertility potential.
2. Known untreatable causes of infertility, such as
following surgery for cryptorchidism or torsion &
some cases of testicular failure.
3. known potentially treatable causes which failed to
adequately respond to specific treatments.
M.A.Wadood Aref

9. Cell destruction by ROS
Reactive oxygen species (ROS)

10.  Cells
living under aerobic conditions
require oxygen to support life.
 Free radicals are highly reactive
molecules with one or more
unpaired electrons that can
oxidatively modify biomolecules.
M.A.Wadood Aref

11.  Reactive oxygen species (ROS) are free radicals
that are derived from the metabolism of oxygen.
 Major ROS in seminal plasma include:
 superoxide (O2).
 hydrogen peroxide (H2O2).
 hydroxyl radical (OH).
 Male germ cells generate ROS at low physiologic
levels to regulate sperm capacitation, acrosome
reaction and sperm–oocyte fusion
M.A.Wadood Aref

12.  ROS React almost immediately with any
substance in surrounding beginning a chain
reaction leading to cellular damage.
 In healthy men, a delicate balance exists
between physiological ROS and antioxidants
in the male reproductive tract
 High levels of seminal ROS have been found
in 30 - 80% of infertile men.
M.A.Wadood Aref

13. Balance status between ROS and antioxidants
Oxidative stress (OS)

14.  Oxidative stress is a condition that
occurs when there is an imbalance
between ROS production and
antioxidant capacity (ROS exceed
antioxidant capacity).
 In 1943, John MacLeod first made the
observation that oxidative stress could
be a significant cause of male infertility.
M.A.Wadood Aref

16. High ROS level in subfertile men
Reduced Defective
Sperm Oocyte-sperm
Motility Increased fusion
Cell membrane
Sperm DNA Lipid
Damage
peroxidation

17. Oxidative stress can be measured
Evaluation of oxidative stress

18.  Quantification of the level of oxidative
stress is important to determine if it is a
significant contributor to sperm
dysfunction in a given patient.
 Despite the different methods available
to measure ROS, currently no standard
exists for estimating oxidative stress.
M.A.Wadood Aref

19.  oneof the most widely used
methods to assess the level of ROS.
 discriminates between the
production of superoxide and
hydrogen peroxide by spermatozoa
by the reagent used.
M.A.Wadood Aref

20.  uses the principles of light
scattering, light excitation, and
emission of fluorochrome molecules
to generate specific multiparameter
data measures.
M.A.Wadood Aref

21.  determines relation between total
amount of ROS and the total
antioxidant capacity to measure
ultimate amount of oxidative stress and
subsequent damage.
 determined by either the enhanced
chemiluminescence assay or the
calorimetric assay.
M.A.Wadood Aref

23. Treatment of infertility caused by
oxidative stress

24. 1. Behaviors and lifestyle modification may
decrease endogenous ROS production.
2. Antioxidant supplements increase the
antioxidant capacity of semen, thereby
scavenging excess ROS and decreasing OS.
3. Specific laboratory techniques used to
decrease ROS production (when sperm are
handled in the laboratory for ART).
M.A.Wadood Aref

25. Antioxidant kick out free radicals in blood
Antioxidants

26. Antioxidants in seminal plasma
Prevent
Affection of Prevent DNA
Sperm fragmentation
Motility
Prevent
Cryo-damage

27.  Human body uses 3 systems of
antioxidants that constantly scavengers
(inactivate) ROS:
1. Endogenous antioxidants.
2. Metal-binding proteins.
3. Dietary antioxidants.
M.A.Wadood Aref

28. Enzymes Vitamins
Vitamin E
Superoxide Fat-soluble
dismutase (SOD) Vitamin A
Water Vitamin C
Catalase
soluble
Glutathione
peroxidase
M.A.Wadood Aref

29.  Metal-binding proteins include:
1. Albumin.
2. Cerruloplasmin.
3. Metallo-thionein.
4. Transferrin.
5. Ferritin.
6. Myoglobulin.
 They inactivate the transition metal ions
(e.g, iron) that catalyze the production
of free radicals.

30.  present in fruits and vegetables and
daily dietary supplements.
 The mechanism of action of the dietary
antioxidants includes:
 scavenging free radicals.
 chain-breaking effect of lipid pero-
xidase enzyme.
M.A.Wadood Aref

31. 1. Carnitine 6. N-acetyl cysteine
2. vitamin C 7. CoEnzyme Q10
3. vitamin E 8. Pentoxyfilline
4. Carotenoids 9. Zinc
(vitamin A) 10.Selenium
5. Glutathione
M.A.Wadood Aref

32. Carnitine
L-Carnitine & L-acetyl carnitine
Daily requirement: 200 - 500 mg
Oral supplemntation dose:
L-Carnitine 1 gm 1 x2 or 1x3 (3 mnths)
Found in: red meat & dairy products
Available in Egypt

33.  Water soluble Dietary antioxidant (75%
derived from diet).
 decreases ROS by removing
extracellular toxic acetyl-CoA that is
responsible for mitochondrial ROS.
 Stabilizes spermatozoal membrane.
M.A.Wadood Aref

34.  The highest concentration of
carnitine occurs in the epididymis
(2000-fold that of plasma).
 50% of carnitine In seminal fluid
exists as acetyl-carnitine.
M.A.Wadood Aref

35.  Some studies showed beneficial effect.
Study Parameters
Moncada et al., 1992 Improved sperm motility
Costa et al., 1994 Improved sperm motility and count
Vitali et al., 1995 Improved sperm motility in 60%
Lenzi et al., 2003 Improved sperm motility , no effect on
morphology
Lenzi et al., 2004 Improved sperm motility , no effect on
morphology
Cavallini et al., 2004 Improved sperm morphology

36.  Other studies showed no effect.
Study Parameters improved
Sigman et al., 2006 L-carnitine and L-acetyl carnitine
Lenzi et al., 2004 no effect on motility, morphology
or count
M.A.Wadood Aref

37. Vitamin E
Daily requirement:
50 - 800 mg /d.
Oral supplemntation dose:
400 or 600 mg /d
.
Found in: fats and oils (olive, and sunflower oils)
Rx: R/ Vitamin E 400 mg tab 1 x 1 (available in Egypt)

38.  Vitamin E which is present within
the cell membrane, is one of the
major membrane protectants
against ROS.
 It neutralizes hydrogen peroxide
and protects the plasma membrane
from lipid peroxidation.
M.A.Wadood Aref

39. Study Parameters improved
Ross et al., 2010 decrease ROS
Comhaire et al, 2000 vit.E, vit.A, acetyl cysteine,
decreased ROS
Suleiman et al., 1996 improving motility and the
probability of achieving pregnancy
Kessopoulou et al, decrease ROS, improved zona-binding assay
1995a
de Lamirande & protect spermatozoa from OS and loss of motility
Gagnon, 1992
Dawson et al., 1987 protect spermatozoa from OS during
cryopreservation and thawing

40. Vitamine C
Daily requirement: 60 mg /d
Oral supplemntation dose
200-1000 mg /d
Found in: citrous fruits, vegetables, fish
Rx: R/ Vitamin C 500 mgcaps. 1X1 available in Egypt

41.  Vitamin C (ascorbic acid) is a water-soluble,
highly potent antioxidant.
 concentrations 10-fold higher in seminal plasma
than serum.
 It also has been shown to recycle vitamin E by
repairing its tocopheroxyl radical, thereby
allowing vitamin E to function as a free radical
chain-breaking antioxidant
 The hydrophilicity and lipophilicity of vitamins C
and E may act synergistically to protect against
peroxidative attack on spermatozoa
M.A.Wadood Aref

42.  Some studies showed beneficial effect.
Study Parameters improved
Greco et al., 2005a reduction in DNA-fragmented
spermatozoa
Greco et al., 2005b improvement in pregnancy
and implantation rates
Kodama et al, 1997 improved concentration,
decreased DNA damage
M.A.Wadood Aref

43.  Other studies showed no effect.
Study Parameters
Rolf et al, 1999 no improvement in
Kessopoulou et al., 1995b • concentration,
• motility,
Moilanen & Hovatta,1995
• morphology, or
Giovenco et al., 1987 • viability
M.A.Wadood Aref

44. Carotenoids
(beta-carotene and lycopene)
Daily requirement: 1000 mg / d
Oral supplemntation dose:
Lycopene 5-10 mg /d
Found in: tomatoes, watermelon, guava.
available in Egypt

45.  Beta-carotenes protect the plasma
membrane against lipid peroxidation.
 Lycopene is twice as potent as beta-carotene
and ten times more potent than vitamin E in
scavenging singlet oxygen and inhibiting lipid
peroxidation in serum plasma
 Astaxanthin, a newly studied carotenoid
extracted from the algae Hemaococcus
pluvialis.

46. Study Parameters improved
Comhaire et al., 2005 improved pregnancy rates
Gupta & Kumar, 2002 improved sperm concentration
& motility
M.A.Wadood Aref

47. Glutathione (GSH)
essential nutrient synthesized
in the body (not obtained via food)
Oral supplemntation dose:
600 mg /d IM
IM form is not available in Egypt

48. Glutathione (GSH)
 one of the most common antioxidant &
reducing agent in the body.
 It plays an important role in protecting lipids,
proteins, and nucleic acids against oxidative
damage.
 It combines with vitamin E and selenium to
form glutathione peroxidase (the main
enzyme involved in removing H2O2 in the
epididymis).
M.A.Wadood Aref

49. N-acetyl cysteine (NAC)
Derived from Cysteine amino acid
Oral supplemntation dose: 30 mg /d
Should be given with IM Glutathione

50.  NAC is a precursor that replenishes
Glutathione .
 scavenges free radicals and reduces ROS
production in human ejaculate.
 NAC plays an important role in germ cell
survival in human seminiferous tubules in
vitro.
M.A.Wadood Aref

51. Study Parameters improved
Comhaire et al., 2000 NAC improved sperm concentration
and acrosome reaction & reduced ROS
Oeda et al., 1997 improved sperm motility
Lenzi et al., 1993 improved sperm motility (forward
progression)
M.A.Wadood Aref

52. Zinc
Daily requirement: 8 - 11 mg/d
Oral supplemntation dose:
Zinc Gluconate 50 mg /d
Found in: oysters, wheat, beans
available in Egypt only with multivitamins

53.  Adequate zinc intake is needed to
maintain the optimal functioning level
of antioxidant enzymes, such as
superoxide dismutase.
 Seminal plasma zinc concentrations is
significantly lower in subfertile men
 Zinc deficiency is associated with
abnormal flagella .

54. Study Parameters improved
Ross et al, 2010 progressive motility, sperm integrity and
pregnancy rates
Omu et al., 2008 Decreased apoptosis & DNA fragmentation
Wong et al., 2002 Increased total sperm count
Omu et al., 1999 Improved sperm concentration
Tikkiwal et al., 1987 Improved sperm concentration
Hartoma et al., 1977 Improved sperm concentration
M.A.Wadood Aref

55. Selenium
Daily requirement: 55 mg/d
Oral supplemntation dose:
225 mcg /d
Found in: Garlic, seeds, meat, eggs.
available in Egypt

56.  Selenium is a trace element
necessary for the synthesis of
glutathione peroxidase.
 selenium acts with Vitamin E
synergistically as antiperoxidants.
M.A.Wadood Aref

57. Study Parameters improved
Kesker-Ammar et al., 2003 improved motility
Vezina et al., 1996 improved motility but not
pregnancy rates
M.A.Wadood Aref

58. Co-enzyme Q 10
(Ubiquinol)
Daily requirement: 3 -6 mg /d
Oral supplemntation dose: 30 mg /d
Found in: meat and fishes
available in Egypt

59.  an energy-promoting agent in the
mitochondria of sperm mid-piece.
 CoQ10 recycles vitamin E and prevents
its pro-oxidant activity.
 The reduced form of CoQ10, ubiquinol,
also acts as an antioxidant preventing
lipid peroxidation.
M.A.Wadood Aref

60. Study Parameters improved
Thomas et al., 1997 improve fertilisation rate
Alleva et al., 1997 inhibit hydrogen peroxide ROS
M.A.Wadood Aref

61. Pentoxifylline
Synthetic Drug
Oral supplemntation dose: 400 mg 1x3
available in Egypt

62.  competitive non-selective phospho-
diesterase inhibitor that raises
intracellular cAMP, so improves
testicular micro-circulation.
 decrease ROS production.
 preserve sperm motility in vitro
 improve semen parameters in vivo.
M.A.Wadood Aref

63. Study Parameters improved
Okada et al., 1997 decreased ROS and preserved sperm
motility in vitro
Pang et al., 1993 preserve sperm motility in vitro
Tesarik et al., 1992 Improved sperm motility but not %
Gavella & Lipovac, decrease ROS
1992
Gavella et al., 1991 decrease ROS
Yovich et al., 1990 improve semen parameters in vivo
Marrama et al., 1985 improve semen parameters in vivo

64. Antioxidant role in vitro
In Vitro Antioxidant supplements

65.  In vitro supplements of antioxidants
used during sperm preparation and ART
(assisted reproductive technique)
1. Neutralizes ROS produced by
leukocytes and immature
spermatozoa
2. Improves sperm-oocyte fusion.
M.A.Wadood Aref

66.  In vitro supplementation with
superoxide dismutase and catalase leads
to:
1. prevention of lipid peroxidation of the
sperm plasma membrane caused by
ROS.
2. recovery of high-quality spermatozoa
after freezing-thawing procedures.
M.A.Wadood Aref

67. Antioxidant role in female reproductive pathologies
Antioxidants for Females

69.  OS is implicated in pathological
processes of the female reproductive
tract that contribute to infertility and
poor pregnancy outcome.
 Antioxidants are used in order to
improve fertility and pregnancy rates.
M.A.Wadood Aref

70.  PCOS: N-acetyl cysteine improve glucose
control and peripheral insulin sensitivity in
hyperinsulinemic patients.
 Endometriosis: The antioxidants catalase,
RU486, curcumin, melatonin, and
catechins have anti-proliferative and anti-
angiogenic effects stopping disease
progression.
M.A.Wadood Aref

71.  Unexplained infertility: Selenium
normalize patient’s RBC Mg levels and
can result in clinical pregnancy after 8
months of treatment.
 recurrent pregnancy loss: folic acid,
melatonin, and omega-3 fatty acids are
now investigated (particularly in
antiphospholid antibody syndrome).
M.A.Wadood Aref

72. Antioxidant kick out free radicals
Take Home Message

73.  Spermatozoa are under a continuous
influence of OS because of excessive
generation of ROS but natural
antioxidants decrease the progression
of the damage.
M.A.Wadood Aref

74.  However, when an imbalance exists
between levels of ROS and the natural
antioxidant defenses, various
measures can be used to protect
spermatozoa against the OS-induced
injury .
M.A.Wadood Aref

75.  For patients suspected to have high
levels of ROS and Oxidative stress
(OS), antioxidant supplements can
be considered.
M.A.Wadood Aref

76.  Oxidativestress can be also
reduced by modifying certain
lifestyle behaviors and
environmental factors.
M.A.Wadood Aref

77.  Another important method for
decreasing OS is the use of
antioxidants during various sperm
processing techniques (ARTs).
M.A.Wadood Aref

78.  Combined therapy is much more
beneficial because antioxidants act
by different mechanisms on different
free radicals.
M.A.Wadood Aref

79.  Although further studies are required
to validate use of antioxidants, The
low cost and relatively low risk of
toxicity of the antioxidants is
appealing to both patients and
clinicians.
M.A.Wadood Aref

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