The eff ects of extenders containing proline and glutamine on oxidative stress and motion parameters of stallion semen during cold storage

Document Type : Research Articles

Authors

Shahrekord University of Shahrekord

Abstract

This study examined the effects of skim-milk based extenders supplemented with proline and glutamine on motility, lipid peroxidation and enzymatic antioxidant status of cooled-stored equine sperm and determined the role of seminal plasma as well. The semen was collected with artificial vagina. In experiment 1, native semen was diluted in skim-milk based extender containing 5mM glutamine and 3mM proline, stored at 5°C and analyzed at 4, 24 and 48 hours storage for motion parameters. In experiment 2, semen was centrifugated, sperm pellet resuspended in the extenders and stored at 5°C for 4 hours to determine motion parameters. The level of catalase, glutathione peroxidase activity and malondialdehyde formation was determined for all samples at 4 hours. Glutamine and proline signifi cantly preserved the percentage of motile sperm (76.5 ± 2.7 and 79.4 ± 1 vs 69 ± 1.4), increased the progressive motility of cold-stored semen (66.1 ± 2.5 and 73.7 ± 2.9 vs 56.2 ± 1.4), increased catalase activity, and decreased malondialdehyde. However these effects were disappeared after seminal plasma removal. We conclude that glutamine and proline would amplify the antioxidant activity of equine cold-stored semen and preserve its motility. This effect seems to be related to interactions with seminal plasma.

Keywords

Main Subjects


1. Aurich J, Aurich C. Developments in European horse breeding
and consequences for veterinarians in equine reproduction.
Reproduction in Domestic Animals. 2006;41:275-9.
2. Aitken RJ. Pathophysiology of human spermatozoa. Current
opinion in obstetrics & gynecology. 1994;6:128-35.
3. Aurich C. Factors aff ecting the plasma membrane function of
cooled-stored stallion spermatozoa. Animal Reproduction
Science. 2005;89:65-75.
4. Squires E, Barbacini S, Matthews P, Byers W, Schwenzer K,
Steiner J, et al. Retrospective study of factors aff ecting fertility
of fresh, cooled and frozen semen. Equine Veterinary Education.
2006;18:96-9.
5. Chatterjee S, Gagnon C. Production of reactive oxygen species
by spermatozoa undergoing cooling, freezing, and thawing.
Molecular Reproduction and Development: Incorporating
Gamete Research. 2001;59:451-8.
6. Ball BA. Oxidative stress, osmotic stress and apoptosis: impacts
on sperm function and preservation in the horse. Animal reproduction
science. 2008;107:257-67.
7. Ball B, Medina V, Gravance C, Baumber J. Eff ect of antioxidants
on preservation of motility, viability and acrosomal integrity
of equine spermatozoa during storage at 5 C. Th eriogenology.
2001;56:577-89.
8. Pickett B, Sullivan J, Byers W, Pace M, Remmenga E. Eff ect of
centrifugation and seminal plasma on motility and fertility
of stallion and bull spermatozoa. Fertility and sterility.
1975;26:167-74.
9. Baas JW, Molan P, Shannon P. Factors in seminal plasma of bulls
that aff ect the viability and motility of spermatozoa. Journal
of reproduction and fertility. 1983;68:275-80.
10. Jasko D, Moran D, Farlin M, Squires E. Eff ect of seminal plasma
dilution or removal on spermatozoal motion characteristics
of cooled stallion semen. Th eriogenology. 1991;35:1059-67.
11. Varner D, Blanchard T, Love C, Garcia M, Kenney R. Eff ects
of semen fractionation and dilution ratio on equine spermatozoal
motility parameters. Th eriogenology. 1987;28:709-23.
12. Love C, Brinsko S, Rigby S, Th ompson J, Blanchard T, Varner
D. Relationship of seminal plasma level and extender
type to sperm motility and DNA integrity. Th eriogenology.
2005;63:1584-91.
13. Backman T, Bruemmer J, Graham J, Squires E. Pregnancy rates
of mares inseminated with semen cooled for 18 hours and
then frozen. Journal of animal science. 2004;82:690-4.
14. Melo C, Zahn F, Martin I, Orlandi C, Dell’Aqua Jr J, Alvarenga
MA, et al. Infl uence of semen storage and cryoprotectant on
post-thaw viability and fertility of stallion spermatozoa. Journal
of equine veterinary science. 2007;27:171-5.
15. Kareskoski M, Katila T. Components of stallion seminal plasma
and the eff ects of seminal plasma on sperm longevity. Animal
reproduction science. 2008;107:249-56.
16. Purdy P. A review on goat sperm cryopreservation. Small Ruminant
Research. 2006;63:215-25.
17. Kobayashi T, Miyazaki T, Natori M, Nozawa S. Protective role
of superoxide dismutase in human
sperm motifi ty: superoxide dismutase
activity and lipid peroxide in human
seminal plasma and spermatozoa.
Human Reproduction. 1991;6:987-91.
18. Kankofer M, Kolm G, Aurich J, Aurich
C. Activity of glutathione peroxidase,
superoxide dismutase and catalase
and lipid peroxidation intensity in
stallion semen during storage at 5 C.
Th eriogenology. 2005;63:1354-65.
19. Crockett E, Graham J, Bruemmer J,
Squires E. Eff ect of cooling of equine
spermatozoa before freezing on postthaw
motility: preliminary results.
Th eriogenology. 2001;55:793-803.
20. Pellicer-Rubio M, Combarnous Y.
Deterioration of goat spermatozoa
in skimmed milk-based extenders
as a result of oleic acid released by
the bulbourethral lipase BUSgp60.
Journal of reproduction and fertility.
1998;112:95-105.
21. Khalili B, Jafaroghli M, Farshad A,
Paresh-Khiavi M. 43 the eff ects of
diff erent concentrations of Glycine
and cysteine on the freezability
of Moghani ram spermatozoa.
Asian-Australasian Journal of Animal
Sciences. 2010;23:318.
22. Bilodeau J-F, Blanchette S, Gagnon
C, Sirard M-A. Th iols prevent
H2O2-mediated loss of sperm motility
in cryopreserved bull semen. Th eriogenology.
2001;56:275-86.
23. Bucak MN, Tuncer PB, Sarıözkan S,
Ulutaş PA. Comparison of the eff ects
of glutamine and an amino acid solution
on post-thawed ram sperm parameters,
lipid peroxidation and anti-
oxidant activities. Small Ruminant
Research. 2009;81:13-7.
24. Amirat-Briand L, Bencharif D, Vera-
Munoz O, Ali HBH, Destrumelle
S, Desherces S, et al. Eff ect of glutamine
on post-thaw motility of bull
spermatozoa aft er association with
LDL (low density lipoproteins) extender:
preliminary results. Th eriogenology.
2009;71:1209-14.
25. Martins-Bessa A, Rocha A, Mayenco-
Aguirre A. Incorporation of taurine
and hypotaurine did not improve
the effi ciency of the Uppsala Equex
II extender for dog semen freezing.
Th eriogenology. 2007;68:1088-96.
26. El-Sheshtawy R, El-Sisy G, El-Nattat
W. Use of selected amino acids to
improve buff alo bull semen cryopreservation.
Global Veterinaria.
2008;2:146-50.
27. Renard P, Grizard G, Griveau J-F,
Sion B, Boucher D, Le Lannou D.
Improvement of motility and fertilization
potential of postthaw human
sperm using glutamine. Cryobiology.
1996;33:311-9.
28. Trimeche A, Renard P, Le Lannou D,
Barriere P, Tainturier D. Improvement
of motility of post-thaw Poitou jackass
sperm using glutamine. Th eriogenology.
1996;45:1015-27.
29. Aitken R, West K, Buckingham D.
Leukocytic infi ltration into the human
ejaculate and its association with
semen quality, oxidative stress, and
sperm function. Journal of andrology.
1994;15:343-52.
30. Davoodian N, Kadivar A, Ahmadi
E, Mohebbi A. Eff ects of Two Amino
Acids on Motion Parameters and
Enzymatic Antioxidant Activity of
Freeze-Th awed Stallion Spermatozoa.
Journal of Equine Veterinary Science.
2017;59:49-56.
31. Trimeche A, Yvon J, Vidament M,
Palmer E, Magistrini M. Eff ects of
glutamine, proline, histidine and betaine
on post-thaw motility of stallion
spermatozoa. Th eriogenology.
1999;52:181-91.
32. Khlifaoui M, Battut I, Bruyas JF, Chatagnon
G, Trimeche A, Tainturier D.
Eff ects of glutamine on post-thaw
motility of stallion spermatozoa: an
approach of the mechanism of action
at spermatozoa level. Th eriogenology.
2005;63:138-49.
33. Brinsko S, Crockett E, Squires E. Eff ect
of centrifugation and partial removal
of seminal plasma on equine spermatozoal
motility aft er cooling and storage.
Th eriogenology. 2000;54:129-36.
34. Jasko D, Hathaway J, Schaltenbrand V,
Simper W, Squires E. Eff ect of seminal
plasma and egg yolk on motion characteristics
of cooled stallion spermatozoa.
Th eriogenology. 1992;37:1241-
52.
35. Oliveira RAd, Piersanti RL, Wolf CA,
Viu MAdO, Gambarini ML. Glutathione
for the freezing of cooled equine
semen, using diff erent protocols.
2014.
36. Oliveira R, Duarte F, Gambarini M.
Comparison between the conventional
and automated systems for frozen
cooled equine semen. ANIMAL REPRODUCTION.
2015;12:324-7.
37. Begley A, Quinn P. Decapacitation factors
in semen. Clinical reproduction
and fertility. 1982;1:167-75.
38. Watson P. Th e causes of reduced fertility
with cryopreserved semen. Animal
reproduction science. 2000;60:481-
92.
39. Moore A, Squires E, Graham J. Eff ect
of seminal plasma on the cryopreservation
of equine spermatozoa. Th eriogenology.
2005;63:2372-81.
40. Katila T, Andersson M, Reilas T, Koskinen
E. Post-thaw motility and viability
of fractionated and frozen stallion
ejaculates. 2002.
41. Lopes S, Jurisicova A, Sun J-G, Casper
RF. Reactive oxygen species: potential
cause for DNA fragmentation in human
spermatozoa. Human Reproduction
(Oxford, England). 1998;13:896-
900.
42. Potts R, Notarianni L, Jeff eries T. Seminal
plasma reduces exogenous oxidative
damage to human sperm, determined
by the measurement of DNA
strand breaks and lipid peroxidation.
Mutation Research/Fundamental and
Molecular Mechanisms of Mutagenesis.
2000;447:249-56.
43. Leboeuf B, Restall B, Salamon S. Production
and storage of goat semen for
artifi cial insemination. Animal reproduction
science. 2000;62:113-41.
44. Roy A. Egg yolk-coagulating enzyme
in the semen and Cowper’s gland of
the goat. Nature. 1957;179:318.
45. Mann T, Lutwak-Mann C. Male reproductive
function and the composition
of semen: general considerations.
Male Reproductive Function and Semen:
Springer; 1981. p. 1-37.
46. Aitken RJ. Free radicals, lipid peroxidation
and sperm function. Reproduction,
Fertility and Development.
1995;7:659-68.
47. Filho IB, Pederzolli C, Sgaravatti A,
Gregory R, Filho CD, Jobim M, et
al. Skim milk-egg yolk based semen
extender compensates for non-enzymatic
antioxidant activity loss during
equine semen cryopreservation.
Anim Reprod. 2009;6:392-9.
48. Sangeeta S, Arangasamy A, Kulkarni
S, Selvaraju S. Role of amino acids as
additives on sperm motility, plasma
membrane integrity and lipid peroxidation
levels at pre-freeze and postthawed
ram semen. Animal reproduction
science. 2015;161:82-8.
49. Rudolph AS, Crowe JH. Membrane
stabilization during freezing: the
role of two natural cryoprotectants,
trehalose and proline. Cryobiology.
1985;22:367-77.
50. Pagl R, Aurich C, Kankofer M. Antioxidative
Status and Semen Quality
during Cooled Storage in Stallions.
Transboundary and Emerging Diseases. 2006;53:486-9.
51. Baumber J, BALL BA, GRAVANCE
CG, Medina V, DAVIES‐MOREL MC.
Th e eff ect of reactive oxygen species
on equine sperm motility, viability,
acrosomal integrity, mitochondrial
membrane potential, and membrane
lipid peroxidation. Journal of andrology.
2000;21:895-902.
52. Lalonde RJ, Lepock JR, Kruuv J. Site
of freeze-thaw damage and cryoprotection
by amino acids of the calcium
ATPase of sarcoplasmic reticulum.
Biochimica et Biophysica Acta
(BBA)-Protein Structure and Molecular
Enzymology. 1991;1079:128-38.
53. Anchordoguy T, Carpenter JF, Loomis
SH, Crowe JH. Mechanisms of interaction
of amino acids with phospholipid
bilayers during freezing. Biochimica
et Biophysica Acta (BBA)-Biomembranes.
1988;946:299-306.
54. Buege JA, Aust SD. [30] Microsomal
lipid peroxidation. Methods in enzymology:
Elsevier; 1978. p. 302-10.
55. Lawrence RA, Burk RF. Glutathione
peroxidase activity in selenium-defi -
cient rat liver. Biochemical and biophysical
research communications.
1976;71:952-8.
56. Goth L. A simple method for determination
of serum catalase activity and
revision of reference range. Clinica
chimica acta. 1991;196:143-51.
CAPTCHA Image