اثر افزودن گلوتاتیون احیا به رقیق کننده بر پایه نانوذره لسیتین بر انجمادپذیری اسپرم گاو

نوع مقاله : مقاله پژوهشی

نویسندگان

1 دانشجوی دکتری، دانشگاه تهران، پردیس کشاورزی کرج

2 دانشیار،پردیس کشاورزی و منابع طبیعی، دانشگاه تهران، تخصص: فیزیولوژی دام

3 استاد،پردیس کشاورزی و منابع طبیعی، دانشگاه تهران، تخصص: فیزیولوژی دام

4 استاد، گروه بیوشیمی،دانشگاه تهران

5 دانشیار،پردیس کشاورزی و منابع طبیعی، دانشگاه تهران، تخصص: فیزیولوژی تولیدمثل

6 استادیار،البرز. کرج. دانشکده کشاورزی دانشگاه تربیت مدرس

چکیده

هدف از این پژوهش ، بررسی اثر افزودن گلوتاتیون احیا به رقیق­کننده بر پایه نانوذره لسیتین بر کیفیت اسپرم گاو پس از فرایند انجماد و یخ‌گشایی بود به همین منظور چهار سطح آنتی­اکسیدان گلوتاتیون (صفر، یک، 2/5 و پنج میلی­مولار) در رقیق­کننده بر پایه نانوذرات لسیتین ارزیابی شد. اندازه ذرات لیستین تهیه شده با استفاده از دستگاه سونیکاتور به ابعاد نانو کاهش یافت. 48 انزال طی سه هفته از شش رأس گاو نر هلشتاین با میانگین سنی سه تا چهار سال جمع­آوری و منجمد شد. صفات مورد ارزیابی پس از انجماد و یخ‌گشایی شامل، فراسنجه­های جنبایی با CASA، فعالیت غشای سلول­ (هاست) و یکپارچگی غشا (ائوزین- نگروزین) ریخت‌شناختی اسپرم (محلول هانکوک) بود. نتایج نشان داد که استفاده از2/5 میلی­مولار گلوتاتیون در رقیق­کننده جنبایی کل و جنبایی پیش‌رونده را  افزایش داد (به‌ترتیب 1/5±63/38 و 1/1±43/1) (p <0/05). درصد زنده­مانی و سلامت غشای اسپرم در تیمار حاوی 2/5 میلی­مولار بیش‌تر از سایر تیمارها بود. به‌نظر می­رسد، غلظت 2/52 میلی­مولار گلوتاتیون می‌تواند غلظت بهینه رقیق­کننده انجمادی گاو بر پایه نانوذره لسیتین باشد.

کلیدواژه‌ها


عنوان مقاله [English]

Effect of supplementation of lecithin nanoparticles-based extender with reduced glutathione on freezability of bull sperm

نویسندگان [English]

  • Touba Nadri 1
  • saeed zeinoaldini 2
  • Armin towhodi 3
  • Gholamhossein Riazi 4
  • Mahdi zhandi 5
  • Mohsen Sharafi 6
1 دانشجوی دکتری فیزیولوژی تولید مثل دام
2 مدرس
3 مدرس
4 مدرس دانشگاه
5 مدرس
6 مدرس
چکیده [English]

This study was designed to add the reduced glutathione to a lecithin nanoparticle-based extender and evaluate the quality of bull sperm after freezing and thawing. In the present study, the effect of four different levels of glutathione (0, 1, 2.5 and 5 mM) in extender-based on lecithin nanoparticles was evaluated. The lecithin nanoparticles were prepared and the particle size was reduced by using a sonicator device. During three weeks, 48 ejaculates from six Holstein bulls were collected and frozen. Properties evaluated after freezing and thawing were kinetic parameters (CASA), membrane activity (HOST), membrane integrity (eosin-nigrosine) and morphology (Hancock solution). The results showed that using 2.5 mM glutathione in the extender significantly increased total and progressive motility (63.38±1.5 and 43.1±1.1 respectively, P <0.05). The results of eosin-nigrosine staining and Host test showed that the highest viability and cell membrane functionality were related to 2.5 mM treatment (64.8±1.5 and 58.1±1.1 respectively) (p < 0.05). In general, 2.5 mM glutathione could improve the quality of bull sperm compared with other concentrations after freezing and thawing process. It seems that the 2.5 mM glutathione is optimum concentration for bull extender based on the lecithin nanoparticles.
 

کلیدواژه‌ها [English]

  • Antioxidants
  • Bull
  • Freeze-Thaw
  • Semen
  • Nanomicelle
1. Agarwal A, Nallella KP, Allamaneni SS, Said
TM (2004) Role of antioxidants in treatment of
male infertility: an overview of the literature.
Reproductive Biomedicine Online 1; 8(6):
616-27.
2. Bhosrekar MR, Purohit JR, Mangurkar BR
(1990) Studies on the effect of additives to
semen diluent. Indian Journal of Animal
Reproduction 11(2): 85-8.
3. Chatterjee S, and Gagnon C (2001) Production of
reactive oxygen species by spermatozoa
undergoing cooling, freezing, and thawing.
Molecular Reproduction and Development:
Incorporating Gamete Research 59(4): 451-458 .
4. Cohen R, Mukai C, Travis AJ, (2016) Lipid
regulation of acrosome exocytosis. InSperm
acrosome biogenesis and function during
fertilization 107-127 .
5. de Oliveira RA, Wolf CA, de Oliveira Viu
MA, Gambarini ML (2013) Addition of
glutathione to an extender for frozen equine
semen. Journal of Equine Veterinary Science
1; 33(12): 1148-52 .
6. Donnelly ET, McClure N, and Lewis SE
(2000) Glutathione and hypotaurine in vitro:
effects on human sperm motility, DNA
integrity and production of reactive oxygen
species. Mutagenesis 15(1): 61-68.
7. Estrada E, del Álamo MM, Rodríguez-Gil JE,
and Yeste M (2017) The addition of reduced
glutathione to cryopreservation media induces
changes in the structure of motile
subpopulations of frozen-thawed boar
sperm. Cryobiology 1; 78: 56-64.
8. Gadea J, Molla M, Selles E, Marco MA,
Garcia-Vazquez FA, Gardon JC (2011)
Reduced glutathione content in human sperm
is decreased after cryopreservation: effect of
the addition of reduced glutathione to the
freezing and thawing extenders. Cryobiology
1; 62(1): 40-6.
9. Gadea J, Sellés E, Marco MA, Coy P, Matás C,
Romar R, Ruiz S, (2004) Decrease in glutathione
content in boar sperm after cryopreservation:
Effect of the addition of reduced glutathione to
the freezing and thawing extenders.
Theriogenology 1; 62(3-4): 690-701.
10. Gardón J, Rodriquez J, and Gadea J (2006)
Addition of reduced glutathione to thawing
medium improved the sperm motility and
reduced ros generation in frozen ovine and
caprine spermatozoa. Reproduction, Fertility
and Development 18(2): 155-155 .
11. Kothari S, Thompson A, Agarwal A, du Plessis
SS (2010) Free radicals: their beneficial and
detrimental effects on sperm function. 201.
12. Kvist U, Björndahl L, and Kjellberg S (1987)
Sperm nuclear zinc, chromatin stability, and
male fertility. Scanning Microscopy 1(3):
1241-1247 .
13. Li Z, Lin Q, Liu R, Xiao W, and Liu W (2010)
Protective effects of ascorbate and catalase on
human spermatozoa during cryopreservation.
Journal of Andrology 10; 31(5): 437-44.
14. Mari M, Morales A, Colell A, García-Ruiz C,
Fernández-Checa JC, (2009) Mitochondrial
glutathione, a key survival antioxidant.
Antioxidants & Redox Signaling 1; 11(11):
2685-700.
15. Meister A (1994) Glutathione, ascorbate, and
cellular protection. Cancer research 1; 54(7
Supplement):1969s-75s.
16. Nadri T, Towhidi A, Zeinoaldini S, Martínez-
Pastor F, Mousavi M, Noei R, Tar M, and
Sangcheshmeh AM (2019) Lecithin
nanoparticles enhance the cryosurvival of
caprine sperm. Theriogenology 15; 133: 38-44.
17. Najafi A, Zhandi M, Towhidi A, Sharafi M,
Sharif AA, Motlagh MK, and Martinez-Pastor
F (2013) Trehalose and glycerol have a dosedependent
synergistic effect on the postthawing
quality of ram semen cryopreserved in
a soybean lecithin-based extender.
Cryobiology 1; 66(3):275-82.
18. Triwulanningsih E, Situmorang P, Sugiarti T,
Sianturi RG, Kusumaningrum DA, (2010) Effect
of glutatione addition to the sperm diluent
medium on quality of bovine chilled semen.
Indonesian Journal of Agriculture 3(1): 60- 5 .
19. Niki E (2014) Antioxidants: basic principles,
emerging concepts, and problems. Biomed
Journal 37(3): 106- 111 .
20. Ogata K, Sasaki A, Kato Y, Takeda A,
Wakabayashi M, Sarentonglaga B, Yamaguchi
M, Hara A, Fukumori R, and Nagao Y (2015)
Glutathione supplementation to semen
extender improves the quality of frozenthawed
canine spermatozoa for transcervical
insemination. Journal of Reproduction and
Development 2014-130.
21. Salmani H, Nabi MM, Vaseghi-Dodaran H,
Rahman MB, Mohammadi-Sangcheshmeh A,
Shakeri M, Towhidi A, Shahneh AZ, and
Zhandi M (2013) Effect of glutathione in
soybean lecithin-based semen extender on goat
semen quality after freeze-thawing. Small
Ruminant Research 1; 112(1-3): 123-7.
22. Varghese AC, Das S, Bhattacharya AK,
Bhattacharya SM, Mandal M, Agarwal A
(2005) Effect of cryoprotective additivesreduced
glutathione, acetyl-L-carnitine on
sperm membrane lipid peroxidation, DNA
integrity and recovery of motile human sperm.
Fertility and Sterility. Sep 1; 84: S410-1.
23. Wang AW, Zhang H, Ikemoto I, Anderson DJ,
Loughlin KR (1997) Reactive oxygen species
generation by seminal cells during
cryopreservation. Urology 1; 49(6): 921-5.
24. Zhandi M, and Ghadimi V (2014) Effect of
glutathione-supplemented INRA82 extender
on miniature Caspian stallion sperm quality
during storage at 5 C. Journal of Equine
Veterinary Science 34(5): 606-610 .