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اثر سطوح مختلف گلوتاتیون درون‌پوشانی‌شده بر‎ ‎کیفیت انجماد اسپرم گاو

نوع مقاله : مقاله پژوهشی

نویسندگان

1 دانشجوی دکتری فیزیولوژی دام، پردیس کشاورزی و منابع طبیعی دانشگاه تهران، کرج

2 دانشیار، پردیس کشاورزی و منابع طبیعی دانشگاه تهران، کرج

3 استاد، پردیس کشاورزی و منابع طبیعی دانشگاه تهران، کرج

4 استاد، گروه بیوشیمی، دانشگاه تهران

5 استادیار، گروه علوم دامی، دانشکده کشاورزی، دانشگاه تربیت مدرس

چکیده

هدف از این مطالعه، ارزیابی اثر سطوح مختلف گلوتاتیون احیا در رقیق­کننده بر کیفیت اسپرم گاو پس از فرایند انجماد و ذوب بود. در پژوهش حاضر، رقیق­کننده حاوی نانولیپوزوم­های غنی‌شده با چهار سطح صفر، 1، 5/2 و 5 میلی‌مولار آنتی اکسیدان گلوتاتیون بود. لیپوزوم­های حاوی گلوتاتیون با استفاده از روش فیلم نازک تهیه شد. اندازه ذرات با استفاده از دستگاه سونیکاتور به ابعاد نانو کاهش یافت. چهل انزال طی شش هفته از شش راس گاو نر هلشتاین جمع­آوری و مورد انجماد قرار گرفت. صفات مورد ارزیابی پس از انجماد و ذوب شامل، فراسنجه­های جنبایی، فعالیت غشا  و یکپارچگی غشا و ریخت­شناختی اسپرم بود. نتایج این مطالعه نشان داد که استفاده از 1 میلی‌مولار گلوتاتیون جنبایی کل و جنبایی پیشرونده را به‌طور معنی‌داری افزایش می‌دهد (7/1±5/47 و 2±7/32  درصد)، اگرچه، تفاوت معنی‌داری با غلظت 5/2 میلی‌مولار نداشت (7/1±0/45 و 2±2/26 درصد). همچنین، غلظت 5/2 میلی‌مولار بالاترین میزان یکپارچگی و سلامت غشا را نشان داد، اما تفاوت معنی‌داری با غلظت 1 میلی‌مولار نشان نداد. با این‌حال، 5 میلی­مولار گلوتاتیون درون پوشانی شده سبب کاهش فراسنجه­های کیفی اسپرم شد. بنابراین، استفاده از سطح 1 میلی‌مولار گلوتاتیون به شکل درون­پوشانی شده در رقیق­کننده می­تواند حفاظت بهتری از اسپرم گاو نسبت به سایر سطوح به عمل آورد.

کلیدواژه‌ها


عنوان مقاله [English]

Effect of different levels of encapsulated glutathione on cryopreservation of bull ‎sperm

نویسندگان [English]

  • Touba Nadri 1
  • Saeed Zeinoaldini 2
  • Armin Towhidi 3
  • Gholamhossein Riazi 4
  • Mahdi Zhandi 2
  • Mohsen Sharafi 5
1 Ph.D. Candidate of Animal Physiology, Department of Animal Science, College of Agriculture ‎and Natural Resources, University of Tehran, Karaj, Iran
2 Associate Professor, Department of Animal Science, College of Agriculture and Natural Resources, University of ‎Tehran, Karaj, Iran
3 Professor, Department of Animal Science, College of Agriculture and Natural Resources, University of Tehran, Karaj, Iran
4 Professor, IBB Center, University of Tehran, Tehran, Iran
5 Assistant Professor of Animal Physiology, Department of Animal Science, Tarbiat Modares University, Tehran, Iran
چکیده [English]

This study was designed to evaluate the effects of encapsulated reduced glutathione added to extender on the quality of bull sperm after freezing and thawing process. Extender contains lecithin nanoliposomes supplemented with different concentrations of glutathione (0, 1, 2.5 and 5 Mm concentration). The supplemented nanoliposomes with glutathione were prepared by thin-film preparation method.  The particle size decreased to nanometer by sonication. 40 ejaculations were collected from 6 Holstein bulls and frozen for 6 weeks. Evaluated characteristics after freezing and thawing process were kinetic parameters (CASA), membrane activity (HOST), membrane integrity (eosin-nigrosin) and morphology of sperm ( Hancock's solution). Results showed that level 1 mM significantly increased the total and progressive motility (47.5±1.7, 32.7±2 %).  Although, there was no significant difference with level 2.5 mM glutathione (45.0±1.7, 26.2±2 %). Also, the concentration of 2.5 mM showed the highest value of cell membrane integrity and cell membrane functionality but did not differ with the concentration of 1mM. However, supplementation of nanoliposomes with 5 mM glutathione reduced the quality of post-thaw sperm. Thus, level 1 M encapsulated glutathione can provide better protection compare to the other level of that for bull sperm.

کلیدواژه‌ها [English]

  • Bull semen
  • Encapsulation
  • Freezing
  • Glutathione
  • nanoparticels
Aitken, R.J., Gordon, E., Harkiss, D., Twigg, J.P., Milne, P., Jennings, Z. & Irvine, D.S. )1998(. Relative impact of oxidative stress on the functional competence and genomic integrity of human spermatozoa. Biology of Reproduction 59, 1037-1046.
Ansari, M. S., Rakha, B. A., Ullah, N., Andrabi, S. M. H., Iqbal, S., Khalid, M. & Akhter, S. (2010). Effect of exogenous glutathione in extender on the freezability of Nili-Ravi buffalo (Bubalus bubalis) bull spermatozoa. Animal Science, 28, 235-244.
Belala, R., Delay, J., Amirat, L., Ropers, M.-H., Le Guillou, J., Anton, M. & Kaidi, R. (2016). The benefits of liposomes for chilling canine sperm for 4 days at 4 C. Animal Reproduction Science, 168, 100-109.
Bilodeau, J. F., Chatterjee, S., Sirard, M. A. & Gagnon, C. (2000). Levels of antioxidant defenses are decreased in bovine spermatozoa after a cycle of freezing and thawing. Molecular Reproduction and Development: Incorporating Gamete Research, 55, 282-288.
Bucak, M.N. & Tekin, N. (2007). Protective effect of taurine, glutathione and trehalose on the liquid storage of ram semen. Small Ruminant Research 73, 103-108.
Bucak, M.N., Tuncer, P.B., Sarıözkan, S., Başpınar, N., Taşpınar, M., Çoyan, K., Bilgili, A., Akalın, P.P., Büyükleblebici, S. & Aydos, S. (2010). Effects of antioxidants on post-thawed bovine sperm and oxidative stress parameters: antioxidants protect DNA integrity against cryodamage. Cryobiology, 61, 248-253.
Câmara, D., Silva, S., Almeida, F., Nunes, J. & Guerra, M. (2011). Effects of antioxidants and duration of pre-freezing equilibration on frozen-thawed ram semen. Theriogenology, 76, 342-350.
Chapman, D., Jones, M. N. & Jones, M. N. (1995). Micelles, monolayers, and biomembranes. Wiley-Liss.
Circu, M. L. & Aw, T. Y. (2012). Glutathione and modulation of cell apoptosis. Biochimica et Biophysica Acta (BBA)-Molecular Cell Research, 1823, 1767-1777.
Darin-Bennett, A., Poulos, A. & White, I. (1973). The effect of cold shock and freeze-thawing on release of phospholipids by ram, bull, and boar spermatozoa. Australian Journal of Biological Sciences, 26, 1409-1420.
de Oliveira, R. A., Wolf, C. A., de Oliveira Viu, M. A. & Gambarini, M. L. (2013). Addition of glutathione to an extender for frozen equine semen. Journal of Equine Veterinary Science, 33, 1148-1152.
Estrada, E., del Álamo, M. M. R., Rodríguez-Gil, J. E. & 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, 78, 56-64.
Gadea, J., Molla, M., Selles, E., Marco, M., Garcia-Vazquez, F. & Gardon, J. (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, 62, 40-46.
Gadea, J., Sellés, E., Marco, M.A., 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, 62, 690-701.
Gardón, J., Rodriquez, J. & 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, 155-155.
Garrett, F. E., Goel, S., Yasul, J. & Koch, R. A. (1999). Liposomes fuse with sperm cells and induce activation by delivery of impermeant agents. Biochimica et Biophysica Acta (BBA)-Biomembranes, 1417, 77-88.
Graham, J., Foote, R. (1987). Effect of several lipids, fatty acyl chain length, and degree of unsaturation on the motility of bull spermatozoa after cold shock and freezing. Cryobiology, 24, 42-52.
Irvine, D.S. (1996). Glutathione as a treatment for male infertility. Reviews of Reproduction, 1, 6-12.
Lev, S. (2010). Non-vesicular lipid transport by lipid-transfer proteins and beyond. Nature reviews Molecular Cell Biology, 11, 739.
Meister, A. (1994). Glutathione, ascorbate, and cellular protection. Cancer Research, 54(7 Supplement), 1969s-1975s.
Munsi, M., Bhuiyan, M., Majumder, S. & Alam, M. (2007). Effects of exogenous glutathione on the quality of chilled bull semen. Reproduction in Domestic Animals, 42, 358-362.
Nadri, T., Towhidi, A., Zeinoaldini, S., Martínez-Pastor, F., Mousavi, M., Noei, R. & Sangcheshmeh, A. M. (2019). Lecithin nanoparticles enhance the cryosurvival of caprine sperm. Theriogenology, 133, 38-44.
Najafi, A., Zhandi, M., Towhidi, A., Sharafi, M., Sharif, A. A., Motlagh, M. K. & Martinez-Pastor, F. (2013). Trehalose and glycerol have a dose-dependent synergistic effect on the post-thawing quality of ram semen cryopreserved in a soybean lecithin-based extender. Cryobiology, 66(3), 275-282.
Ogata, K., Sasaki, A., Kato, Y., Takeda, A., Wakabayashi, M., Sarentonglaga, B. & Nagao, Y. (2015). Glutathione supplementation to semen extender improves the quality of frozen-thawed canine spermatozoa for transcervical insemination. Journal of Reproduction and Development, 2014-2130.
Quinn, P., Chow, P. & White, I. (1980). Evidence that phospholipid protects ram spermatozoa from cold shock at a plasma membrane site. Reproduction, 60, 403-407.
Rawash, Z. M., Ibrahim, E. A. & El-Raey, M. (2018). Effects of reduced glutathione on Boer goat semen freezability. Asian Pacific Journal of Reproduction, 7, 33.
Said, T. M., Gaglani, A. & Agarwal, A. (2010). Implication of apoptosis in sperm cryoinjury. Reproductive Biomedicine Online, 21, 456-462.
Sakkas, D. & Alvarez, J. G. (2010). Sperm DNA fragmentation: mechanisms of origin,   impact on reproductive outcome, and analysis. Fertility and Sterility 93, 1027-1036.
Salamon, S. & Maxwell, W. (2000). Storage of ram semen. Animal Reproduction Science, 62, 77-111.
Schäfer, S. & Holzmann, A. (2000). The use of transmigration and Spermac™ stain to evaluate epididymal cat spermatozoa. Animal Reproduction Science, 59, 201-211.
Salmani, H., Nabi, M. M., Vaseghi-Dodaran, H., Rahman, M. B., Mohammadi-Sangcheshmeh, A., Shakeri, M. & Zhandi, M. (2013). Effect of glutathione in soybean lecithin-based semen extender on goat semen quality after freeze-thawing. Small Ruminant Research, 112, 123-127.
Sinha, M., Sinha, A., Singh, B. & Prasad, R. (1996). The effect of glutathione on the motility, enzyme leakage and fertility of frozen goat semen. Animal Reproduction Science, 41, 237-243.
Sinha, R., Sinha, I., Calcagnotto, A., Trushin, N., Haley, J. S., Schell, T. D. & Richie Jr, J. P. (2018). Oral supplementation with liposomal glutathione elevates body stores of glutathione and markers of immune function. European Journal of Clinical Nutrition, 72, 105-111.
Sharafi, M., Zhandi, M., Shahverdi, A., Shakeri, M., 2015. Beneficial effects of nitric oxide induced mild oxidative stress on post-thawed bull semen quality. International Journal of Fertility & Sterility, 9, 230.
Stradaioli, G., Noro, T., Sylla, L. & Monaci, M. (2007). Decrease in glutathione (GSH) content in bovine sperm after cryopreservation: comparison between two extenders. Theriogenology, 67, 1249-1255.
Ugur, M. R., Saber Abdelrahman, A., Evans, H. C., Gilmore, A. A., Hitit, M., Arifiantini, R. L., Purwantara, B., Kaya, A. & Memili, E. (2019). Advances in Cryopreservation of Bull Sperm. Frontiers in Veterinary Science, 6, 268.
Thomson, L. K., Fleming, S. D., Barone, K., Zieschang, J.-A. & Clark, A. M. (2010). The effect of repeated freezing and thawing on human sperm DNA fragmentation. Fertility and Sterility, 93(4), 1147-1156.
Triwulanningsih, E., Situmorang, P., Sugiarti, T., Sianturi, R. & Kusumaningrum, D. (2010). Effect of glutatione addition to the sperm diluent medium on quality of bovine chilled semen. Indonesian Journal of Agriculture, 3, 60-65.