SELİM ALÇAY, Mehmet Berk TOKER, ELİF GÖKÇE, Nail Tekin ÖNDER, BURCU ÜSTÜNER, ZEKARİYA NUR

Long Term Incubation Resilience of Post-Thaw Ram Semen Diluted with Lecithin-Based Extender Supplemented with Bovine Serum Albumin

The objective of the study was to determine the optimum concentration of BSA in lecithin-based extender for post-thawing quality and incubation resilience (0 h, 6 h and 10 h) of ram sperm. Ejaculates were collected from fie rams via electro ejaculation. Ejaculates were mixed to obtain pooled semen. Then, pooled semen was diluted with soybean lecithin-based extender without BSA (control) or supplemented with diffrent concentrations of BSA (2.5 mg/mL, 5 mg/mL, 7.5 mg/mL and 10 mg/mL), at a fial concentration of 150x106 spermatozoon/mL. Sperm motility, plasma membrane functional integrity (HOST), mitochondrial activity (rhodamine123), capacitation status (CTC), and DNA integrity (TUNEL) were evaluated. At the 10 h incubation, motility, plasma membrane functional integrity and mitochondrial function were better preserved in the BSA5 group compared to the control group. It was determined that high doses of BSA (5 mg/mL, 7.5 mg/mL and 10 mg/mL) affcted acrosome reaction. The highest acrosome reaction rates were obtained in BSA10 groups in 6 h and 10 h incubation (P

Sığır Serum Albumini İlave Edilmiş Lesitin Bazlı Sulandırıcı İle Sulandırılan Koç Spermasının Dondurma-Çözdürme Sonrası Uzun Süreli İnkübasyon Direnci

Bu çalışmanın amacı koç spermasının dondurma çözdürme sonrası ve inkübasyon direnci göz önüne alınarak (0, 6 ve 10. saatler) lesitin bazlı sulandırıcı için uygun Sığır Serum Albumini BSA konsantrasyonunun belirlenmesidir. Bu amaçla 5 koçtan sperma örnekleri alınmıştır. Alınan spermalar bir araya getirildikten sonra fial konsantrasyonu 150x106 spermatozoon/mL olacak şekilde farklı dozlarda BSA içeren (2.5 mg/mL, 5 mg/mL, 7.5 mg/mL and 10 mg/mL) ve içermeyen (kontrol) soya fasülyesi lesitini bazlı sulandırıcılarla sulandırılmıştır. Spermanın değerlendirilmesi amacıyla motilite, plazma membran fonksiyonel bütünlüğü (HOST), mitokondriyal aktivite (Rhodamine123), kapasitasyon statüsü (CTC) ve DNA bütünlüğü değerlerine bakılmıştır. İnkübasyonun sonunda (10. saat) BSA5 grubunun motiliteyi, plazma membran fonksiyonel bütünlüğü ve mitokondriyal fonksiyonu diğer gruplara göre daha fazla koruduğu görülmüştür. BSA’nın yükselen dozları (5 mg/ mL, 7.5 mg/mL and 10 mg/mL) akrozom reaksiyonunu etkilemektedir. İnkubasyonun 6 ve 10. saatlerinde en yüksek akrozom reaksiyonu oranları BSA10 grubunda gözlemlenmiştir (P

PDF

___

Salamon S, Maxwell WM: Storage of ram semen. Anim Reprod Sci, 62, 77-111, 2000. DOI: 10.1016/S0378-4320(00)00155-X

Holt WV, Penfold LM: Fundamental and practical aspects of semen cryopreservation. In, Chenoweth P, Lorton S (Eds): Animal Andrology: Theories and Applications, 76-99, CAB International CABI, Wallingford Oxfordshire, 2014. DOI: 10.1079/9781780643168.0076

Fiser PS, Fairfull RW: The effects of rapid cooling (cold shock) of ram semen, photoperiod, and egg yolk in diluents on the survival of spermatozoa before and after freezing. Cryobiology, 23, 518-524, 1986. DOI: 10.1016/0011-2240(86)90061-1

O’Connell M, McClure N, Lewis SEM: The effcts of cryopreservation on sperm morphology, motility and mitochondrial function. Hum Reprod, 17 (3): 704-709, 2002. DOI: 10.1093/humrep/17.3.704

Ustuner B, Alcay S, Toker MB, Nur Z, Gokce E, Sonat FA, Gul Z, Duman M, Ceniz C, Uslu A, Sagirkaya H, Soylu MK: Effct of rainbow trout (Oncorhynchus mykiss) seminal plasma on the post-thaw quality of ram semen cryopreserved in a soybean lecithin-based or egg yolkbased extender. Anim Reprod Sci, 164, 97-104, 2016. DOI: 10.1016/J. ANIREPROSCI.2015.11.017

Yildiz S, Öztürkler Y, Ari UC, Lehimcioglu NC, Atakisi E, Kulaksiz R: The effcts of l-ergothioneine, n-acetylcystein and cystein on freezing of ram semen. Kafkas Univ Vet Fak Derg, 21 (1): 81-86, 2015. DOI: 10.9775/ kvfd.2014.11792

Ari UC, Kulaksiz R, Ozturkler Y, Lehimcioglu NC, Yildiz S: Effct of n-acetylcysteine (NAC) on post-thaw semen quality of Tushin rams. Kafkas Univ Vet Fak Derg, 22 (6): 883-887, 2016. DOI: 10.9775/kvfd.2016.15558

Ari UC, Kulaksiz R, Ozturkler Y, Yildiz S, Lehimcioglu NC: Effect of L -(+)-ergothioneine (EGT) on freezability of ram semen. Int J Anim Vet Adv, 4 (6): 378-383, 2012.

Ozturkler Y, Ari UC: Addıtives used ın extenders to ımprove the freezabılıty of ram semen ın recent years: A mini review. Agric Biol, 52 (2): 242-250, 2017.

Ari UC, Kulaksiz R, Ozturkler Y: Freezability of tushin ram semen extended with goat or cow milk based extenders. Reprod Domest Anim, 46 (6): 975-979, 2011. DOI: 10.1111/j.1439-0531.2011.01769.x

Ustuner B, Alcay S, Nur Z, Soylu MK: Effct of egg yolk and soybean lecithin on tris-based extender in post-thaw ram semen quality and in vitro fertility. Kafkas Univ Vet Fak Derg, 20, 393-398, 2014. DOI: 10.9775/ kvfd.2013.10248

Toker MB, Alcay S, Gokce E, Ustuner B: Cryopreservation of ram semen with antioxidant supplemented soybean lecithin-based extenders and impacts on incubation resilience. Cryobiology, 72, 205-209, 2016. DOI: 10.1016/J.CRYOBIOL.2016.05.001

Masoudi R, Sharafi M, Zare Shahneh A, Towhidi A, Kohram H, Zhandi M, Esmaeili V, Shahverdi A: Effect of dietary fish oil supplementation on ram semen freeze ability and fertility using soybean lecithin-and egg yolk-based extenders. Theriogenology, 86, 1583-1588, 2016. DOI: 10.1016/j.theriogenology.2016.05.018

Mehdipour M, Daghigh Kia H, Nazari M, Najafi A: Effct of lecithin nanoliposome or soybean lecithin supplemented by pomegranate extract on post-thaw flow cytometric, microscopic and oxidative parameters in ram semen. Cryobiology, 78, 34-40, 2017. DOI: 10.1016/J. CRYOBIOL.2017.07.005

Motlagh MK, Sharafi M, Zhandi M, Mohammadi-Sangcheshmeh A, Shakeri M, Soleimani M, Zeinoaldini S: Antioxidant effct of rosemary (Rosmarinus officinalis L.) extract in soybean lecithin-based semen extender following freeze–thawing process of ram sperm. Cryobiology, 69, 217-222, 2014. DOI: 10.1016/J.CRYOBIOL.2014.07.007

Najafi A, Daghigh-Kia H, Dodaran HV, Mehdipour M, AlvarezRodriguez M: Ethylene glycol, but not DMSO, could replace glycerol inclusion in soybean lecithin-based extenders in ram sperm cryopreservation. Anim Reprod Sci, 177, 35-41, 2017. DOI: 10.1016/J.ANIREPROSCI.2016.12.004

Uysal O, Bucak MN: Effcts of oxidized glutathione, bovine serum albumin, cysteine and lycopene on the quality of frozen-thawed ram semen. Acta Vet Brno, 76, 383-390, 2007. DOI: 10.2754/avb200776030383

Uysal O, Bucak MN, Yavas I, Varisli O: Effct of various antioxidants on the quality of frozen-thawed bull semen. J Anim Vet Adv, 6, 1362-1366, 2007.

Davis BK, Byrne R, Hungund B: Studies on the mechanism of capacitation. II. Evidence for lipid transfer between plasma membrane of rat sperm and serum albumin during capacitation in vitro. Biochim Biophys Acta, 558, 257-266, 1979. DOI: 10.1016/0005-2736(79)90260-8

Xia J, Ren D: The BSA-induced Ca 2+ inflx during sperm capacitation is CATSPER channel-dependent. Reprod Biol Endocrinol, 7:119, 2009. DOI: 10.1186/1477-7827-7-119

Hossain MS, Hyeong LJ, Mıah AG, Tsujıı H: Effct of fatty acids bound to bovine serum albumin-V on acrosome reaction and utilization of glucose in boar spermatozoa. Reprod Med Biol, 6, 109-115, 2007. DOI: 10.1111/j.1447-0578.2007.00173.x

Gokce E, Alcay S, Gul Z: Positive effct of BSA supplemented soybean lecithin based extender on liquid storage of ram semen at 5°C. Ankara Univ Vet Fak Derg, 64, 313-320, 2017.

Amidi F, Farshad A, Khor AK: Effcts of cholesterol-loaded cyclodextrin during freezing step of cryopreservation with TCGY extender containing bovine serum albumin on quality of goat spermatozoa. Cryobiology, 61, 94-99, 2010. DOI: 10.1016/j.cryobiol.2010.05.006

Anghel A, Zamfirescu S, Dragomir C, Nadolu D, Elena S, Florica B: The effcts of antioxidants on the cytological parameters of cryopreserved buck semen. Rom Biotechnol Lett, 15 (3): 26-32, 2010.

Ball BA, Medina V, Gravance CG, Baumbe J: Effct of antioxidants on preservation of motility,viability and acrosomal integrity of equine spermatozoa during storage at 5°C. Theriogenology, 56, 577-589, 2001. DOI: 10.1016/S0093-691X(01)00590-8

Akhter S, Rakha BA, Iqbal R, Ansari MS: Effct of bovine serum albumin on motility, plasmalemma, viability and chromatin ıntegrity of bufflo bull spermatozoa. Pakistan J Zool, 46, 115-120, 2014.

Sariozkan S, Turk G, Canturk F, Yay A, Eken A, Akcay A: The effct of bovine serum albumin and fetal calf serum on sperm quality, DNA fragmentation and lipid peroxidation of the liquid stored rabbit semen. Cryobiology, 67, 1-6, 2013. DOI: 10.1016/j.cryobiol.2013.04.002

Bakst MR, Cecil HC: Effct of bovine serum albumin on motility and fecundity of turkey spermatozoa before and after storage. J Reprod Fertil, 94, 287-293, 1992. DOI: 10.1530/jrf.0.0940287

Leboeuf B, Le Vern Y, Furstoss V, Kerboeuf D, Guıllouet P, Magıstrını M: Response of goat sperm to hypoosmotic steps modelled probit analysis. Anim Reprod Sci, 91, 265-274, 2006. DOI: 10.1016/j. anireprosci.2005.04.014

Alcay S, Gokce E, Toker MB, Onder NT, Ustuner B, Uzabaci E, Gul Z, Cavus S: Freeze-dried egg yolk based extenders containing various antioxidants improve post-thawing quality and incubation resilience of goat spermatozoa. Cryobiology, 72, 269-273, 2016. DOI: 10.1016/j. cryobiol.2016.03.007

Fraser L, Lecewicz M, Strzezek J: Fluorometric assessments of viability and mitochondrial status of boar spermatozoa following liquid storage. Pol J Vet Sci, 5, 85-92, 2002.

Pérez LJ, Valcárcel A, de las Heras MA, Moses DF, Baldassarre H: In vitro capacitation and induction of acrosomal exocytosis in ram spermatozoa as assessed by the chlortetracycline assay. Theriogenology, 45, 1037-1046, 1996. DOI: 10.1016/0093-691X(96)00031-3

Nur Z, Zik B, Ustuner B, Sagirkaya H, Ozguden CG: Effcts of diffrent cryoprotective agents on ram sperm morphology and DNA integrity. Theriogenology, 73, 1267-1275, 2010. DOI: 10.1016/J.THERIOGENOLOGY. 2009.12.007

Matsuoka T, Imai H, Kohno H, Fukui Y: Effcts of bovine serum albumin and trehalose in semen diluents for improvement of frozenthawed ram spermatozoa. J Reprod Develop, 52, 675-683, 2006. DOI: 10.1262/jrd.18033

Maxwell WM, Salamon S: Liquid storage of ram semen: A review. Reprod Fertil Dev, 5, 613-638, 1993. DOI: 10.1071/RD9930613

El-Kon I: Testing usability of bovine serum albumin (BSA) for preservation of egyptian bufflo semen. Am Eurasian J Agric Environ Sci, 11, 495-502, 2011.

Fang L, Bai C, Chen Y, Dai J, Xiang Y, Ji X, Huang C, Dong Q: Inhibition of ROS production through mitochondria-targeted antioxidant and mitochondrial uncoupling increases post-thaw sperm viability in yellow catfish. Cryobiology, 69, 386-393, 2014. DOI: 10.1016/j.cryobiol.2014.09.005

Bedford JM: Significance of the need for sperm capacitation before fertilization in eutherian mammals. Biol Reprod, 28, 108-120, 1983.

Baek S, Lee ST, Hwang JY, Park KH, Yun JI: Identification of capacitation inducers customized to sperm retrieved from inbred mouse epididymis. Biochem Biophys Res Commun, 488, 273-277, 2017. DOI: 10.1016/J.BBRC.2017.04.152

Visconti PE: Understanding the molecular basis of sperm capacitation through kinase design. Proc Natl Acad Sci U S A, 106, 667-668, 2009. DOI: 10.1073/pnas.0811895106

López-Fernández C, Fernández JL, Gosálbez A, Arroyo F, Vázquez JM, Holt WV, Gosálvez J: Dynamics of sperm DNA fragmentation in domestic animals III. Ram. Theriogenology, 70, 898-908, 2008. DOI: 10.1016/j.theriogenology.2008.04.055

Alcay S, Ustuner B, Nur Z: Effects of low molecular weight cryoprotectants on the post-thaw ram sperm quality and fertilizing ability. Small Ruminant Res, 136, 59-64, 2016. DOI: 10.1016/J.SMALLRUMRES. 2016.01.009

Alcay S, Toker MB Gokce E, Gul Z, Onder NT, Ustuner B, Nur Z, Sagirkaya H, Soylu MK: Improvement of incubation resilience with various antioxidants in cryopreserved ram semen. Erciyes Üniv Vet Fak Derg, 14 (3): 183-190, 2017.