title: Raffinose and hypotaurine improve the post-thawed Merino ram sperm parameters.
abstract: The aim of this study was to determine the effects of raffinose and hypotaurine on sperm parameters after the freeze-thawing of Merino ram sperm. Totally 40 ejaculates of five Merino ram were used in the study. Semen samples, which were diluted with a Tris-based extender containing 10mM raffinose, 5mM hypotaurine, 5mM raffinose +2.5mM hypotaurine (H+R) and no antioxidant (control), were cooled to 5 °C and frozen in 0.25 ml French straws and stored in liquid nitrogen. Frozen straws were then thawed individually at 37 °C for 25s in a water bath for evaluation. The addition of raffinose led to higher percentages of subjective and CASA motilities (47.5 ± 12.2%, 46.3 ± 13.6%) compared to controls (38.8 ± 13.8%, 30.5 ± 11.7%, P<0.05). For the CASA progressive motility, 5mM raffinose (20.12 ± 8.82%) had increasing effect in comparison to control (10 ± 7.94%, P<0.05) following the freeze-thawing process. Raffinose and hypotaurine led to higher viability (40.8 ± 4.68%, 40.8 ± 4.7%), high sperm mitochondrial activity (29.5 ± 5.4%, 27.3 ± 4.9%) and acrosome integrity (50.8 ± 8.1, 50.7 ± 4.4) percentages, compared to control groups (31.5 ± 3.5%, 9.5 ± 8.2%, 42.8 ± 7.3%, P<0.05). H+R group only led to high sperm mitochondrial activity when compared to control group. In the comet test, raffinose and hypotaurine resulted in lower sperm with damaged DNA (6.2% and 3.9%) than that of control (9.1%), reducing the DNA damage. For TUNEL assay, The TUNEL-positive cell was distinguished by distinct nuclear staining. Raffinose and H+R groups resulted in lower sperm with TUNEL-positive cell (1.5 ± 1.2% and 2.1 ± 0.9%) than that of control (4.9 ± 2.5%) (P<0.05). In conclusion, findings of this study showed that raffinose and hypotaurine supplementation in semen extenders provided a better protection of sperm parameters against cryopreservation injury, in comparison to the control groups.
authors: Veterinary Faculty, Department of Reproduction and Artificial Insemination, Selcuk University, Konya, Turkey. mustafanbucak@yahoo.com
authors: Mustafa Numan
authors: MN
authors: Bucak
authors: Nazan
authors: N
authors: Keskin
authors: Mehmet
authors: M
authors: Taşpınar
authors: Kenan
authors: K
authors: Çoyan
authors: Nuri
authors: N
authors: Başpınar
authors: Mihai C
authors: MC
authors: Cenariu
authors: Ali
authors: A
authors: Bilgili
authors: Caner
authors: C
authors: Öztürk
authors: Ahmet Nuri
authors: AN
authors: Kurşunlu
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000187:drug effects; D015925:Cryopreservation / Q000379:methods* / Q000662:veterinary; D004249:DNA Damage / Q000187:drug effects; D008297:Male; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D011887:Raffinose / Q000494:pharmacology*; D012662:Semen Preservation / Q000379:methods* / Q000662:veterinary; D012756:Sheep; D013081:Sperm Motility / Q000187:drug effects; D013094:Spermatozoa; D013654:Taurine / Q000031:analogs & derivatives* / Q000494:pharmacology
chemical_list: D013654:Taurine; C003949:hypotaurine; D011887:Raffinose