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Öğe Comparison of cryoprotective effects of iodixanol, trehalose and cysteamine on ram semen(Elsevier, 2013) Cirit, Umut; Bagis, Haydar; Demir, Kamber; Agca, Cansu; Pabuccuoglu, Serhat; Varisli, Omer; Clifford-Rathert, CharlotteThis study was conducted to improve cryosurvival of electroejaculated (EE) ram semen in the presence of iodixanol (OptiPrep (TM)), trehalose or cysteamine. A tris-based extender was used to prepare 12 extenders containing OptiPrep (TM) (Op), trehalose (Tr) or cysteamine (Cy) alone, or different combinations of these compounds. Extenders were designated as follows: Tris (control), Op1.25 (1.25% Op, v/v), Op2.5 (2.5% Op, v/v), Op5 (5% Op, v/v), Tr50 (50 mM Tr), Tr100 (100 mM Tr), Cy (5 mM Cy), OpTr (2.5% Op and 100 mM Tr), OpCy (2.5% Op and 5 mM Cy), TrCy (100 mM Tr and 5 mM Cy), OpTrCy1 (2.5% Op, 100 mM Tr and 5 mM Cy) and OpTrCy2 (1.25% Op, 50 mM Tr and 2.5 mM Cy). A two-step dilution was used and glycerol was added at 5 degrees C in the second step. Diluted samples were equilibrated for 1 h, loaded in 0.25 mL straws and frozen in a programmable freezing machine. Supplementation of 5% OptiPrep (TM) significantly protected post-thaw progressive motility, membrane integrity, acrosomal integrity and morphological damages. Trehalose supplementation protected membrane integrity of ram sperm; however, it did not help post-thaw motility and morphology. Supplementation of 5 mM cysteamine had detrimental effect on cryosurvival of EE ram semen. These results demonstrate that the supplementation of iodixanol increases the cryosurvival of EE ram semen in a dose-dependent manner. (c) 2013 Elsevier B.V. All rights reserved.Öğe Comparison of Two Different Media for In vitro Production of Dog Embryos(Kafkas Univ, Veteriner Fakultesi Dergisi, 2010) Evecen, Mithat; Cirit, Umut; Demir, Kamber; Karaman, Elif; Bakirer, Gul; Hamzaoglu, Asiye Izem; Birler, SemaEmbryo production via in vitro fertilization and nuclear transfer has been accomplished in the dog, and the transfer of the cloned embryos has recently resulted in the birth of puppies. However, the efficiency of these technologies is still very limited. Until now, only two morulas and single blactocyst have been achieved in vitro. Therefore, the aim of the present study was to examine the effects of two different media (mSOF and TCM 199) on in vitro maturation (IVM), in vitro fertilization (IVF) and in vitro culture (IVC) of immature dog oocytes. The study was performed in two steps. At the first step, the effects of two different media, on IVM of dog oocytes were investigated. At the end of the IVM period, the nuclear maturation rates were evaluated by aceto-orcein staining method. At the second step, after the IVM period the oocytes inseminated with fresh spermatozoa for 24 h and left for IVC for 7 d. At the end of the IVC period, embryonic development was assessed by microscopic observation at 24 h intervals and then fixed and stained by the same method. Consequently, maturation rates of oocytes in mSOF medium were significantly higher than those of the TCM 199 (P<0.001). After 7 d of the IVC period, only one oocyte was cleaved in the TCM 199. while eight oocytes cleaved and one of them developed to morula stage in mSOF medium group (P=0.037).Öğe Development of starch based mucoadhesive vaginal drug delivery systems for application in veterinary medicine(Elsevier Sci Ltd, 2016) Gok, Mehmet Koray; Ozgumus, Saadet; Demir, Kamber; Cirit, Umut; Pabuccuoglu, Serhat; Cevher, Erdal; Ozsoy, YildizThe aim of this study was to prepare and evaluate the mucoadhesive, biocompatible and biodegradable progesterone containing vaginal tablets based on modified starch copolymers for the estrus synchronization of ewes. Starch-graft-poly( acrylic acid) copolymers (S-g-PAA) were synthesized and characterized. The vaginal tablets were fabricated with S-g-PAA and their equilibrium swelling degree (Qe) and matrix erosion (ME%) were determined in lactate buffer solution. In vitro, mucoadhesive properties of the tablets were investigated by using ewe vaginal mucosa and in vivo residence time were also investigated. In vitro and in vivo progesterone release profiles from the tablets were compared with two commercial products. Tablet formulation containing wheat starch based grafted copolymer (WS-g-PAA)(gc) indicated promising results and might be convenient as an alternative product to the commercial products in veterinary medicine. (C) 2015 Elsevier Ltd. All rights reserved.Öğe Effect of different transport temperatures on in vitro maturation of oocytes collected from frozen-thawed sheep ovaries(Tubitak Scientific & Technological Research Council Turkey, 2013) Ozdas, Ozen Banu; Baran, Alper; Tas, Muzaffer; Cirit, Umut; Demir, Kamber; Bacinoglu, Suleyman; Pabuccuoglu, SerhatThe aim of this study was to determine the effects of 2 different transport temperatures on the in vitro maturation of oocytes collected from frozen-thawed sheep ovaries. Sheep ovaries were transferred into saline at temperatures of 4 degrees C and 32 degrees C. After the 2 experimental groups (A: fresh cortex, B: frozen-thawed cortex) were formed, each group was divided into 2 subgroups (group A1: 4 degrees C, group A2: 32 degrees C [control]; group B1: 4 degrees C, group B2: 32 degrees C). The cortexes were dissected into slices 1-3 mm thick and pieces of 0.5 cm(2). For groups B1 and B2, 1-2 cortex pieces were placed in cryogenic vials containing 1 mL of freezing medium modified with Earle's salts (TCM-199) and supplemented with 10% fetal calf serum (FCS) (FCS + 2.5 M ethylene glycol + 0.1 M sucrose). The vials were then cooled to 7 degrees C at 2 degrees C/min and held at 7 degrees C for 10 min for manual seeding. The temperature was then lowered by 0.3 degrees C/min to -35 degrees C and thereafter by -10 degrees C/min to -75 degrees C. Vials were plunged into -196 degrees C liquid nitrogen and stored. Cortexes were thawed at 37 degrees C. Collected oocytes were matured in their own groups in 700 mu L of TCM-199 (supplemented with luteinizing hormone, follicle-stimulating hormone, pyruvate, and FCS) for 23 h in a gas mixture of 5% CO2, 5% O-2, and 90% N-2 at 38.8 degrees C. After maturation, oocytes were fixed in acetic acid and ethyl alcohol (1: 3) for 48 h. Oocytes were stained with aceto-orcein and then examined. At the end of the study, maturation rates for reaching metaphase I (MI) were similar in all groups (group A1: 30.76%, group A2: 38.09%, group B1: 30.65%, and group B2: 33.33%). The rates at which metaphase II (MII) was reached were 18.58%, 34.69%, 7.25%, and 6.48%, respectively. The best development was seen in group A2 (P < 0.001). Sheep oocytes obtained from fresh and frozen-thawed cortexes reached the MII stage if transported at 4 degrees C.Öğe Effect of Oocyte Diameter on in vitro Embryo Production in Dogs(Kafkas Univ, Veteriner Fakultesi Dergisi, 2010) Evecen, Mithat; Cirit, Umut; Demir, Kamber; Karaman, Elif; Bakirer, Gul; Hamzaoglu, Asiye Izem; Birler, SemaIn vitro embryo production has not been confidently applied to the dog successfully. Up to date, only one blactocyst have been achieved by in vitro culture. Therefore, the aim of the present study was to examine the effects of different oocyte diameters (<= 100 mu m and >100 mu m) on in vitro maturation (IVM), in vitro fertilization (IVF) and in vitro culture (IVC) of immature dog oocytes. The study was performed in two steps. At the first step (experiment I), effects of two different oocyte diameters on IVM of dog oocytes were investigated. The nuclear maturation rates were evaluated by aceto-orcein staining method at the end of the IVM. At the second step (experiment II), in vitro matured oocytes were fertilized with fresh spermatozoa for 24 h and in vitro cultured for 7 d. At the end of the IVC period, embryonic development was assessed by microscopic observation at 24 h intervals and fixed for staining by aceto-orcein staining method after 7 days. In comparison relating in IVM and IVF rates, larger oocytes have higher maturation (P<0.05) and cleavage (P<0.01) rates than the smaller ones. Unfortunately, none of the oocyte was reached to morula or blactocyst stage in both groups. In conclusion, it is demonstrated that the oocyte diameter may be a helpful selection criteria for dog in vitro embryo production.Öğe Effect of transport and storage temperature of ovaries on in vitro maturation of bitch oocytes(Elsevier Science Bv, 2006) Tas, Muzaffer; Evecen, Mithat; Ozdas, Ozen Banu; Cirit, Umut; Demir, Kamber; Birler, Sema; Pabuccuoglu, SerhatIn this study, the effects of ovary transport and storage temperature on in vitro maturation of bitch oocytes were investigated. Ovaries were collected from 23 mature bitches and one randomly selected ovary of each pair (n = 23 pairs) was transported in physiologic saline at 4 degrees C, while the other one at 35-38 degrees C for 2-4 h. A total of 316 cumulus oocyte complexes (COCs) were obtained from the 4 degrees C group and 301 COCs from the 35-38 degrees C group. All COCs were matured in modified synthetic oviduct fluid (mSOF) supplemented with follicle stimulating hormone (FSH), essential and non-essential amino acids at 38 degrees C in ahumidified 5% CO2, 5% O-2, and 90% N-2 atmosphere for 72 h. At the end of the in vitro maturation period, nuclear maturation of oocytes was classified as germinal vesicle (GV), germinal vesicle breakdown (GVBD), metaphase I (MI), metaphase II (MII), undetermined nuclear maturation (UDNM), and MI + MII. The nuclear maturation rates to MI, MII, and MI + MII stages were 60.44%, 10.75%, and 71.20% in the 4 degrees C group and 37.20%, 7.64%, and 45.85% in the 35-38 degrees C group, respectively. The data demonstrated that oocytes obtained from ovaries transported at 4 degrees C had higher maturation rates than from the ones transported at 35-38 degrees C (p < 0.001). (c) 2005 Published by Elsevier B.V.Öğe Effects of cooling rate on membrane integrity and motility parameters of cryopreserved ram spermatozoa(2015) Pabuccuoğlu, Serhat; Cirit, Ümit; Demir, Kamber; Birler, Sema; Bozkurt, H. Hakan; Ak, Kemal; Bakırer, Gül ÖztürkBu çalışmada koç spermasının 26°Cden +5°Cye indirilmesinde farklı soğutma hızlarının (0.3°C/dk., 0.6°C/dk. ve 0.9°C/dk.) eritme sonrasıspermatolojik özellikler ve spermatozoonların ultrastrüktürel yapısı üzerindeki etkilerinin incelenmesi amaçlanmıştır. Altı adet koçtan elektroejakülatörle alınan spermalar 26°Cdaki su banyosunda pooling işlemine tabii tutuldu. Tris bazlı sulandırıcıyla sulandırılan birleştirilmiş sperma üç eşithacme bölünerek 3 farklı hızda (0.3, 0.6 ve 0.9°C/dk.) +5°C ye soğutuldu. Sperma iki basamakta sulandırıldı, gliserol sperma ısısının +5°Cye indiğiikinci basamakta katıldı. Sulandırma sonrası sperma 1 saat ekilibre edildi daha sonrasında 0.25 ml payetlere çekilerek sıvı azot buharında donduruldu.Sperma pooling, sulandırma, soğutma, ekilibrasyon ve eritme sonrası gibi tüm aşamalarında motilite değerleri Bilgisayar Destekli Analiz Sistemleri(CASA) ile değerlendirildi. Pooling ve soğutma sonrasında elektron mikroskop incelemeleri gerçekleştirildi. 0.3°C/dk. soğutma grubunun, spermanın+5°Cye soğutma, ekilibrasyon ve eritme sonrasındaki hem total motilite hem de progressive motilite değerleri 0.9°C/dk. soğutma grubuna göre önemliderecede yüksek bulundu (P<0.05). Bu grup 0.6°C/dk soğutma hızı ile karşılaştırıldığında ise, 0.3°C/dk. soğutma grubunun soğutma ve ekilibrasyonsonrasındaki total motilite değerleri yüksek bulundu ancak eritme sonrası gruplar arasında fark bulunmadı (P>0.05). Soğutma ve eritme sonrasında iseprogressive motilite değerleri daha yüksek bulunurken(P<0.05), ekilibrasyon aşamasında progresif motilite değerleri arasında fark bulunmadı(P>0.05).Yapılan TEM incelemesinde, tüm soğutma hızı gruplarında eritme sonrasında tespit edilen toplam hasarlı spermatozoit oranı, pooling sonrasına göreönemli derecede yüksek bulunmuştur (P<0.05). Sonuç olarak koç spermasının dondurulması öncesinde +5°Cye soğutulmasında 0.3°C/dk.nın üzerindesoğutma hızlarının kullanılmasının sperma kalitesini olumsuz etkilediği ve soğutma hızı 0.6 ve 0.9°C/dk.ya arttırıldıkça eritme sonrası total ve progresifmotilitenin artan oranlarda etkilendiği sonucu çıkarılmıştır. Ayrıca, koç spermasında düşük sıcaklara bağlı olarak oluşan ultra strüktürel hasarların ilksulandırma aşamasından itibaren başladığı ve ultra strüktürel hasarların, spermanın gördüğü işleme ve soğutma hızlarına göre başın farklı bölgelerindelokalize olma eğiliminde olduğu sonucu çıkarılmıştır.Öğe Effects of Cooling Rate on Membrane Integrity and Motility Parameters of Cryopreserved Ram Spermatozoa(Kafkas Univ, Veteriner Fakultesi Dergisi, 2015) Demir, Kamber; Bakirer Ozturk, Gul; Cirit, Umit; Bozkurt, H. Hakan; Aktas, Abit; Birler, Sema; Ak, KemalIn this study we aimed to determinate the effects of three different cooling rates from +26 degrees C to +5 degrees C at (0.3 degrees C/min 0.6 degrees C/min and 0.9 degrees C/min) on spermatologic and ultrastructure properties of ram semen. For this purpose semen from 6 rams was collected by electroejaculator and was pooled in a +26 degrees C waterbath. Pooled semen was diluated with tris based extender and divided into three equal parts according cooling rates (0.3 degrees C/min., 0.6 degrees C/min. and 0.9 degrees C/min). Cooled semen was reextended with extender B +5 degrees C in the second step. Diluated samples were equilibrated for 1 h and then were loaded in 0.25 mL straws and freezed in liquid nitrogen vapor. After each freezing stage semen was evaluated motility with computer-assisted semen analysis (CASA). Electron microscobic evaluation was done for pooled and chilled samples. It has been observed that 0.3 degrees C/min. cooled group had meaningfully higher values of motility and progressive motility at +5 degrees C after equilibration and post-thaw stages when compared with the 0.9 degrees C/min. group (P<0.05). When compared to the 0.6 degrees C/min., the 0.3 degrees C/min. cooled group had higher total motility values at after cooling to +5 degrees C (P<0.05), equilibration (P<0.05) and post thaw stages (P>0.05) and had higher progressive motility at after cooling to + 5 degrees C (P<0.05), equilibration (P>0.05) and post-thaw stage (P<0.05). The TEM evaluation showed that at cooling to the +5 degrees C increases the total damaged spermatozoa in all groups (P<0.05). In conclusion, cooling the ram semen to +5 degrees C with a rate above 0.3 degrees C/min. affected negatively the spermatological characteristics. Reaching the cooling rates of 0.6 and 0.9 degrees C/min. increasingly deteriorated the post-thaw motility and progressive motility values. Also, low temperature related to ultrastructural damage was observed at the first dilution step and localized at different regions of the sperm head depends upon the processes and cooling rates.Öğe Effects of estrous cycle stage and transport temperature of ovaries on in vitro maturation of canine oocytes(Elsevier Science Bv, 2010) Evecen, Mithat; Cirit, Uemuet; Demir, Kamber; Ozdas, Ozen Banu; Tas, Muzaffer; Birler, Sema; Pabuccuoglu, SerhatUnlike other domestic animals, in vitro maturation (IVM) of canine oocytes still has limited success. The present study investigated the effects of estrous cycle stage and transport temperature of ovaries on in vitro maturation of canine oocytes. The donor bitches were categorized into three groups based on stage of estrus cycle: follicular (proestrus or estrous), luteal (diestrus) and anestrus. One ovary of each pair collected from 39 mature bitches was transported in Phosphate Buffer Saline (PBS) at 4 degrees C while the other was transported at 37 degrees C. A total of 1138 Grade I COCs obtained from all ovaries were grouped and matured in modified synthetic oviduct fluid (mSOF) supplemented with follicle stimulating hormone (FSH), luteinizing hormone (LH), essential and non-essential amino acids at 38.5 degrees C in a humidified 5% CO2, 5% O-2, and 90% N-2 atmosphere for 72 h. The nuclear maturation rates were evaluated by aceto-orcein staining. Oocytes harvested from follicular and luteal ovaries have a significantly higher maturation rates (MI+MII) than the oocytes from anestrual ovaries in the 37 degrees C group (p<0.05). However, oocytes harvested from anestrual ovaries transported at 4 degrees C had the highest maturation (MI + MII) rate, and the difference between anestrual and luteal ovary groups was significant (p < 0.05). The oocytes from anestrual ovaries transported at 4 degrees C have significantly higher maturation rates than those transported at 37 degrees C (p < 0.0001). However, the transport temperature (37 or 4 degrees C) did not significantly affect the maturation (MI + MII) rates of oocytes harvested from the luteal (p = 0.61) and follicular (p = 0.48) stage ovaries. It can be concluded from this study that (1) both transport temperature and transport temperature x estrus cycle stage interaction effected the maturation rates, while estrus cycle stage alone did not, and (2) transporting canine ovaries at 4 degrees C can improve in vitro maturation rates in oocytes harvested from anestrous ovaries. (C) 2009 Elsevier B.V. All rights reserved.Öğe Effects of serum starvation and ionomycin activation on the development of somatic cell nuclear transfer embryos in sheep(Chartered Inst. of Building Services Engineers, 2019) Demir, Kamber; Pabuccuoǧlu, Serhat; Cirit, Ümüt; Evecen, Mithat; Karaman, Elif; Özdaş, Özen Banu; Alkan, S.; Atalla, Hatem; Birler, SemaSynchronization of donor cells and activation of the reconstructed oocytes are important factors affecting the success rate in somatic cell cloning. In this study, it was aimed to investigate the effects of serum starvation in donor cell synchronization and ionomycin treatment in the activation of reconstructed oocytes after somatic cell nuclear transfer in Kıvırcık sheep. Cumulus cells were obtained from a slaughtered sheep ovaries and used as donor cells after serum starvation for 4 days (0.5% FCS; SS) or without serum starvation (10% FCS; S). After reconstruction, oocytes were activated by ionomycin for 5 min plus 6-dimethylaminopurine for 3 h (I+) or only with 6-dimethylaminopurine for 3 h (I-). All cleaved embryos (n= 44) at the second day of in vitro culture were transferred into synchronized recipient ewes (n= 10). Cleavage rates of the embryos were 37.3, 44.1, 34.6 and 44.7% in SS/I+, S/I+, SS/I- and S/Igroups, respectively. Recipient ewes had serum progesterone levels >1 ng/ml at 18th day were 33.3, 50.0, 50.0 and 100.0%, respectively. Only one pregnancy in the S/I- group continued after 40 days however the cloned lamb (7.1%, regarding to embryos transferred) died 10 days before term due to a maternal problem (uterine torsion). The results of this study reveal that somatic cell synchronization by serum starvation and ionomycin treatment for the activation of oocytes can be omitted for the success of somatic cell nuclear transfer in sheep.Öğe The effects of the thiolation with thioglycolic acid and L-cysteine on the mucoadhesion properties of the starch-graft-poly(acrylic acid)(Elsevier Sci Ltd, 2017) Gok, M. Koray; Demir, Kamber; Cevher, Erdal; Ozsoy, Yildiz; Cirit, Umut; Bacinoglu, Suleyman; Ozgumus, SaadetThe aim of this study is to investigate the effects of the thiolation on the mucoadhesion characteristics of the gelatinized and crosslinked wheat starch-graft-poly(acrylic acid) [(WS-g-PAA)(gc)] for potential use in drug delivery via vaginal route. Thiolation of (WS-g-PAA)(gc) was first time realized using L-cysteine hydrochloride monohydrate (CyS) and thioglycolic acid (TGA). These conjugates [(WS-g-PAA)(gcth)] were characterized using FTIR. The free SH group, mucoadhesion, cytotoxicity characteristics and the mechanism of the thiolation were also evaluated. To obtain fundamental data for possible application such as drug carrier, in vitro and in vivo progesterone release profiles from the mucoadhesive tablet formulations were also determined. The results showed that, vaginal tablet containing (WS-g-PAA)(gc)-TGA, which has not contain free SH groups in its structure, displays higher mucoadhesion than (WS-g-PAA)(gc) and (WS-g-PAA)(gc)-CyS. This tablet formulation can also be used as a drug carrier in vaginal applications. (C) 2017 Elsevier Ltd. All rights reserved.Öğe Evaluation of short estrus synchronization methods in dairy cows(Elsevier, 2008) Cirit, Uemuet; Bacinoglu, Suleyman; Tas, Muzaffer; Demir, Kamber; Bas, Ahmet; Ak, Kemal; Ileri, Irfan KamuranIn the present study, two new short estrus synchronization methods have been developed for lactating dairy cows. The study was completed in three consecutive phases. In experiment (Exp) 1, 32 cows, that were not detected in estrus since calving between the 50th and 84th post-partum days, were treated with PGF2 alpha (PGF, D-Cloprostenol, 0.150 mg), estradiol propionate (EP, 2 mg) and GnRH (lecirelina, 50 mu g) at 24 h intervals, respectively, and timed artificial insemination (TAI) was performed 48 h after PGF. Different from Exp 1, EP and GnRH were given at 48 and 60 h, respectively after PGF in Exp 2 (n = 20), instead of 24 and 48 h. Ovulations were investigated by ultrasound for 7 days starting from the day of PGF treatment, and ovulation rates were compared with the ones obtained in Exp 1. In Exp 3, cows were given the same treatments as Exp 2, but treatments started at certain estrus stages. Cows detected in estrus and with a confirmed ovulation (n = 27) after the second PGF given 11 days apart were assigned to three treatment groups. Treatment was initiated at Day 3 (group metestrus, n = 9), Day 12 (group diestrus, n = 9) and Day 18 (group proestrus, n = 9) after ovulation. All cows included in Exp 3 were TAI between 16 and 20 h after GnRH treatment. In Exp 2 and 3, blood samples were obtained once every 2 days, starting from Day 0 to the 10th day after GnRH injection, and once every 4 days between the 10th and the 22nd days after GnRH to examine post-treatment luteal development. During the study, animals exhibiting natural estrus were inseminated and served as controls (n = 85). The rate of estrus was found to be significantly higher in cows with an active corpus luteum (CL) at the start of Exp 1 (72.7% vs. 30.0%, P < 0.05) and the pregnancy rate tended to be higher than cows without an active CL (40.9% vs. 10.0%, P = 0.08). Compared to those in Exp 1, cows in Exp 2 had higher rates of synchronized ovulation (94.1% vs. 59.1%, P = 0.013), In Exp 3, estrus (P < 0.001) and pregnancy rates (P = 0.01) were found to be significantly higher in cows in the proestrus group than in those in the metestrus group. Comparable pregnancy rates were obtained from the first and second inseminations in Exp 1 and 3 with results from those inseminated at natural estrus (P > 0.05). It was concluded from the study that the treatment in Exp 1 and 3 could result in comparable pregnancy rates after timed Al of lactating dairy cows at random stages of the estrus cycle relating to those inseminated at natural estrus, but the stage of the estrus cycle can have significant effects on pregnancy rates. (C) 2007 Elsevier B.V. All rights reserved.Öğe Evaluation of synergic effects of iodixanol and trehalose on cryosurvival of electroejaculated ram semen(Wiley, 2020) Ozmen, Mehmet Ferit; Cirit, Umut; Arici, Ramazan; Demir, Kamber; Kurt, Dogan; Pabuccuoglu, Serhat; Ak, KemalThe primary aim of the study was to investigate whether iodixanol and trehalose would have a synergic effect on the cryosurvival of electroejaculated ram semen. Tris-based diluter was used to prepare 9 different extenders by the addition of iodixanol or trehalose alone or varying combinations of these substances. Diluters were prepared as follows: Tris (control), Io5 (5% iodixanol), Tr25 (25 mmol/L trehalose), Tr50 (50 mmol/L trehalose), Tr50 + Io1.25 (50 mmol/L trehalose and 1.25% iodixanol), Tr50 + Io2.5 (50 mmol/L trehalose and 2.5% iodixanol), Tr50 + Io5 (50 mmol/L trehalose and 5% iodixanol), Tr25 + Io5 (25 mmol/L trehalose and 5% iodixanol) and Tr12.5 + Io5 (12.5 mmol/L trehalose and 5% iodixanol). Supplementation of the freezing extender with trehalose or iodixanol alone supported the protection of both morphological and functional integrity of ram spermatozoa and total motility at 1 and 4 hr post-thawing respectively. However, beyond these positive effects, the combination of trehalose (25 mmol/L) and iodixanol (5%) significantly increased post-thaw sperm longevity and motion properties at the end of 4-hr incubation. The results of the study clearly showed that there was positive synergic effect of iodixanol and trehalose on cryosurvival of ram semen.Öğe İn vitro elde edilen sığır embriyolarının dondurulmasında vitrifikasyon medyumuna maruz kalma sürelerinin çözünme sonrası gelişim üzerine etkisi(2012) Pabuccuoğlu, Serhat; Alkan, Serhat; Demir, Kamber; Ak, Kemal; Cirit, Ümüt; Özdaş, Özen Banu; İrez, TülayGünümüzde dondurulmuş embriyoların transferi ile üstün verim özelliklerine sahip sürülerin oluşturulması, hastalıkların kontrolü ve genetik materyallerin uzun süre saklanması mümkün olabilmektedir. Ancak donmuş embriyolarda eritme sonrası tranfer edilebilir embriyo eldesi ve gebelik oranları istenilen düzeyde değildir. Özellikle embriyoların dondurulması sırasında dejenerasyonlar oluşmaktadır. Dejenerasyonların, embriyoların kriyoprotektif maddelere maruz kalma süreleri ile toksik etkilerinden meydana geldiği düşünülmektedir. Çalışmada mezbahada kesilen sığırların ovaryumları kullanıldı. Aspirasyon yöntemi ile elde edilen oositler (1290 adet) TCM-199 da 22-24 saat süreyle %5 CO2, %5 O2, %90N2 gaz atmosferinde 38,8 ºC de in vitro olarak olgunlaştırıldılar. Olgun oositler IVFTALP medyumunda 18-24 saat süreyle fertilize edildiler. Fertilizasyon sonrası 48. saatte cleavage %67,05 (865/1290) saptandı. Embriyolar %10 FCS’li SOF medyumunda 7 gün süreyle %5 CO2, %5 O2, %90N2 gaz karışımında blastosist (%34,91; 302/865) aşamasına kadar inkübe edildiler. Erken blastosist-blastosist aşamasına ulaşan 302 adet embriyodan 254 tanesi vitrifikasyon solüsyonunda farklı sürelere (15, 30, 60, 90 sn) maruz bırakılarak donduruldular. Bu amaçla sırasıyla 4 grup oluşturuldu. Her grup sırasıyla 67, 64, 63 ve 60 embriyo içerdi (Grup 1, 2, 3, 4). Embriyolar önce % 10 Gliserol + % 10 FCS içeren PBS solüsyonunda (Vs1) 5 dk, sonra %10 Gliserol + %10 FCS+ %20 Etilen Glikollü PBS’de (Vs2) 5 dk. bekletildiler. Daha sonra embriyolar payet içindeki %25 Gliserol + %20 Etilen Glikol + %10 FCS + 0,1 M Sükroz vitrifikasyon solüsyonuna (Vs3) aktarıldılar ve değişik sürelere (15, 30, 60, 90 sn) maruz bırakıldıktan sonra sıvı azota daldırılarak donduruldular. Eritme sonrası (37ºC) embriyolar birkaç kez yıkama medyumunda ve SOF medyumunda yıkandıktan sonra, her bir gruba ait embriyolar 48 saat süreyle tekrar inkübe edildiler. Çalışmada istatistiki analizde ki-kare testi kullanıldı. Eritme sonrası, genişlemiş blastosist-zonadan çıkma safhasında en iyi gelişim %52,2 (35/67) ile Grup 1 de saptanırken, bunu %45,3 (29/64) ile Grup 2, %22,2 (14/63) ile Grup 3 ve %5 (3/60) ile Grup 4 takip etti. Grup I ve II arasında istatiksel bir fark bulunmadı. Grup 1 ile Grup 3 arasındaki istatistiksel fark P<0,01, Grup 1 ile Grup 4 arasında ise P<0,001 düzeyinde anlamlı bulundu.
