Trombositten zengin plazma (TZP) ile muamele edilmiş semen örneklerinde oda ısısı (+23±2°c) ve soğuk ortam şartlarının (+4°c) sperm parametreleri ve kromatin yapısına etkilerinin değerlendirilmesi
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Dosyalar
Tarih
2019
Yazarlar
Dergi Başlığı
Dergi ISSN
Cilt Başlığı
Yayıncı
Necmettin Erbakan Üniversitesi Sağlık Bilimleri Enstitüsü
Erişim Hakkı
info:eu-repo/semantics/openAccess
Özet
Günümüzdeki yaşam şartlarında çeşitli birçok sebepten dolayı çiftlerin doğal yollarla gebelik elde etme şansları azalmaktadır. Gebelik elde etme imkanlarının sağlanabilmesi için birçok çiftin yardımcı üreme tekniklerine (YÜT) başvurması sonrasında yapılan semen analizi ile bireyin fertilizasyonda önemli rolü olan sperm motilite, konsantrasyon ve morfoloji gibi kriterleri değerlendirilmektedir. Bu kriterlere ek olarak fertilizasyonun başarılı bir şekilde gerçekleştirilmesi ve sağlıklı bir embriyonun gelişimi için sperm deoksirio nükleik asitinde (DNA) herhangi bir hasar bulunmaması ve DNA bütünlüğü oldukça önemlidir. Trombositler ise içerisinde yer alan granüllerin hemoostaz, inflamasyon, kemik ve yara iyileşmesinde görevli pek çok protein ve büyüme faktörünü içermesi trombositlerin yoğunlaştırılarak klinikte yara, kemik iyileşmesi gibi alanlarda kullanımının gündeme gelmesine neden olmuştur. Trombositlerden yoğunlaştırılarak elde edilen trombositten zengin plazma (TZP), kandan elde edilmesinin kolay olması ve gerçekleştirilen araştırmalarda herhangi bir yan etkisinin ortaya çıkmamış olması sebebiyle günümüzde kullanımı mevcut, alternatif tedavi yöntemlerinden biri olarak araştırılmaktadır. Çalışmamızda bu düşüncelere dayanarak basit yıkama yöntemine aktive edilerek hazırlanmış TZP (A-TZP) ilavesiyle hazırlanan semen örneklerindeki sperm motilite ve konsantrasyon değerlerinin değerlendirilmesi, hazırlanan örneklerin bir gece farklı ortam sıcaklıklarında saklanmasının sperm motilite parametreleri üzerine olası etkilerinin değerlendirilmesi, sperm DNA kromatin yapı hasarlarının uygulanan anilin mavisi boyaması ile incelenerek istatistiksel analizinin yapılmasıyla A-TZP'nin alternatif bir sperm yıkama medyum suplementi olarak kullanılabilirliğinin araştırılması amaçlanmıştır. Çalışmamızda 20 sağlıklı gönüllüden kan alınıp manuel olarak A-TZP hazırlanmıştır. Gönüllü hastalardan semen için onam formları alındı ve spermiogram analizi sonucunda azospermi durumu göstermeyen 20 hasta örneği çalışmaya dahil edildi. Her bir semen örneği fosfat tamponlu tuz (PBS) ile muamele edeceğimiz kontrol grubu ve A-TZP ile muamele edeceğimiz deney grubu olarak iki gruba ayrıldı. Gruplara basit yıkama yöntemi uygulandı ve elde edilen son örneklerin Makler kamarasında sayımları yapıldı ve sperm konsantrasyon ve motilite verileri kaydedildi. Bu işlemlerden sonra +23±2°C ve +4°C'de bir gece bırakılmak üzere deney ve kontrol grubundan ikişer örnek ayırıldı. Bir gece sonunda her bir gruptaki iki örneğin sperm konsantrasyon ve motilite verileri kaydedildi ve örnekler anilin mavisi ile boyanarak DNA kromatin yapıları incelendi. Örneklerin anilin mavisi boyamasından sonra sonuçları kaydedildi ve elde edilen tüm verilerin istatistiksel analizleri yapıldı. Yapılan değerlendirmede A-TZP ile muamele edilen deney grubu örnekleri hem basit yıkama işlemi sonrasında hem de bir gece saklama sonrasında kontrol grubu örneklerinin verileri ile karşılaştırıldığında motilite değerleri olarak daha anlamlı bulunmuştur (p<0.05). Deney grubu içerisinde +23±2°C'de saklanan örneklerin motilite bulguları +4°C'de saklanan örneklerin bulgularına göre daha iyi çıkmıştır. Bir gece sonrasında anilin mavisi boyama üzerine elde edilen verilere bakıldığında deney grubu +4°C'ye bırakılan örneklerde anilin mavisi negatif boyanan spermler daha fazla olup diğer örneklere göre daha olumlu bulunmuştur (p<0.05). Sonuç olarak; A-TZP kullanımının spermde hem motilite hem de sperm DNA kromatin yapısı üzerine artı yönde bir etkisinin olduğu çalışmamızla gösterilmiş oldu.
The chance of achieving pregnancy for couples with natural ways is decreased recently because of lots of various reasons under today's living conditions. After the appeal of lots of couples to assisted reproductive techniques for a possibility of achieving pregnancy, with semen analysis, the criteria like sperm motility, concentration and morphology which have a significant role in individual fertilization are evaluated. In addition to these criteria, it is very important that the sperm deoxyribio nucleic acid (DNA) is not damaged and that the integrity of DNA is very important for successful fertilization and development of a healthy embryo. Thanks to granules inside thrombocytes, platelet includes a lot of protein and growth factor which are responsible for homeostasis, inflammation, bone and wound healing, so the idea to use platelets in clinics to heal bone and wounds after it is concentrated became popular. Platelet-rich plasma (PRP) obtained by concentration from platelets is being investigated as one of the alternative treatment methods available today because it is easy to obtain from blood and no side effects have been observed in the studies performed. By considering these thoughts, in our work, the purposes are investigating the evaluation of sperm motility and concentration values in semen samples prepared by adding activated PRP(A-PRP) to simple washing method and the evaluation of possible impacts of prepared samples, which are waited a night in different ambient temperatures, on sperm motility parameters, also after statistical analysing of sperm DNA chromatin structure damages by implementing aniline blue painting, the usability of A-PRP as an alternative supplement for sperm washing medium. In our study, blood was drawn from 20 healthy volunteers and A-PRP was prepared manually. Consent forms for semen were obtained from volunteer patients and 20 patients who did not show azoospermia status were included in the study. Each semen sample was divided into two groups. First one is the control group treated with phosphate buffered saline (PBS) and the other is the experimental group treated with A-PRP. Simple washing method was applied to the groups and the final samples were counted in the Makler chamber, also sperm concentration and motility data were recorded. After these procedures, two samples were separated from the experimental and control groups by living theme overnight at +23±2°C and +4°C. At the end of the night, sperm concentration and motility data of two samples in each group were recorded and samples were stained with aniline blue to examine DNA chromatin structures. After the aniline blue staining of the samples, the results were recorded and all data were analysed statistically. In the evaluation, the experimental group samples treated with A-PRP were found to be more significant as motility values when compared with the data of the control group samples after both simple washing and overnight storage (p<0.05). Motility findings of samples stored at +23±2°C in the experimental group were better than those of samples stored at +4°C. After a night, when the data of aniline blue staining obtained, aniline blue negative stained sperm was higher in the samples that were left to +4°C in the experimental group and it was found to be more positive than the other samples (p <0.05). In conclusion, it has been shown in our study that the use of PRP has a positive effect on both motility and sperm DNA chromatin structure of sperm.
The chance of achieving pregnancy for couples with natural ways is decreased recently because of lots of various reasons under today's living conditions. After the appeal of lots of couples to assisted reproductive techniques for a possibility of achieving pregnancy, with semen analysis, the criteria like sperm motility, concentration and morphology which have a significant role in individual fertilization are evaluated. In addition to these criteria, it is very important that the sperm deoxyribio nucleic acid (DNA) is not damaged and that the integrity of DNA is very important for successful fertilization and development of a healthy embryo. Thanks to granules inside thrombocytes, platelet includes a lot of protein and growth factor which are responsible for homeostasis, inflammation, bone and wound healing, so the idea to use platelets in clinics to heal bone and wounds after it is concentrated became popular. Platelet-rich plasma (PRP) obtained by concentration from platelets is being investigated as one of the alternative treatment methods available today because it is easy to obtain from blood and no side effects have been observed in the studies performed. By considering these thoughts, in our work, the purposes are investigating the evaluation of sperm motility and concentration values in semen samples prepared by adding activated PRP(A-PRP) to simple washing method and the evaluation of possible impacts of prepared samples, which are waited a night in different ambient temperatures, on sperm motility parameters, also after statistical analysing of sperm DNA chromatin structure damages by implementing aniline blue painting, the usability of A-PRP as an alternative supplement for sperm washing medium. In our study, blood was drawn from 20 healthy volunteers and A-PRP was prepared manually. Consent forms for semen were obtained from volunteer patients and 20 patients who did not show azoospermia status were included in the study. Each semen sample was divided into two groups. First one is the control group treated with phosphate buffered saline (PBS) and the other is the experimental group treated with A-PRP. Simple washing method was applied to the groups and the final samples were counted in the Makler chamber, also sperm concentration and motility data were recorded. After these procedures, two samples were separated from the experimental and control groups by living theme overnight at +23±2°C and +4°C. At the end of the night, sperm concentration and motility data of two samples in each group were recorded and samples were stained with aniline blue to examine DNA chromatin structures. After the aniline blue staining of the samples, the results were recorded and all data were analysed statistically. In the evaluation, the experimental group samples treated with A-PRP were found to be more significant as motility values when compared with the data of the control group samples after both simple washing and overnight storage (p<0.05). Motility findings of samples stored at +23±2°C in the experimental group were better than those of samples stored at +4°C. After a night, when the data of aniline blue staining obtained, aniline blue negative stained sperm was higher in the samples that were left to +4°C in the experimental group and it was found to be more positive than the other samples (p <0.05). In conclusion, it has been shown in our study that the use of PRP has a positive effect on both motility and sperm DNA chromatin structure of sperm.
Açıklama
Yüksek Lisans Tezi.
Anahtar Kelimeler
Anilin Mavisi, Deoksiribo Nükleik Asit (DNA), Trombositten Zengin Plazma (TZP), Aniline Blue, Deoxyribo Nucleic Acid (DNA), Platelet-Rich Plasma (PRP)
Kaynak
WoS Q Değeri
Scopus Q Değeri
Cilt
Sayı
Künye
Karakuş, Gül. (2019). Trombositten zengin plazma (TZP) ile muamele edilmiş semen örneklerinde oda ısısı (+23±2°c) ve soğuk ortam şartlarının (+4°c) sperm parametreleri ve kromatin yapısına etkilerinin değerlendirilmesi. (Yayımlanmamış Yüksek Lisans Tezi). Necmettin Erbakan Üniversitesi, Sağlık Bilimleri Enstitüsü Histoloji ve Embriyoloji Anabilim Dalı, Konya.
