Dondurulmuş tavşan spermatozoon separasyonu ve dölverimi

Bu çalışmanın amacı; dondurulmuş çözdürülmüş tavşan spermasında glass wool filtrasyon (GWF), swim-down (SD) yöntemleri ile glass wool filtrasyon ve swim - down yöntemlerinin kombinasyonunun (GWFSD), spermatolojik parametreler ve dölverimi üzerine olan etkilerinin araştırılmasıdır. Dondurulmuş-çözdürülmüş tavşan spermaları 4 eşit hacme bölünmüştür. Intrauterin tohumlama (IUT) öncesi spermalardan üçüne sırasıyla GWF tGrup I), SD (Grup II) ve GWFSD (Grup III) uygulanmıştır. Dördüncü spermalar kontrol grubu olarak (Grup IV) direk IUT'da kullanılmıştır. Separasyon öncesi ve sonrası spermatolojik örnekler alınmış ve motilite, ölü/ canlı spermatozoa oranı ve yoğunluk parametreleri değerlendirilmiştir. En yüksek ortalama motilite ve en düşük ölü spermatozoa oranı SD yöntemi sonunda elde edilirken, en yüksek yoğunluklar GWF yöntemi sonunda elde edilmiştir (P < 0,05). Gebelik oranları kontrol grubunda, GWF, SD ve GWFSD sonunda-, sırasıyla; % 70, % 50, % 70 ve % 60 olarak belirlenmiş ve aralarında önemli bir fark bulunamamıştır. Çalışma sonunda dondurulmuş çözdürülmüş tavşan spermasına uygulanan separasyon yöntemleri arasında SD yönteminin spermatolojik parametreler açısından diğer iki yönteme (GWF ve GWFSD) göre daha etkin olduğu sonucuna varılmıştır. Bununla birlikte, gebelik oranları açısından separasyon yöntemleri arasında istatistiki bir farka rastlanmaması bu konu üzerine daha fazla araştırma yapılmasına ihtiyaç olduğunu göstermektedir.

Spermatozoon separation and fertility of frozen-thawed rabbit semen

The aim of this study was to evaluate the effect of glass wool filtration (GWF), swim - down (SD) and combination of glass wool filtration and swim - down techniques (GWFSD) on frozen - thawed rabbit semen parameters and fertility. Frozen - thawed rabbit semen were divided into four equal aliquots. Before intrauterine insemination (IUI) 3 of 4 aliquits were processed by GWF (Group I), SD (Group II) and GWFSD (Group III) techniques. Fourth aliquits were used directly for IUI as a control group (Group IV). Semen samples were taken before and after sperm separation techniques and motility, live-dead spermatozoa ratio and concentration parameters were evaluated. While the highest mean percentage of motility and lowest live-dead spermatozoa ratios were detected after SD technique (P < 0,05), the highest concentrations were obtained by GWF (P < 0,05). Pregnancy rates in the control group, after GWF, SD and GWFSD were detected as 70%, 50 %, 70 % and 60 % respectively (P > 0,05). In conclusion; swim - down technique was found to be more effective in improving the quality of frozen thawed rabbit semen then the other two tecniques (GWF and GWFSD). However there was no significant difference in the pregnacy rates between separation techniques. We have concluded that, further studies need be carried out on this subject.

PDF

___

1. Al-Hasani S, Alpüstün S, Ludwig M, Diedrich K, Bauer O, Küpker W, Wolff A, Krebs D, 1996. The combination of two semen preparation techniques (glass wool filtration and swim-up) and their effect on the morphology of recovered spermatozoa and outcome of İVF - ET. Int J Androl, 19 : 55 - 60.
2. Almagor M, Benbenishti D, Rosenak D, Mogle Y, Yaffe H, 1993. Simultaneous swim-up, swimdown of sperm in assisted reproduction procedures. J Assist Reprod Genet, 10 : 261 -265.
3. Babbo CJ, Hecht MD, Jeyendran RS, 1999. Increased recovery of swim-up spermatozoa by application of 'antigravitational' centrifugation. Fertil Steril, 72:556-558.
4. Baguhl VF, Flieb R, Bernt D, 1989. Effekt der glaswollfiltration auf humane spermatozoen ein vergleich mit der swim up technik. Zentb Gynakol, 111: 1613-1616.
5. Carell DT, Kuneck PH, Peterson M, Hatasaka HH, Jones KP, Campbell BF, 1998. A randomized, prospective analysis of sperm preparation techniques before intrauterine insemination of husband sperm. Fertil Steril., 69: 122 - 126.
6. Chapeau C, Gagnon C, 1987. Nitrosellulose and polyvinyl coatings prevent sperm adhesion to glass without affecting the. motility of intact and demembranated human spermatozoa. J Androl., 8:34-40.
7. Coetzee K, Erasmus EL, Kruger TF, Menkveld R, Lombard CJ, 1994. Glass wool filter preparation of cryopreserved spermatozoa. Andrologia, 26 : 33 - 34.
8. Daya S, Awatkin BL, 1987. Improvement in semen quality using glass bead column. Arch Androl, 18:241-244. ,
9. Dickey R, Pyrzak R, Taylor S, Rye PH, Lu PY, Belinda SM, K, 2001. Decision to do IUI or İVF sperm count? Fertil Steril, 76 : 1086.
10. Dickey RP, Pyrzak R, Lu P, Taylor S, Rye P, 1999. Comprasion of the sperm quality necessary for successful intrauterine insamination with world health orgnization threshold values for normal sperm. Fertil Steril, 71 : 684 - 689.
11. Drobnis EZ, Zhong CQ, Overstreet JW, 1991. Seperation of cryopreserved human semen using sephadex columns, washing, or percoll gradients. J Androl, 12 (3): 201-208.
12. Engel S, Weber H, Petzoldt R, Seidl B, Wieher W, Sperl J, 2001. An improved method of sperm selection by glass wool filtrasyon. Andrologia., 33 :223 - 230.
13. Gonzales GF, Pella RE, 1993. Swim-down: A rapid and easy method to select motile spermatozoa. Arch Androl, 30 : 29 - 34.
14. Hall JA, Fishel SB, Timson JA, Dowell K, Klemtzeris LD, 1995. Human sperm morphology evaluation pre-and post-Percoll gradient centrifugation. Hum Reprod., 10 (2) : 342-346.
15. Hammadeh, ME, Kuhnel, A, Stieber, M, Rosenbaum, P, Schmidt W, 1998. Chromation condensation and morphology recovery rate after sperm selection with Swim - up, percoll and glass wool techniques and their influence on ivf outcome. Fertil Steril, 70 (2S) : 492.
16. Hammadeh ME, Kühnen A, Amer AS, Rosenbaum P, Schmidt W, 2001. Comparison of sperm preparation methods : effect on chromatin and morphology recovery rates and their consequences on the clinical outcome after in vitro fertilizasyon embryo transfer. Int J Androl, 24 : 360 - 368.
17. Jeyendran RS, Perez M, Crabo BG, 1986. Concentration of viable spermatozoa for artificial insemination. Fertil Steril, 45 : 132 -134.
18. Johnson DE, Confino E, Jeyendran RS, 1996. Glass wool colummn filtration versus mini - percoll gradient for processing poor quality semen samples. Fertil Steril., 66 : 459 - 462.
19. Katayama KP, Stehlig E, Jeyendran RS, 1989. In vitro fertilization outcome : glass wool - filtered sperm versus swim - up sperm. Fertil Steril, 52 : 670 - 672.
20. Katila, T, 2001 In vitro evaluation of frozenthawed stallion semen : arewiev. Actavet. Scand. 42:199-217.
21. Kobayaski T, Kaneko S, Hara I, Aoki R, Ohno T, Nozawa S, 1991. Swim- down sepatration of progressively motile sperm from poor quality human semen by the modified funnel procedure. Andrologia, 23 : 17 - 20.
22. Larson KL, Brannian JD, Timm BK, Lost LK, Evenson DP, 1999. Density gradient centrifugation and glass wool filtration of semen remove spermatozoa with damaged chromatin structure. Hum Reprod., 14(8): 2015-2019.
23. Levay PF, Fourie R, Bezuidenhout C, Koch A, 1995. Efectiveness of various sperm processing methods in removing seminal plasma from insemination media. Human Reprod., 10 : 2056 - 2060.
24. Makler A, Stoller J, Blumenfeld Z, Feigin PD, Brandes JM, 1993. Investigation in real time of the effect of gravitation on human spermatozoa and their tendency to swim-up and swim-down. Int. J. Androl, 16 : 251 -257.
25. Makler A, Stoller J, Makler - Shiran E,' 1998. Dynamic aspects concerned with the mechanism of seperating motile sperm from nonmotile sperm, leukocytes, and debris with the use of high density percoll gradients. Fertil Steril, 70 : 961-966.
26. Pereira RJTA, Tuli RK, Wallenhorst S, Holtz WK, 2000. The effect of heparin, caffeine and calcium ionophore a 23187 on in vitro induction of the acrosome reaction in frozen-thawed bovine and caprine spermatozoa. Theriogenology, 54 : 185-192.
27. Pyrzak R, Dickey RP, Taylor DN, Curole DN, Rye PH, Potts AS, 2001 Effectiveness of swimdown sperm preparation on the pregnancy outcome of intra uterine insemination and art procedures. Erişim: http://www.google.com.
28. Raymond MY, Li DQ, Harding AM, Hons B, Jones WR. 1991. A comparison of swim-down and swim-up methods for the extraction of high motility sperm. Fertil Steril, 55 : 817 - 819.
29. Rhemrev J, Jeyendran RS, Vermeidan JPW, Zanaveld LJD, 1989. Human sperm selection by glass wool filtration and two - layer, discontinous percoll gradient centrifugation. Fertil Steril, 51 : 685 - 690.
30. Rho GJ, 1998. Effect of sperm preparation technique on subsequent in vitro development of bovine embryos. J Emb Trans., 13(2) : 117 -125.
31. Rodriguez - Martinez H, Larsson B, Pertoft H, 1997. Evaluation of sperm damage and techniques for sperm clean - up. Reprod Fertil Dev., 9: 297 - 308.
32. Sanchez R, Stalf T, Khanaga O, Turley H, Gips H, Schill W, 1996. Sperm selection methods for intracytoplasmic sperm injection (ICSI) in andrological patients. J Assist Reprod Genet., 13 : 228 - 233.
33. Sherman JK, Paulson JD, Liu KC, 1981. Effect of glass wool filtration on ultrastructure of human spermatozoa. Fertil Steril, 36 (5) : 643 -647.
34. Szerman-Joly E, Sauvalle A, Izard J, Drossowsky M, 1989. Comparasion of two techniques osed to collect normal and motile sperm. Int J Androl, 12 : 199 - 205.
35. Valcarcel A, De Las HeraS MA, Moses DF, Perez LJ, Baldasarre H, 1996. Comparison between Sephadex G - 10 and Percoll for preparation of normospermic, asthenozoo spermic and frozen / thawed ram semen. Anim Reprod Sel, 41: 215 - 224.
36. Vicente JS, Viudes-De-Castro, M.P, 1996. A sucrose-DMSO extender for freezing rabbit semen. Reprod. Nutr. Dev., 36: 485 - 492.
37. Vijayakumar R, Ndubisi B, Prien S, Leon F, Heine , Wayne, 1986. Quantitative ultramorphological evaluation of swim-up spermatozoa used in human in vitro fertilization and transcervical intrauterine insemination. Arch Androl, 17 : 223 - 230.