Post-ejaculation of sperms into the female reproductive tract, acquisition of sperm capacitation is an essential step in the fertilization process. Accordingly, during in-vitro fertilization, the successful fertilization requires necessarily induction of capacitation in the retrieved sperms. To date, many candidate substances have been considered as capacitation inducers. However, there were no reports about the comparison of efficiency inducing sperm capacitation among diverse capacitation inducers. Therefore, we tried to determine an inducer showing the best capacitation performance in mouse sperms by comparing the preimplantation development of in-vitro-fertilized embryos using sperms experiencing capacitation by a variety of capacitation inducers. For these, calcium, progesterone, bovine serum albumin (BSA), heparin, lysophosphaticylcholine (Lyso-PC) were used as candidate capacitation inducers. Optimized concentration of each inducer were determined by accessing ratio of sperms experiencing acrosome reaction using coomassie G-250 blue staining. Subsequently, in vitro fertilization was performed using sperms incubated in each optimized concentration inducer. The ratio of fertilized oocytes was observed. As the results, Calcium at 2.7 mM and 0.3% (w/v) BSA showed the highest fertilization rates compared to 15 μM progesterone, 50 mM heparin, and 100 μM Lyso-PC. From these results, we found that 2.7 mM calcium and 0.3% (w/v) BSA were the most effective sperm capacitation inducers of mouse sperm for in vitro fertilization. From these results, we could identify that, among diverse sperm capacitation inducers, 2.7mM calcium and 0.3% (w/v) BSA were the most effective inducers for in vitro fertilization.

This study was carried out to elucidate whether sperm contain a factor inducing second polar body extrusion and to search for an effective collection method of the sperm factor Thus, sperm extract, dialyzed sperm-extract or liquid chromatographic fractions of sperm extract was microinjected into ovulated oocytes. And the microinjected oocytes were incubated for 24 hours to investigate about the extrusion of second polar body. The results obtained were as follows; 1. Sperm extract significantly increased the second polar body extrusion. 2. Sperm extract showed five major fractions at retention volumes (RVs) 1.25, 1.37, 1.84, 2.10 and 2.67ml after separation with Superose 12 column. These sperm extract fractions did not significantly increase the second polar body extrusion. 3. Dialyzed sperm-extract significantly increased the second polar body extrusion 4. Dialyzed sperm-extract showed three maior fractions at RVs 1.88, 2.14 and 2.77ml after separation with Superose 12 column. Of these fractions, the fraction RV2.14 significantly increased the second polar body extrusion. In conclusion, sperm extract contained a factor inducing the second polar body extrusion and the factor was contained largely in fraction RV2.14 after dialysis and liquid chromatographic fractionation of sperm extract.

Interleukin(IL)은 cytokines의 group으로 주로 macrophages에 의해 생산되어지며, 그 중 IL-1 system 은 염증반응과 면역 및 항상성을 조절한다. Interleukin-1은 Interleukin-1α(IL-1α)와 Interleukin-1β (IL-1β) 2개의 isoform이 존재하며, 이는 IL-1Receptor1(IL-1R1)과 IL-1Receptor2(IL-1R2) 두 가지 타입의 receptor에 결합하여 작용한다. 또한 IL-1R1의 신호전달에는 IL-1 receptor accessory protein (IL-1RAcP)가 필요로 한다. 또한 IL-1RAcP와 IL-1R1은 Toll interacting protein(TOLLIP)과도 작용한 다고 알려져 있다. Interleukin-1 cytokine family member 중 Interleukin-1 receptor antagonist(IL-1Ra)는 IL-1α와 IL-1β의 작용을 저해한다. 선행된 다른 연구에서는 IL-1의 과발현이 정자형성에 대한 부정 적인 효과를 주는 것이 밝혀진바 있다. 따라서 본 연구에서는 정자에서 IL-1α, IL-1β, IL-1R1, IL-1RAcP와 TOLLIP의 발현 여부와 IL-1α, IL-1β와 IL-1Ra가 정자에 미치는 영향을 알아보고자 하였으며, 정자에게 미치는 영향 중에서도 정자의 운동성에 관여한다고 알려져 있는 GSK3의 억제성 인산화 형태인 p-GSK3에 대해서 알아보고자 하였다. 실험 결과, 생쥐의 정자에서 IL-1α, IL-1β, IL-1R1, IL-1RAcP와 TOLLIP이 모두 정자에서 발현되는 것을 확인하였다. Western blot 실험 결과, IL-1α, IL-1β가 단독 처리된 생쥐 정자에서 p-GSK3의 발현이 증가한 것을 확인하였다. 그리고 IL-1Ra가 처리된 경우 p-GSK3의 발현이 생쥐 정자에서 대조군에 비해 감소한 것을 확인하였다. IL-1 α, IL-1β를 IL-1Ra와 함께 처리 시에는 IL-1α, IL-1β를 단독처리 했을때 보다 생쥐의 정자에서 p-GSK3의 발현이 감소한 것을 확인하였다. 이러한 결과를 토대로 IL-1과 IL-1R1이 정자에서 발현되며, IL-1 및 IL-1Ra 처리시 p-GSK3의 발현에 영향을 미치는 것으로 보아 IL-1이 정자운동성에도 관여하는 것으로 사료된다.

Glycan epitopes of cellular glycoconjugates act as versatile biochemical signals, and this sugar coding plays an important role in cell-to-cell recognition processes. In the present study, our aims were to determine the distribution of sperm receptors with activity for fucosyl- and galactosyl glycans and to address whether mono sugar neoglycoproteins functionally mimic the binding between zona pellucida (ZP) glycoproteins and sperm. In mouse epididymal spermatozoa with intact acrosomes, fucopyranosyl bovine serum albumin (BSA-Fuc) bound to the segment of the acrosome, the equatorial segment, and the postacrosome region of the sperm head. Galactosyl BSA (BSA-Gal) binding activity was similar to that of BSA-Fuc, but was weaker. In acrosome-reacted spermatozoa treated with the Ca2+ ionophore A23187, BSA-Fuc binding was lost in the apical segment of the acrosome but remained in the equatorial segment and postacrosome regions. BSA-Gal binding to the equatorial region was increased. In the presence of 2.5 μg/ml BSA-Fuc, in vitro sperm - ZP binding was significantly decreased, indicating that fucosyl-BSA functionally mimics ZP glycoproteins during sperm-egg ZP interactions. At the same concentration, BSA-Gal was not effective. Fucosyl BSA which efficiently inhibited the sperm-ZP binding can mimic the ZP glycoconjugate and has potential for use as a sperm fertility control agent in mouse.

정자의 수정능력획득과 첨체반응은 Ca에 의존적으로 일어나며, 수정능력획득 과정에서는 원형질막으로부터 cholesterol이 방출되어 첨체반응이 일어나기 쉬운 상태로 전환된다. 최근 세포막으로부터 cholesterol 방출을 촉진하는 methyl beta cyclodextrin (MBCD)이 첨체반응을 유발함이 알려졌으나 정자 주변의 Ca 농도와 관계없이 cholesterol 방출만으로 첨체반응이 유발되는지의 여부는 확인되지 않았다. 본 연구에서는 생쥐

생쥐 정자의 수정능력획득과 첨체반응에 작용하는 /H antiporter의 역할을 조사하고자 하였다. /H antiporter를 특이적으로 억제하는 dimethyl amiloride는 정자의 자발적인 첨체반응을 농도 의존적으로 억제한 반면 난포액 및 calcium ionophore인 A23l87에 의해 유도된 첨체반응은 억제하지 못하였다. 이러한 결과는 정자내 /H antiporter에 의한 1가이온의 출입과 이에 따른 세포질내 pH 조절이 정자의 수정능