간이동결법에 의한 운동성 불량정액과 정자감소증 정액의 정자냉동보존 = Simple Method of Cryopreservation for Poor Motility and Oligozoospermic Sperm

국문 초록 (Abstract)

배우자간인공수정(AIH)시의 정자동결과 냉동보존의 응용을 목적으로, 융해시의 정자소생율 향상과 동결법의 간편화를 시도했다.
정자운동율 50%이하의 정액은 Continuous-step density gradient centrifugation method를 이용해 세정 농축한 결과, 정자수와 운동율이 향상되어 정자소생율의 지표가 되는 Fertility Index{FI=정자수(/ml)X운동율(%)X10-8}가 5.6배 향상되었다. 또한, Oligozoospermia중에서도 정자수가 10X106/ml이하인 정액은 Cushion method를 이용해 농축한 결과, 정자수는 4.2 ± 2.1배, 운동율은 2.3 ± 1.9배 증가되었다.
세정과 농축시킨 정자의 간이동결법에 의한 냉동보존에 있어서, 간이동결기내에 액체질소(-196℃)를 넣고 수분간 방치시켜, 액체질소증기에 의해 동결기 내부가 온도평형(-150℃)에 도달되도록 했다. KS-II 보존액을 혼합한 0.5ml의 세정 농축정자를 냉동보존용 serum tube에 넣고, 액체질소 수면위 2cm에서 옆으로 뉘여 동결시킨 후, 액체질소저장용기(Liquid Nitrogen container)에 옮겨 보관했다. 인공 수정시에 30℃의 온수에 융해시킨 결과, 각각 70 ± 12%, 64 ± 11%의 정자소생율을 얻을 수가 있었다. 융해후에도(Continuous-step density gradient centrifugation method의 경우는 원래정자의 평균 2배 이상의 FI가 유지되었다.
이상의 결과에서 운동성 불량정액과 Oligozoospermia의 AIH에 있어서 정자의 세정 및 농축과, KS-II 냉동보존액을 혼합시킨 후 간이동결법을 이용한 냉동보존법으로 원래정액 이상의 정자수와 운동율을 얻을 수 있음이 확인되었다.

다국어 초록 (Multilingual Abstract)

It is well known that the cryopreservation of human semen plays an important role in the field of clinical Artificial Insemination by Husband semen (AIH) and Donor semen (AID). However, semen dilution with cryopreservation medium and decrease in sperm motility during freeze-thaw processes make lowering semen qualities and survival rate. The author undertook this experiments to pursue the higher survival rate of cryopreserved semen and more simple method of freezing technique.
To improve the qualities of cryopreserved semen after thawing, it is effective to concentrate the sperm from whole ejaculate by two method, continuous-step density gradient centrifugation method and Cushion method, as well as to achieve higher survival rate.
A : Prior to freezing, the continuous-step density gradient centrifugation method was undertaken for semen with poor motility (below 50 %) sperm. As a result of this, the semen was given 5.8 times improvement of Fertility Index (sperm density (106/ml) X motility (%) X 10-8)
B : The concentration of severe oligozoospermic semen by the Cushion method was improved the sperm density 4 times than the original semen.
A simple liquid nitrogen vapor freezing was employed in the present study : the concentrated sperm was mixed with KS-II cryopreservation medium and them placed at 2 cm up the surface of liquid nitrogen. After freezed by liquid nitrogen vapor at -150℃, the specimen was stored in liquid nitrogen container.
At the time of AIH, thawed with shaker in 30℃ water, the sperm survival rate of A and B were found to be 7-±12 % and 64 ± 11 %, respectively. The post thaw Fertility Index was as twice as that of the original semen by the continuous-step density gradient centrifugation method, and the survival rate was also high.
It is consequently suggested that simple liquid nitrogen vapor freezing method using KS-II cryopreservation medium, after sperm washing and concentration, is the very simple and convenient method with good results for cryopreservation of sperm.