난소관련병변에서 복강체액내의 cytokine 농도

전상식(Sang Sik Chun),조영래(Young Lae Cho),구태본(Tae Bon Koo),서장수(Jang Soo Suh) 대한산부인과학회 2001 Obstetrics & Gynecology Science Vol.44 No.2

Objective : Recent studies have shown that multifunctional cytokines have been implicated in the pathogenesis and pathophysiology of various ovarian lesions including ovarian cancer and endometriosis. This study was performed to determine the possibility of the clinical usefulness of peritoneal cytokines in patients with various ovarian lesions. Methods : Peritoneal fluid was obtained from patients with benign cystic adenoma(n=11), benign cystic teratoma(n=10), endometriosis(n=23), malignant ovarian tumor(n=11), and women without evidence of any pathology(n=7) at the time of laparotomy or operative laparoscopic surgery and were examined for the levels of IFN-γ, IL-1β, IL-5, IL-6, IL-10 and TNF-α using enzyme-linked immunosorbent assay(ELISA). Statistical analysis was performed with the one way ANOVA, Scheffe test or Mann-Whitney test. Results : The mean concentration of IL-6 and IL-10 was significantly higher in peritoneal fluid specimens with ovarian cancer than other benign pathology(p<0.05). Both IL-6 and IL-10 levels in peritoneal fluid specimens with endometriosis tended to be higher than normal, and the levels of IL-6 and IL-10 were significantly higher in peritoneal fluid of women with severe endometriosis compaired to women with mild endometriosis (p<0.05). Conclusion : This study supports the concept that regional immunologic dysfunction might be involved in the disease process of various ovarian pathology such as ovarian cancer and endometriosis. A larger study would help in evaluating the potential use of local peritoneal cytokines in differentiating ovarian cancer from other benign pathology, and demonstrating any association between concentrations of cytokines and severity of endometriosis.

불임환자에서 sonohysterography의 이용

이춘희(Chun Hee Lee),김광수(Gwang Soo Kim),이동영(Dong Young Lee),박철민(Chul Min Park),오영은(Young Eun Oh),이택후(Taek Hoo Lee),전상식(Sang Sik Chun) 대한산부인과학회 1999 Obstetrics & Gynecology Science Vol.42 No.9

Objective: The purpose of this study was to assess the usefulness of sonohysterography in the detection of abnormalities of the uterine cavity in infertile patients, compared with other diagnostic methods, such as transvaginal sonography, hysterosalpingography and hysteroscopy. Methods: Three intrauterine pathologies including intrauterine synechiae, endometrial polyp and submucosal myoma, that could be possible causes of infertility were diagnosed. Results: Transvaginal ultrasonography and hysterosalpingography were able to detect 41.7% and 83.3% of uterine pathologies respectively comparing with sonohysterography to detect all of the uterine pathologies. Conclusion: Our results show that sonohysterography is easy, inexpensive, well-tolerated and non-invasive technique with high sensitivity in the detection of uterine pathologies. Therefore the use of sonohysterography for the diagnosis of intrauterine pathologies in infertile patients is highly recommended.

마우스에서 배반포 형성, 세포 수 및 ICM의 비율에 미치는 배양액의 효과

박기상,이택후,전상식,송해범,Park, Kee-Sang,Lee, Taek-Hoo,Chun, Sang-Sik,Song, Hai-Bum 대한생식의학회 2002 Clinical and Experimental Reproductive Medicine Vol.29 No.4

Objective : The aim of this study was to evaluate the influence of different media on blastulation, mean cell number, percentage of inner cell mass (ICM) of total cells and ICM : trophectoderm (TE) ratio in mice. Materials and methods: A total 552 two cell embryos were retrieved from ICR female mice (4 weeks old) at 48 hr after hCG injection (mated just after hCG injection) and cultured in MEM (n=276) or TCM (n=276) supplemented with 20% hFF. The grading of blastocysts from zona-intact (ZiB) to -escape (hatching and hatched, ZeB) was performed at 72 hours after culture. Total, TE and ICM cell numbers were analyzed by differential staining of blastocyst. Statistical analysis was performed using t-test with SigmaPlot-2001. P-values < 0.05 were accepted as statistically significant. Results: The blastulation rate in MEM ($64.9{\pm}4.95%$) was significantly higher (p=0.0031) than that in TCM ($57.2{\pm}5.22%$). No differences were found in the number of ZiB and ZeB between MEM ($31.9{\pm}2.62$, $33.0{\pm}4.58%$), and TCM ($27.2{\pm}4.28$, $30.1{\pm}4.58%$). A total 314 blastocysts (MEM=166; TCM=148) were stained differentially. Mean cell number of blastocysts was significantly higher (p=0.0002) in TCM ($73.1{\pm}3.3$) than in MEM ($61.7{\pm}2.5$). Differential staining was successfully performed in 155 blastocysts (MEM=77; TCM=78). The percentage of ICM was significantly higher in MEM than in TCM ($20.9{\pm}1.3$ vs. $17.1{\pm}1.2%$, p=0.0281). The ICM : TE ratio was higher in TCM than in MEM (1 : $4.85{\pm}0.68$ vs. 1 : $3.78{\pm}0.78$, NS). Conclusion: These results show that MEM increase the blastocyst formation and percentage of ICM of total cells comparing with TCM in mice.

자궁내유착의 자궁경 치료와 그 결과

주응식(Eung Sik Ju),김일규(Il Gyu Kim),오영은(Young Eun Oh),이현정(Hyun Jung Lee),이택후(Taek Hu Lee),전상식(Sang Sik Chun) 대한산부인과학회 2001 Obstetrics & Gynecology Science Vol.44 No.2

One hundred sixty three patients were hysteroscoped for various reasons at our department over 10 year period. Ninety eight patients out of 163 who were diagnosed and treated for their intrauterine adhesions(IUAs) by hysteroscopy were analysed retrospectively. Patients were classified as having mild, moderate or severe stage of IUAs according to American Fertility Society classification. Twenty nine patients(29.6%) had mild IUAs, 43 patients (43.9%) had moderate IUAs, and 26 patients(26.5%) were classified as having severe stage IUAs. The predisposing causal factors of IUAs were abortion, full-term delivery, H. mole, pelvic tuberculosis and myomectomy in order of frequency but in eight cases no cause was identified. After hysteroscopic surgery, although high rate(83.9%) of restoration of normal menstruation was attained but term pregnancy rate was only 53.3% and rather high rate (32%) of preterm or abortion were identified and those pregnancy outcomes were correlated with the stage of IUAs. Complications of surgery consisted of uterine or bladder perforation, pulmonary edema and hemorrhage.

The Clinical Outcomes after Embryo Transfer (ET) on Day 2 and Day 5 or Subsequent ET on Day 2-5, 2-6, 2-7, 3-5 and 4-7 in In Vitro Fertilization-ET Cycles

김대원,박기상,송해범,이택후,전상식,Kim, Dae-Won,Park, Kee-Sang,Song, Hai-Bum,Lee, Taek-Hoo,Chun, Sang-Sik The Korean Society for Reproductive Medicine 2003 Clinical and Experimental Reproductive Medicine Vol.30 No.1

연구목적: 일반적으로 IVF-ET에서 가장 높은 임신율을 얻는 방법은 5 day ET (배반포기 배아 이식)이지만 장기간 배양이 적절하지 못한 경우에는 $2{\sim}4$일째에 ET를 실시하고 나서 $5{\sim}7$일째에 배반포기에 도달한 배아를 재이식 (SET)하여, SET의 효용성에 대하여 조사하고자 실시하였다. 연구재료 및 방법: 48주기의 환자에서 회수한 난자와 수정란은 10%와 20% hFF가 첨가한 DMEM에서 이식 직전까지 각각 공배양하였다. 채란 2일 (group I, day 2 ET), 5일째 이식 (group II, day 5 ET) 또는 재이식 (group III, SET; 2-5, 2-6, 2-7, 3-5, 4-7일)을 실시하면서 수정률, 할구분할률 및 임신율을 각각 비교하였다. 결과에 대한 통계 분석을 SAS (version 6.2)를 이용한 Duncan's Multiple Range Test를 이용하여 p값이 0.05 보다 작을 때 통계적으로 유의차가 있는 것으로 하였다. 결 과: 수정률은 group II (90.5%)가 다른 군에 비하여 높게 (p<0.05) 나타났다 (group I: 80.6%; group III: 82.9%). 할구분할률은 군간에 차이가 없었다 (수정란 당 $93.3{\sim}99.1%$). 임상적 임신율은 group II와 III (각각 58.3%)가 group I (33.3%) 보다 높게 나타났다. 그러나 처리군이 적어서 통계적인 차이는 없었다. 결 론: 배반포기 배아를 단독 이식하는 것이 임신율을 높일 수 있는 최선의 방법으로 나타났지만, 채란수가 적거나 수정률이 저조한 경우에는 $2{\sim}4$일째에 ET를 실시한 후 여분의 배아를 배반포기까지 배양한 다음 $5{\sim}7$일에 재이식 (SET)하면 blastocyst ET에서 나타날 수 있는 이식 자체의 실패를 방지할 수 있으면서 임신율을 높일 수 있는 이식 기법이 될 것이다.

Glutamine 함유 배양액에 첨가한 에너지원이 마우스의 배 발달에 미치는 영향

김주환,박기상,이택후,전상식,송해범,Kim, Ju-Hwan,Park, Kee-Sang,Lee, Taek-Hoo,Chun, Sang-Sik,Song, Hai-Bum 대한생식의학회 2000 Clinical and Experimental Reproductive Medicine Vol.27 No.1

Objective: Mammalian embryos undergo changes of energy environment for transfer from oviduct to uterus. Also, the human reproductive organ (oviduct, uterus) contains energy sources of different concentration (oviduct - glucose: 0.5 mM, pyruvate: 0.32 mM, lactate: 10.5 mM; uterus - goucose: 3.15 mM, pyruvate: 0.1mM, lactate: 5.87 mM, respectively). This study was conducted to examine the effect of these energy sources added in DMEM with glutamine on the mouse embryo development. Methods: There was used ICR female mouse. Two cell embryos of mouse are collected by method of 'flushing'. Flushing fluid was used Ham's F-10 added to 20% FBS. The collected 2 cell embryos were cultured in media such as Control (only DMEM), group A and B (DMEM supplemented with 0.5 mM and 3.15 mM glucose), and group C and D (DMEM supplemented with 0.1 mM and 0.32 mM pyruvate), and group E and F (DMEM supplemented with 5.87 mM and 10.5 mM lactate). All experimental media supplemented with 20% hFF, respectively. Pattern of embryo development was observed to interval at 24hr during 96hr. Results : The media with glutamine added glucose (group A: 51.0%; group B: 48.4%) was significantly (p<0.05) higher than other experimental group in development into the morula stage after 24 hr in culture, but not significantly different compared with control and the rate of development into the blastocyst was significantly (p<0.05) low in the both of pyruvate (group C: 7.9% group D: 6.8%) and lactate (group E: 7.1%, group F: 7.1%) treatment group after 48 hr in culture. Development into the blastocyst and hatched balstocyst after 72 hr in culture revealed similarly in control (81.9%) and glucose treatment group (group A: 83.3%, group B: 82.8%). However, development into the hatched and attached blastocyst after 96hr in culture revealed significantly (p<0.05) development in the glucose treatment group (group A: 82.3%, group B: 78.5%) than control (63.2%), and its of pyruvate (group C: 34.1%, group D: 34.1%) and lactate (group E: 25.9%, group F: 33.3%) treatment group were significantly (p<0.05) lower than control similar to previous observations. Conclusion : The glucose added to the DMEM with only glutamine, as energy source, was highly to the rate of development compared with control, but the other energy sources were not, synthetically. Above refer to, the human reproductive organ (oviduct, uterus) contains energy sources of different concentration. Thus, further studies are will examine continuously to effects by interaction of different energy sources in the mouse embryo development, and these results will provide to foundation on the human embryo culture.

인간양수에 의한 생쥐 난자 투명대의 정자수용능력 억제의 관찰

박기상,이택후,송해범,전상식,Park, Kee-Sang,Lee, Taek-Hoo,Song, Hai-Bum,Chun, Sang-Sik 대한생식의학회 2000 Clinical and Experimental Reproductive Medicine Vol.27 No.1

Objective: Zona pellucida (ZP) has been thought to be the barrier of egg to sperm penetration before and after fertilization. The phenomenon of ZP hardening has been considered as a post-fertilization event until now, and it is generally accepted that it is caused by the secretory products of cortical granules released during the cortical reaction. Hardening of ZP could occur "spontaneously" in mammalian oocytes in standard culture conditions, and that it is probably not a consequence of cortical reaction. The purpose of our study was to investigate the effect of human amniotic fluid (HAF) on nuclear maturation (NM) and fertilization ability of mouse immature oocytes. Methods: HAF was obtained from patients undergoing amniocentesis at $16{\sim}20$ weeks of gestation. HAF from five to ten patients was centrifuged and the supernatants was pooled. Cumulusenclosed mouse immature oocytes were incubated in the medium containing HAF, and examined to confirm NM and fertilization. Female ICR mice (about 3 weeks old) were stimulated with 7.5 IU PMSG. Immature oocytes were isolated at $48{\sim}52$ hrs post PMSG injection and cultured in TCM-199 supplemented with 20% HAF for 18 hrs. FBS was used as a control for the examination. Matured oocytes (MII) were fertilized with sperms collected from the epididymis of male mice (over 10 weeks old). Fertilization was in conducted T6 medium containing 15 mg/ml BSA, and confirmed at 6 hrs post-insemination. Fertilization rate was assessed in zona-intact or zona-free oocytes (denuded by trypsin). Evaluation of NM and fertilization was carried out by rapid staining method. ZP hardening was evaluated by incubating cumulus cell-free mature oocytes in 0.001% chymotrypsin at $37^{\circ}C$ for 10 min. Results: There was no significant difference between the effects of HAF (86.6%) and FBS (87.7%) supplements on NM of immature oocytes. When maturation medium was supplemented with HAF, total fertilization rates (7%) were significantly lower (p<0.01) than that of FBS (85.1%). In HAF group, fertilization rate was increased (p<0.01) in zona-free oocytes (7% versus 100%). The resistance of mouse oocyte ZP to digestion by chymotrypsin after maturation in vitro was significantly higher (p<0.01) in HAF group (86.7%) than in FBS (6.7%). To culture oocytes in FBS were very effective in preventing ZP hardening. However cultured oocytes in HAF showed high rate of ZP hardening (p<0.01). Conclusions: These results suggest that HAF can be used as a supplement for the NM of mouse immature oocytes in vitro. However, HAF induces spontaneous hardening of ZP of mouse immaure oocytes during maturation in vitro.

난자내 정자 직접주입술에서 난자의 처리방법이 난자의 발생능력에 미치는 영향

박기상,이택후,송해범,전상식,Park, Kee-Sang,Lee, Taek-Hoo,Song, Hai-Bum,Chun, Sang-Sik 대한생식의학회 1999 Clinical and Experimental Reproductive Medicine Vol.26 No.3

Objective: In the preparation of ICSI, cumulus and corona cells should be removed from the oocytes by using a combination of enzymatic (hyaluronidase) and mechanical (pipetting) methods. But little is known about the effects of different degrees of oocyte denudation and incubation time between denudation and sperm injection on the outcomes of ICSI. The aim of this study was to evaluate the effects of varying the degrees of oocyte denudation and the lengths of incubation time from denudation to sperm injection on the outcomes of ICSI. Methods: In experiment 1, patients (oocytes) were grouped into group A and B according to the degree of denudation, complete and partial, respectively. In experiment 2, patients (oocytes) were grouped into group I, II and III according to the length of incubation time of denuded oocytes until sperm injection as < 1, $1{\sim}2$ and >2 hours, respectively. Results: There was no significant difference between the degree of oocyte denudation on the survival, fertilization and development rates after ICSI procedure. In case of the incubation time of denuded oocytes until ICSI, survival rates was higher in group III (83.1 %) than in group I (61.5%, p<0.05) or group II (64.3%). However no statistically significant differences were found between incubation time and fertilization or development rates. Conclusions: This study reveals that the outcomes of ICSI are not affected by the degree (complete or partial) of oocyte denudation. However, the denuded oocytes with incubation period of more than 2 hours show better outcomes of ICSI than those with the incubation period of less than 2 hours.

생쥐 난소에서 Preantral Follice의 단순 분리법

김주환,박기상,송해범,전상식,Kim, Ju-Hwan,Park, Kee-Sang,Song, Hai-Bum,Chun, Sang-Sik 대한생식의학회 2000 Clinical and Experimental Reproductive Medicine Vol.27 No.3

Objective: Our present studies were conducted to examine more effective isolating method of preantral follicles from mouse ovaries. Methods: ICR mice (3-6 weeks old) were sacrificed through cervical dislocation and their ovaries were removed and put into watch glasses containing Hams F-10 supplemented with 10% fetal bovine serum (FBS). Preantral follicles were isolated by three different methods; 1) enzymatical method and 2) mincing method, and 3) scraping method. Enzymatical method was carried out as following. Ovaries were bisected with a pair of fine 30G needles. Bisected ovaries were incubated at $37^{\circ}C$ and 5% $CO_2$ incubator in 2-well dish containing Hams F-10 supplemented with collagenase 600 lU/ml and DNAse 20 lU/ml. After 20 min., follicles were isolated by repeated pipetting. Isolated preantral follicles were collected, and the remnant of tissues was placed in incubator and previous procedure was repeated. Mincing method was carried out with a pair of fine 30G needles attached to 1 ml syringes and minced ovary. Scraping method was carried out with a pair of fine 30G needles and scratched to surface of ovary. The differences between isolating methods were analyzed using Student's t-test and Chi-square. Results were considered statistically significant when ${\rho}$ value was less than 0.05. Results: In handling time, mincing or scraping method ($28{\pm}3.42$ min or $16{\pm}1.58$ min) were significantly (p<0.00001) shorter than enzymatical method ($72{\pm}1.69$ min), and scraping method was significantly (p<0.01) shorter than mincing method. Total number of isolated follicles was significantly (p<0.0001) higher in enzymatical method ($49.8{\pm}3.91$) than in mincing or scraping method ($25.3{\pm}2.33$ or $20.5{\pm}1.75$). Isolated follicles in ${\leq}$90${\mu}m$ were significantly (p<0.005) higher in enzymatical method ($15{\pm}1.71$) than in mincing or scraping method ($7.8{\pm}0.98$ or $8.1{\pm}1.31$). In 91~130 ${\mu}m$, isolated follicles were significantly (p<0.0005) higher in enzymatical method ($33{\pm}3.27$) than in mincing or scraping method ($16.3{\pm}1.82$ or $10.7{\pm}1.38$). In ${\geq}$ 131 ${\mu}m$, isolated follicles were not significantly differences between all groups. In equal sizes, the rate of isolated follicles in ${\leq}$ 90 ${\mu}m$ was highest in scraping method (39.6% vs. enzymatical method: 30.1%, p<0.05; mincing method: 30.9%, p=0.11719, NS). Rate of follicles in $91{\sim}130$ ${\mu}m$ was significantly (p<0.05) lower in scraping method (52.7%) than in enzymatical or mincing method (66.3% or 64.5%). Rate of follicles in ${\geq}$131 ${\mu}m$ was highest in scraping method (8.3% vs. enzymatical or scraping method: 3.6%, p<0.05 or 4.6%, p=0.19053, NS). Conclusions: This study suggests that scraping method is simple and useful for isolation of preantral follicles, because this method reduced handling time and recovered enough follicles. The recovered rate of isolated follicles in diameter of 91 ~ 130 ${\mu}m$ was highest in all methods.

마우스 배반포 배의 Differential staining에서 Propidium Iodide와 Bisbenzimide의 노출이 미치는 영향

박기상,박성백,이택후,전상식,송해범,Park, Kee-Sang,Park, Sung-Baek,Lee, Taek-Hoo,Chun, Sang-Sik,Song, Hai-Bum 대한생식의학회 2002 Clinical and Experimental Reproductive Medicine Vol.29 No.4

Objective: These experiments were conducted to investigate the optimal expose length of propidium iodide (PI) and bisbenzimide on differential staining of mouse blastocysts. Materials and methods: A total 964 blastocysts (early${\sim}$hatched) was exposed to PI (n=831) (group I: $\leq$ 10; II: $11{\sim}15$; III: $16{\sim}20$; IV: $\geq$21 sec) and bisbenzimide (n=133) (group A: $\leq$1; B: $1{\sim}$3; C: $\geq$ 4 hr) in several periods for differential staining. Statistical analysis was performed using t-test with SigmaPlot-2001. P-values < 0.05 were accepted as statistically significant. Results: In case of PI exposure, differential staining rates were significantly higher (p<0.05) in group I (89.8%) than in any others (group II: 77.6%; III: 29.6%; IV: 22.2%) and higher (p<0.05) in group II than in group III and IV. In case of bisbenzimide exposure, differential staining rates were not statistically differences in three groups (group A: 97.4%; B: 87.8%; C: 93.3%). Conclusion: The differential staining rates of mouse blastocysts are not affected by the exposure length of bisbenzimide. However, blastocysts were exposed to PI with period of shorter than 15 sec show best outcomes of differential staining rates.