The purpose of this study is compare IVF cycle outcome in poor responders between clomiphene citrate (CC) stimulated and controlled ovarian hyperstimulation (COH) protocol. A total of 94 patients responding poorly in previous IVF cycles (estradiol<600 pg/ml or less than 3 oocytes retrieved) subsequently underwent either COH (COH group: 122 cycles, 68 patients) or CC-stimulated cycles (CC group: 43 cycles, 26 patients). CC was administered for five consecutive days starting on cycle day 3 at a dose of 100 mg daily. Serial transvaginal ultrasound examination was done from cycle day 8. Urine was collected 3~4 times before hCG injection for the detection of LH surge. The hCG was administered when serum estradiol reached greater than 150 Pg/ml and mean follicle diameter>16 mm. In COH group, ovarian stimulation was done using short protocol (GnRH-a/FSH/HMG/hCG). No difference in age or number of transferred embryos was found between CC group and COH group. COH group had significantly (p<0.05) higher mean peak level of E2 (810+/-112 vs 412+/-55 pg/ml) and greater number of retrieved oocytes (3.0+/-0.2 vs 2.0+/-0.2) than CC group. CC group had significantly embryos (1.8+/-0.2) compared with (2.1+/-0.2) in COH group. However, CC group had higher pregnancy rate than COH group per retrieval [26.9% (7/26) vs 6.2% (6/97)], or per transfer [31.8% (7/22) vs 7% (6/86)]. Although cycle cancellation rate in CC group (48.8%) was higher than that of COH group (21.3%), the pregnancy rate per cycle in CC group was still higher (16.3%) than COH group (4.9%). In addition, implantation rate in CC group was 17.5% (7/40), which was significantly p<0.01) higher than 3.9% (7/180) in COH group. These data suggest that oocyte and embryo quality are lower in COH cycles of poor responders than CC cycles. We suggest that clomiphene citrate stimulated IVF cycle may be more efficient than COH IVF cycle in poor responders in terms of lower costs and higher pregnancy performance.
Clomiphene* ; Embryonic Structures ; Estradiol ; Fertilization* ; Humans ; Oocytes ; Ovulation Induction ; Pregnancy ; Pregnancy Rate ; Ultrasonography

OBJECTIVE: Human embryonic stem (ES) cells have a great potential in regenerative medicine and tissue engineering. The human ES cells could be differentiated into specific cell types by treatments of growth factors and alterations of gene expressions. However, the efficacy of guided differentiation and isolation of specific cells are still low. In this study, we characterized isolated cells from differentiated human ES cells by magnetic activated cell sorting (MACS) system using specific antibodies to cell surface markers. METHODS: The undifferentiated hES cells (Miz-hESC4) were sub-cultured by mechanical isolation of colonies and embryoid bodies were spontaneously differentiated with DMEM containing 10% FBS for 2 weeks. The differentiated cells were isolated to positive and negative cells with MACS system using CD34, human epithelial antigen (HEA) and human fibroblast (HFB) antibodies, respectively. Observation of morphological changes and analysis of marker genes expression were performed during further culture of MACS isolated cells for 4 weeks. RESULTS: Morphology of the CD34 positive cells was firstly round, and then it was changed to small polygonal shape after further culture. The HEA positive cells showed large polygonal, and the HFB positive spindle shape. In RT-PCR analysis of marker genes, the CD34 and HFB positive cells expressed endodermal and mesodermal genes, and HEA positive cells expressed ectodermal genes such as NESTIN and NF68KD. The marker genes expression pattern of CD34 positive cells changed during the extension of culture time. CONCLUSION: Our results showed the possibility of successful isolation of specific cells by MACS system from undirected differentiated human ES cells. Thus, MACS system and marker antibodies for specific cell types might be useful for guided differentiation and isolation of specific cells from human ES cells.
Antibodies ; Ectoderm ; Embryoid Bodies ; Endoderm ; Fibroblasts ; Gene Expression ; Humans* ; Intercellular Signaling Peptides and Proteins ; Mesoderm ; Nestin ; Regenerative Medicine ; Tissue Engineering

No abstract available.

The aim of this study is to determine whether or not hydrosalpinx affects pregnancy rate and implantation rate adversely in human IVF-ET program. 110 infertile couples with tubal factor undertaken IVF-ET treatment from May 1995 through April 1996 were included. Patients with tubal factor combined with other factors were excluded from this study. The hydrosalpinx group included 35 patients who had unilateral or bilateral hydrosapinx. Fifty four patients with proximal tubal occlusion without hydrosalpinx served as a control. The corrected hydrosalpinx group included 21 patients hydrosalpinx served as a control The corrected hydrosalpinx group included 21 patients who had undertaken either salpingectomy(n=5) or salpingoneostomy(n=16) before IVF-ET cycle. Controlled ovarian hyperstimulation was performed using GnRH agonist/human menopausal gonadotropin or follicular stimulation hormone(FSG). Thirty four hours after intramuscular injection of 10,000 IU human chorionic gonadotropin(hCG), trasvaginal sonography-guided oocyte retrieval was done, The cleaved embryos were transferred to the uterus on day two or three after fertilization. There was no significant difference in age(mean +/- SEM; 32.9 +/- 0.4, 32.7 +/- 0.6, 31.4 +/- 0.6), basal FSH level(7.1 +/- 0.3, 7.2 +/- 0 0.3, 7.0 +/- 0.4 mIU/ml) and estradiol on the day of hCG injection(2674 +/- 219, 3239 +/- 304, 3376 +/- 360 pg/ml) among the control, hydrosalpinix and corrected hydrosalpinx group, respectively(p > 0.05). The number of trasferred embryos(5.1 +/- 0.3, 5.6 +/- 0.3 and 5.4 +/- 0.6) were similar between the groups. The clinical pregnancy rate of 8.3% in hydrosalpingeal group was significantly lower than 25.4% of control group (p=0.057). However, in corrected hydrosalpinx group, pregnancy rate of 27.3% compared well with the control group. The implantation rate showed a similar pattern with pregnancy rate, i.e., hydrosalpingeal group had the lowest implantation rate of 2.0%, which was significantly lower than 11.6% of control group. In the corrected hydrosalpinx group, implantation rate(9.3%) recovered as that of the control group. The ectopic pregnancy rate(11.1%) of the hydrosalpingeal group was higher than that of the control group(1.7%) and was intermediate(4.5%) in corrected hydrosalpinx group(p > 0.05). In conclusion, these data show that hydrosalpinx affects the pregnancy rate adversely in IVF-ET cycyle. Thus, it is suggested that surgical correction of the hydrosalpinx before the initiation of IVF-ET cycle may be beneficial in increasing the pregnancy rate as well as decreasing the ectopic pregnancy.
Chorion ; Embryo Transfer* ; Embryonic Structures* ; Estradiol ; Family Characteristics ; Female ; Fertilization ; Gonadotropin-Releasing Hormone ; Gonadotropins ; Humans ; Injections, Intramuscular ; Oocyte Retrieval ; Pregnancy Rate* ; Pregnancy* ; Pregnancy, Ectopic ; Sterilization, Tubal ; Uterus

OBJECTIVE: To evaluate the effects of recombinant FSH (rFSH) and urinary FSH (uFSH) on the gene expressions of human endometrial stromal cells in vitro. METHODS: Endometrial tissue was obtained from a pre-menopausal women undergoing hysterectomy. Primary endometrial stromal cells were isolated and in vitro cultured with FBS-free DMEM/F-12 containing 0, 10, 100, and 1,000 mIU/ml of rFSH and uFSH for 48 hours, respectively. Total RNA was extracted from the cultured cells and subjected to real time RT-PCR for the quantitative analysis of progesterone receptor (PR), estrogen receptor alpha/beta (ER-alpha/beta), cyclooxygenase 2 (Cox-2), leukemia inhibitory factor (LIF), homeobox A10-1 and -2 (HoxA10-1/-2). RESULTS: Both hormone treatments slightly increased (< 3 folds) the expressions of PR, ER-beta and HoxA10-1/-2 gene. However, ER-alpha expression was increased up to five folds by treatments of both FSH for 48 hours. The LIF expression by the 10 mIU/ml of uFSH for 12 hours was significantly higher than that of rFSH (p<0.01). After 24 hours treatment of two kinds of hormones, the expression patterns of LIF were similar. The 100 and 1,000 mIU/ml of rFSH induced significantly higher amount of Cox-2 expression than those of uFSH, respectively (p<0.05). CONCLUSION: This study represents no adversely effect of exogeneous gonadotropins, rFSH and uFSH, on the expression of implantation related genes. We suggest that rFSH is applicable for the assisted reproductive technology without any concern on the endometrial receptivity.
Cells, Cultured ; Cyclooxygenase 2 ; Estrogens ; Female ; Gene Expression* ; Genes, Homeobox ; Gonadotropins ; Humans* ; Hysterectomy ; Leukemia Inhibitory Factor ; Receptors, Progesterone ; Reproductive Techniques, Assisted ; RNA ; Stromal Cells*

No abstract available.
Epidermolysis Bullosa ; Muscular Dystrophy, Duchenne ; Ornithine Carbamoyltransferase ; Polymerase Chain Reaction* ; Preimplantation Diagnosis*

OBJECTIVE: To investigate whether polymorphisms of gene encoding CYP1B1 is associated with the risk of endometriosis in Korean women. METHODS: We investigated 199 patients with histopathologically confirmed endometriosis rAFS stage III/IV and 183 control group women who were surgically proven to have no endometriosis. The genetic distribution of four different CYP1B1 polymorphisms at G119-T, G432-C, T449-C, and A453-G were analyzed by polymerase chain reaction (PCR) and restriction fragment length polymorphism of PCR products. RESULTS: We found no overall association between each individual CYP1B1 genotype and the risk of endometriosis. The odds ratio of genotype GG/GC+GG/TC+TT/AA compared to GG/CC/CC/AA (reference) was calculated as 2.06 with a 95% confidence interval of 1.003~4.216. CONCLUSIONS: This results suggest that CYP1B1 genetic polymorphism may be associated with development of endometriosis in Korean women.
Endometriosis* ; Female ; Genotype ; Humans ; Odds Ratio ; Polymerase Chain Reaction ; Polymorphism, Genetic ; Polymorphism, Restriction Fragment Length

OBJECTIVE: To investigate the association between endometriosis and polymorphisms of N-acetyl transferase 2 (NAT2), glutathione S-transferase M1 (GSTM1), and cytochrome P450 (CYP) 1A1 genes in Korean infertile patients. MATERIALS AND METHODS: A total of 303 infertile patients who had undertaken diagnostic laparoscopy during January, 2001 through December, 2003 at Samsung Cheil Hospital enrolled in this study. The patients were grouped according to laparoscopic findings: minimal to mild endometriosis (group I: n=147), moderate to severe endometriosis (group II: n=57), normal pelvic cavity (n=99). Peripheral blood was obtained and genomic DNA was extracted. The genotypes of each genes were analyzed using polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP). For NAT2, RFLP was used to detect the wild type (wt) and mutant (mt) alleles, enabling classification into slow (mt/mt) or fast (wt/wt or wt/mt) acetylation genotypes. For GSTM1, PCR was used to distinguish active (+/- or +/+) from null (-/-) genotypes. For CYP1A1, MspI digestion was used to detect the wild type (A1A1), heterozygote (A1A2) or mutant (A2A2) genotypes. RESULTS: The genotype frequencies of NAT2 slow acetylator was 12.8%, 10.9%, 12.8% in group I, group II and control, respectively. The genotype frequencies of GSTM1 null mutation was 55.3%, 41.8%, 53.2% in group I, group II and control, respectively. The genotype frequencies of CYP1A1 MspI polymorphism was 16.3%, 9.1%, 18.1% in group I, group II and control, respectively. No significant difference was observed between endometriosis and normal controls in the genotype frequencies of the NAT2, GSTM1, CYP1A1 MspI polymorphism. CONCLUSION: The NAT2, GSTM1, CYP1A1 gene polymorphism may not be associated with the susceptibility of endometriosis in Korean women.
Acetylation ; Alleles ; Classification ; Cytochrome P-450 CYP1A1 ; Cytochrome P-450 Enzyme System* ; Cytochromes* ; Digestion ; DNA ; Endometriosis* ; Female ; Genotype ; Glutathione Transferase* ; Glutathione* ; Heterozygote ; Humans ; Laparoscopy ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length ; Transferases*

OBJECTIVE: To evaluate the viability and the characteristics of shed endometrial tissues obtained from menstrual fluid during in-vitro culture. METHODS: The menstrual fluids were collected using Wallace catheter from uterine cavity in 10 women with regular menstruation. The menstrual fluids were washed twice, and the pellets, containing blood cells and shed endometrium, were collected and diluted fivefold with Ham's F-10 medium containing 10% fetal bovine serum. The cell suspension was placed on culture dishes, and cultured for 7 days in an incubator. To evaluate the characteristics of the cultured endometrial cells, immunohistochemical (IHC) staining was performed using anti-cytokeratin and anti-vimentin antibody. RESULTS: The mean volume of menstrual fluids and pellets were 0.7ml and 0.3ml, respectively. Only 15% of the shed endometrial tissues were attached and proliferated in culture dishes, which was considered to have viability. Initially, endometrial epithelial cells and fibroblasts were attached and proliferated, and the area of these cells was increased according to prolong the culture time. Stromal cell colonys were located and proliferated on the epithelial cells. IHC staining showed strongly positive for cytokeratin in epithelial cells and for vimentin in stromal cells. In the confocal microscopic observation of 3-dimensional structure of cultured endometrium, cytokeratin-positive cells (epithelial cells) were located in the pheriphery and cytokeratin-negative cells (stromal cells) inside of the structure. CONCLUSION: From our study, shed endometrial tissues in menstrual fluid showed meaningful viability and closed relationship between epithelial cells and stromal cells during in-vitro culture. Thus, we suggest that the in-vitro culture system of shed endometrium is a suitable model for researches of endometriosis.
Blood Cells ; Catheters ; Endometriosis ; Endometrium ; Epithelial Cells ; Female ; Fibroblasts ; Humans ; Incubators ; Keratins ; Menstruation ; Stromal Cells ; Vimentin

OBJECTIVE: The aim of this study was to evaluate the efficacy of frozen-thawed ET in poor prognosis patients such as the old age (38~44 years; OA group) and the patients who did not achieve clinical pregnancy with the first fresh ET cycle (non-pregnant patients; NP group). METHODS: Laboratory and clinical data were collected from fresh and frozen-thawed ET cycles of OA and NP group. Controlled ovarian hyperstimulation (COH) and conventional insemination or ICSI, in vitro culture and ET were performed by routine procedures. Supernumerary embryos were frozen by the slow freezing method, and frozen embryos were thawed by the rapid thawing method. Embryo development, pregnancy and implantation rates were statistically analyzed by Student t-test and chi square test. RESULTS: Mean ages were similar between fresh ET (40.0+/-1.8 years, n=206) and frozen-thawed ET (39.9+/-1.9 years, n=69) cycles in OA group. However, the clinical pregnancy and implantation rate of subsequent frozen-thawed ET significantly higher than those of fresh ET cycles (29.0% and 11.2% vs. 16.5% and 7.0%, p<0.05). In NP group, there was no difference in the mean age between fresh ET (31.2+/-2.3 years, n=40) and frozen-thawed ET (31.9+/-3.1 years, n=119) in subsequent cycles. The clinical pregnancy and implantation rates were similar between the subsequent fresh ET (42.5% and 22.6%) and the frozen-thawed ET (40.3% and 18.8%). CONCLUSION: In old age patients, higher pregnancy rate of frozen-thawed ET compared to fresh ET cycles in this study. It may be related that better uterine environments for implantation in frozen-thawed ET cycles than that of non-physiological hormonal condition in uterus of fresh COH cycles.
Embryonic Development ; Embryonic Structures ; Female ; Freezing ; Humans ; Insemination ; Pregnancy ; Pregnancy Rate ; Prognosis ; Sperm Injections, Intracytoplasmic ; Uterus