D.H. Choi

FREEZING IMMATURE OOCYTES

The establishment of a long-term preservation system for mammalian oocytes is important for the development of both biological and medical sciences. A number of efforts have been made to develop this system. In human reproductive medicine, the development of an oocyte cryopreservation system can improve the efficacy of the current assisted reproductive technology (ART) for infertile patients with severe reproductive disorders. In this article, the technical development of cryopreservation programs for human oocytes and its biological background were reviewed. Clinical outcome after the use of this technology was further introduced.

A molecular basis for embryo apposition at the luminal epithelium

To obtain a gene expression profile during embryo apposition to the luminal epithelium, we isolated mouse luminal epithelium from implantation (IM) and interimplantation (INTER) sites using laser capture microdissection (LCM), and analyzed their gene expression by microarray analysis. IM and INTER sites were sampled on day 4.5 after mating of female mice with fertile males (day 0.5 = vaginal plug). RNA was extracted, amplified, labeled, and hybridized to microarrays and results were analyzed using the significance analysis of microarrays (SAM) method. Comparison of IM and INTER sites by SAM identified 73 genes most highly ranked at IM, while 13 genes most highly expressed at the INTER sites, within the estimated false discovery rate (FDR) of 0.163. Among 73 genes at IM, 20 were ESTs or were of unknown function, and the remain 53 genes had known functions mainly relating to cellular structuring and others such as cell cycling, gene/protein expression, immune responses, invasion, metabolism, oxidative stress, or signal transduction. Specifically, of the 24 structural genes, 14 were implicated in extracellular matrix and tissue remodeling. Meanwhile, of the 13 genes that were highly expressed at INTER, eight were ESTs or of unknown function, and the remaining five were implicated in metabolism, signal transduction, and gene/protein expression. Among these 58 (53 + 5) genes with known functions, 13 genes (22.4%) were associated with Ca2+ for their function. Results of the present study suggest that (1) at IM sites, active tissue remodeling is occurring for embryo invasion while the INTER sites are relatively quiescent and (2) Ca2+ may be a vital regulatory factor in the apposition process. Investigations of human homologues of those genes expressed in the mouse luminal epithelium during apposition may help to understand the implantation process and/or implantation failure in humans.

The apolipoprotein A-I level is downregulated in the granulosa cells of patients with polycystic ovary syndrome and affects steroidogenesis.

Polycystic ovary syndrome (PCOS) is the most common endocrine disorder found in women. The etiology of PCOS is still not clear, and there are no available studies on the proteome analysis of granulosa cells (GCs) in PCOS patients. To identify the pathogenic mechanisms and potential diagnostic markers for PCOS, we conducted proteomic profiling of GCs in PCOS patients by two-dimensional gel electrophoresis and liquid chromatography coupled with mass spectrometry (LC-MS/MS) analyses. The proteomic analysis yielded eight downregulated and 12 upregulated proteins in PCOS patients, among which apolipoprotein A-I (ApoA-I) showed significant downregulation in PCOS patients as confirmed by Western blotting. Knockdown of ApoA-I decreased the number of transcripts of steroidogenic enzymes in a granulosa cell line (KGN), while its overexpression generally increased the level of expression of these enzymes. Furthermore, modulation of the expression level of ApoA-I in the granulosa cells altered progesterone production. Therefore, this study suggests that ApoA-I can be useful as a granulosa cell biomarker of PCOS patients and that downregulated ApoA-I may be related to the disturbed production of steroid hormones in PCOS patients.

mRNA expression pattern of insulin-like growth factor components of granulosa cells and cumulus cells in women with and without polycystic ovary syndrome according to oocyte maturity

The mRNA expression of insulin-like growth factor (IGF) components was investigated in granulosa cells (GCs) and cumulus cells (CCs) sampled from immature and mature follicles in women with and without polycystic ovary syndrome (PCOS). Gene expression for IGF-binding protein (IGFBP) 3 in GCs definitely differed between normal women and women with PCOS, whereas increase in IGF-I and IGF receptor (IGFR) 2 mRNA in mature-follicle GCs, increase in IGF-II, IGFR-1, and IGFBP-4 mRNA in immature-follicle GCs, increase in IGFR-2 and IGFBP-2 mRNA in mature-follicle CCs, and increase IGFBP-6 mRNA in immature-follicle CCs were observed in both normal women and women with PCOS.

Ruptured pyosalpinx following a fetal reduction procedure

Pyosalpinx is an accumulation of pus in a fallopian tube. The conditions under which a pyosalpinx can occur vary greatly. Although pyosalpinx has been reported during controlled ovarian hyperstimulation for in vitro fertilization and embryo transfer (IVF-ET) [1], it has also been reported in a 13-year-old virgin [2] and after uterine arterial embolization [3]. A 30-year-old woman with a history of a right salpingectomy performed because of a tubal pregnancy before 20 weeks of gestation visited the CHA Fertility Center in March 2008 for IVF-ET. Left tuboplasty was performed by laparoscopy after 1 cycle of IVF-ET and thawed ET had failed. Three embryos were transferred after an additional IVF cycle and the patient became pregnant. Transvaginal ultrasound-guided selective abortion of monochorionic diamniotic (MCDA) twins was performed under analgesia with fentanyl at 8 weeks of gestation because of a quadruple pregnancy. The patient presented with generalized weakness after the procedure. The following day, the patient presented with chills and myalgia. Her body temperature was 37.6 °C and lower abdominal rebound tenderness was present. Leukocytosis was evident by a leukocyte count of 12620 cells/μL (segmentation 95.0%) and the patients C-reactive protein concentration was 5.96 mg/dL. Blood, cervical, and urine cultures were obtained and intravenous antibiotics (cephalosporin andmetronidazole) were administered. The following day, the patients diffuse abdominal pain was aggravated. A follow-up ultrasound detected 2 fetuses with heart beats and an enlarged left ovary (5.7×3.7 cm) that showed intact blood flow from the ovarian vessels. Fluid in the pouch of Douglas was also noted. Brown pus and debris were found on explorative laparoscopy of the pelvic cavity. On close inspection, the left salpinx was ruptured and yellowish pus was found to be leaking from the lesion. After aspiration of the pus for peritoneal fluid cytology, the left salpinx was excised with suction and irrigation and a drain was inserted into the pelvic cavity. On the first postoperative day the patients body temperature and leukocyte count had decreased, and her abdominal pain had subsided. Culture results for the pus, blood, urine, and cervix showed no microbial growth. Many neutrophils were present in the peritoneal fluid cytology specimen. By the sixth postoperative day, the patient had made an uneventful recovery and left the hospital with the 2 remaining fetuses healthy and viable. An immunocompromised status due to poor general health can cause a latent pathogen in a hydrosalpinx to precipitate a pyosalpinx. The cause is uncertain in the present case. Clinicians should consider the possibility of pyosalpinx in the differential diagnosis of patients presenting with symptoms of abdominal pain and fever following fetal reduction.