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The presence of pelvic pain, a pelvic/paravaginal mass, and purulent vaginal discharge in primigravida should raise the possibility of obstructed hemivagina and uterine didelphys. Though conservative management could result in successful pregnancy outcomes, early excision of vaginal septum and adequate drainage offer a shorter course of management and complication avoidance.

Progressive loss of ovarian estrogen (E2) production is a hallmark feature of, if not a driving force behind, reproductive aging and the menopause. Recent genetic studies in mice have shown that female germline or oogonial stem cells (OSCs) contribute to maintenance of adult ovarian function and fertility under physiological conditions through support of de-novo oogenesis. Here we show that mouse OSCs express E2 receptor-α (ERα). In the presence of E2, ERα interacts with the stimulated by retinoic acid gene 8 (Stra8) promoter to drive Stra8 expression followed by oogenesis. Treatment of mice with E2 in vivo increases Stra8 expression and oogenesis, and these effects are nullified by ERα (Esr1), but not ERβ (Esr2), gene disruption. Although mice lacking ERα are born with a normal quota of oocytes, ERα-deficient females develop premature ovarian insufficiency in adulthood due to impaired oogenesis. Lastly, mice treated with reversible ER antagonists show a loss of Stra8 expression and oocyte numbers; however, both endpoints rebound to control levels after ceasing drug treatment. These findings establish a key physiological role for E2-ERα signaling in promoting OSC differentiation as a potential mechanism to maintain adequate numbers of ovarian follicles during reproductive life.

MUi028-A human induced pluripotent stem cell (hiPSC) line was generated from normal fetal skin fibroblasts using a non-integrative reprogramming method. Reprogramming factors OCT4, SOX2, KLF4, L-MYC, and LIN28, and TP53 shRNA in three episomal vectors were delivered by electroporation. The MUi028-A cell line had embryonic stem cell characteristics with consistent expression of specific pluripotency markers and the capability of in vitro differentiation into the three germ layers. This iPSC line can be used as a control to study disease mechanisms.

Aims: The aim is to study the relation and distribution in gene expression level of the luteinizing hormone receptor (LHR) gene and regulator of G-protein signaling 2 (RGS2) gene expression with oocyte maturation. Setting and Design: This cross-sectional study was undertaken in an instruction-based tertiary care infertility unit, department of obstetrics and gynecology. Materials and Methods: After controlled ovarian hyperstimulation, cumulus granulosa cells (CCs) from 59 oocytes among 18 women being treated by in vitro fertilization/intracytoplasmic sperm injection cycle technique from November 2015 to January 2016 were collected on the day of oocyte retrieval. Total RNA was extracted and converted to cDNA in individual oocytes. LHR and RGS2 gene levels were measured and analyzed using digital droplet polymerase chain reaction. Statistical Analysis: Gene expression level was analyzed using software STATA, version 14.0 (College Station, TX: StataCorp LP, USA). Results: CCs were obtained from 59 cumulus-oocyte complexes (COC), 46 COC from metaphase II (CCMII), 13 COC from metaphase I, and GV oocyte (CCMI + GV). The RGS2 gene expression level, when compared with the housekeeping gene in CCMII and CCMI + GV, was 0.15 (0.05–0.52) and 0.08 (0.02–0.27), respectively. The LHR gene expression when compared with the housekeeping gene in CCMII and CCMI + GV did not differ and was quite in the same value that was 0.02 (0.00–0.11) and 0.02 (0.00–0.06), respectively. Conclusion: This study showed that LHR gene expression did not differ in between oocyte groups. Even though the median of RGS2 gene expression was more in the mature oocyte group, the result was inconclusive due to scattering and overlapping of gene expression data between oocyte groups.

Objectives: To study the predictive value of post-processing total motile sperm count (TMSC) on pregnancy rate in intrauterine insemination (IUI). Materials and Methods: Medical records were reviewed from August 2013 to January 2015. All unexplained infertile women or anovulation less than or equal to 35 years old, and initially treated with IUI were included. Small myoma, adenomyosis, adenomyoma, endometrial thickness less than 6 mm at 2-3 days before IUI and patients with previous pelvic surgery were excluded. The predictive values of post-processing TMSC parameters were analyzed regarding pregnancy outcomes with a receiver operating characteristic (ROC) curve. results: The overall pregnancy rate was 9.5% for all 218 couples with 314 IUI cycles. The cause of female infertility was unexplained for 75.16% of the sample population with anovulation at 24.84%. All parameters including duration of infertility, number of dominant follicles, largest dominant follicle diameter and endometrial thickness were unrelated to pregnancy outcomes. The ROC showed sensitivity, specificity, positive predictive value and negative predictive value of post-processing TMSC at 20x106/mm, with pregnancy outcomes of 50, 45, 48, and 47%, respectively. Conclusion: Post-processing TMSC was an unreliable predictive factor for IUI outcome.

Thalassemia and hemoglobinopathy is a group of hereditary blood disorder with diverse clinical manifestation inherited by autosomal recessive manner. The Beta thalassemia/Hemoglobin E disease (HbE/βthal) causes a variable degree of hemolysis and the most severe form of HbE/βthal disease develop a lifelong transfusion-dependent anemia. Preimplantation genetic testing (PGT) is an established procedure of embryo genetic analysis to avoid the risk of passing on this particular condition from the carrier parents to their offspring. Preimplantation genetic testing for chromosomal aneuploidy (PGT-A) also facilitates the selection of embryos without chromosomal aberration resulting in the successful embryo implantation rate. Herein, we study the clinical outcome of using combined PGT-M and PGT-A in couples at risk of passing on HbE/βthal disease. The study was performed from January 2016 to December 2017. PGT-M was developed using short tandem repeat linkage analysis around the beta globin gene cluster and direct mutation testing using primer extension-based mini-sequencing. Thereafter, we recruited 15 couples at risk of passing on HbE/βthal disease who underwent a combined total of 22 IVF cycles. PGT was performed in 106 embryos with a 3.89% allele drop-out rate. Using combined PGT-M and PGT-A methods, 80% of women obtained satisfactory genetic testing results and were able to undergo embryo transfer within the first two cycles. The successful implantation rate was 64.29%. PGT accuracy was evaluated by prenatal and postnatal genetic confirmation and 100% had a genetic status consistent with PGT results. The overall clinical outcome of successful live birth for couples at risk of producing offspring with HbE/βthal was 53.33%. Conclusively, combined PGT-M and PGT-A is a useful technology to prevent HbE/βthal disease in the offspring of recessive carriers.

We aim to present a case of a healthy infant born after intracytoplasmic sperm injection-in vitro fertilization (ICSI-IVF) with a preimplantation genetic diagnosis (PGD) for pantothenate kinase-associated neurodegeneration (PKAN) due to PANK2 mutation.ICSI-IVF was performed on a Thai couple, 34-year-old female and 33-year-old male, with a family history of PKAN in their first child. Following fertilization, each of the embryos were biopsied in the cleavage stage and subsequently processed for whole-genome amplification. Genetic status of the embryos was diagnosed by linkage analysis and direct mutation testing using primer extension-based mini-sequencing. Comprehensive chromosomal aneuploidy screening was performed using a next-generation sequencing-based strategy.Only a single cycle of ICSI-IVF was processed. There were seven embryos from this couple-two were likely affected, three were likely carriers, one was likely unaffected, and one failed in target genome amplification. Aneuploidy screening was performed before making a decision on embryo transfer, and only one unaffected embryo passed the screening. That embryo was transferred in a frozen thawed cycle, and the pregnancy was successful. The diagnosis was confirmed by amniocentesis, which presented with a result consistent with PGD. At 38 weeks of gestational age, a healthy male baby was born. Postnatal genetic confirmation was also consistent with PGD and the prenatal results. At the age of 24 months, the baby presented with normal growth and development lacking any neurological symptoms.We report the first successful trial of PGD for PKAN in a developing country using linkage analysis and mini-sequencing in cleavage stage embryos.

BackgroundA key event in human development is the establishment of erythropoietic progenitors in the bone marrow, which is accompanied by a fetal-to-adult switch in hemoglobin expression. Understanding of this event could lead to medical application, notably treatment of sickle cell disease andβ-thalassemia. The changes in gene expression of erythropoietic progenitor cells as they migrate from the fetal liver and colonize the bone marrow are still rather poorly understood, as primary fetal liver (FL) tissues are difficult to obtain.MethodsWe obtained human FL tissue and adult peripheral blood (AB) samples from Thai subjects. Primary CD34+cells were culturedin vitroin a fetal bovine serum-based culture medium. After 8 days of culture, erythroid cell populations were isolated by flow cytometry. Gene expression in the FL- and AB-derived cells was studied by Affymetrix microarray and reverse-transcription quantitative PCR. The microarray data were combined with that from a previous study of human FL and AB erythroid development, and meta-analysis was performed on the combined dataset.ResultsFL erythroid cells showed enhanced proliferation and elevated fetal hemoglobin relative to AB cells. A total of 1,391 fetal up-regulated and 329 adult up-regulated genes were identified from microarray data generated in this study. Five hundred ninety-nine fetal up-regulated and 284 adult up-regulated genes with reproducible patterns between this and a previous study were identified by meta-analysis of the combined dataset, which constitute a core set of genes differentially expressed between FL and AB erythroid cells. In addition to these core genes, 826 and 48 novel genes were identified only from data generated in this study to be FL up- and AB up-regulated, respectively. Thein vivorelevance for some of these novel genes was demonstrated by pathway analysis, which showed novel genes functioning in pathways known to be important in proliferation and erythropoiesis, including the mitogen-activated protein kinase (MAPK) and the phosphatidyl inositol 3 kinase (PI3K)-Akt pathways.DiscussionThe genes with upregulated expression in FL cells, which include many novel genes identified from data generated in this study, suggest that cellular proliferation pathways are more active in the fetal stage. Erythroid progenitor cells may thus undergo a reprogramming during ontogenesis in which proliferation is modulated by changes in expression of key regulators, primarily MYC, and others including insulin-like growth factor 2 mRNA-binding protein 3 (IGF2BP3), neuropilin and tolloid-like 2 (NETO2), branched chain amino acid transaminase 1 (BCAT1), tenascin XB (TNXB) and proto-oncogene, AP-1 transcription factor subunit (JUND). This reprogramming may thus be necessary for acquisition of the adult identity and switching of hemoglobin expression.

Human iPSC line MU011.A-hiPS was generated from homozygous α-thalassemia (−SEA/−SEA) fetal skin fibroblasts using a non-integrative reprogramming method. Reprogramming factors OCT3/4, SOX2, KLF4, L-MYC, LIN28, and shRNA of TP53 contained in three episomal vectors were delivered using electroporation.

Multiple labs have reported that mammalian ovaries contain oogonial stem cells (OSCs), which can differentiate into oocytes that fertilize to produce offspring. However, the physiological relevance of these observations to adult ovarian function is unknown. Here we performed targeted and reversible ablation of premeiotic germ cells undergoing differentiation into oocytes in transgenic mice expressing the suicide gene, herpes simplex virus thymidine kinase (HSVtk), driven by the promoter of stimulated by retinoic acid gene 8 (Stra8), a germ cell-specific gene activated during meiotic commitment. Over a 21-day ablation phase induced by the HSVtk pro-drug, ganciclovir (GCV), oocyte numbers declined due to a disruption of new oocyte input. However, germ cell differentiation resumed after ceasing the ablation protocol, enabling complete regeneration of the oocyte pool. We next employed inducible lineage tracing to fate map, through Cre recombinase-mediated fluorescent reporter gene activation only in Stra8-expressing cells, newly-formed oocytes. Induction of the system during adulthood yielded a mosaic pool of unmarked (pre-existing) and marked (newly-formed) oocytes. Marked oocytes matured and fertilized to produce offspring, which grew normally to adulthood and transmitted the reporter to second-generation offspring. These findings establish that oocytes generated during adulthood contribute directly to ovarian function and natural fertility in mammals.

Context: In controlled ovarian hyperstimulation cycles, 15% of oocytes have been proven to be immature. Key factors include failure in signal transmission from the cumulus cell to the oocyte, insufficient level of luteinizing hormone, and internal conditions of the oocyte itself. Aims: The aim of the present study was to investigate the effect of human chorionic gonadotropin (hCG) on the in vitro maturity of partially cumulus-denuded immature oocytes collected after controlled ovarian stimulation for in vitro fertilization (IVF). Settings and Design: This was a prospective, randomized controlled design at the department of obstetrics and gynaecology, university hospital. Subjects and Methods: Infertile women underwent gonadotropin-releasing hormone antagonist stimulated protocol for IVF with final maturation triggered by hCG, partially cumulus-denuded immature human oocytes were allocated to two groups: the first was treated with fertilization medium and the second was treated with fertilization medium and hCG. They were cultured for 24 h. Outcomes measured were the oocyte maturation rates to metaphase II (MII) and glucose-6-phosphate dehydrogenase (G6PD) activity of in vitro maturation (IVM) mature oocytes which represent the oocyte quality. Statistical Analysis Used: The Mann–Whitney U-test and One-way ANOVA were used to compare continuous variables, and Chi-square was used for categorical data. Results: In all, 250 immature stimulated oocytes were allocated (125 per group). The maturation rate was higher in the hCG supplement group (48% vs. 39.2%) without significance. The positive brilliant cresyl blue results among the MII oocytes developed from the metaphase I (MI) were significantly higher in the hCG group (P = 0.001). Conclusions: Rescue IVM in fertilization culture medium plus hCG was slightly better than that in the only fertilization culture. MII oocytes developed from MI in hCG supplemented medium had a higher quality based on the measured G6PD activity.

Objective:To study the effectiveness of a pre-operative single dose sublingual misoprostol for reducing blood loss of abdominal hysterectomy with/without salpingo-oophorectomy performed for symptomatic uterine leiomyomas.Materials and methods: A total of 70 women undergone total abdominal hysterectomy with/without salpingo-oophorectomy for symptomatic uterine leiomyoma between June 2012 to January 2013 was double-blindly randomized into two groups to receive two tablets of misoprostol (200 microgram each) sublingual (n= 35) or two placebo tablets vitamin B6 (100 milligram each) sublingual (n=35) at 30-60 minutes before the operation. The amount of intra-operative blood loss, the change in haemoglobin level after the operation, the requirement for blood transfusion and the incidence of side effects were recorded.Result:Our data showed no significant difference between two groups in term of intraoperative blood loss. (misoprostol, mean 549.6 ± 391.9 ml versus placebo, mean 445.0 ± 270.6 ml, p=0.20) Moreover there were no observed differences in the need for postoperative blood transfusion, the change in hemoglobin level after the operation and side effects profiles between the two groups.Conclusion: We concluded that a single preoperative dose of sublingual misoprostol is not effective in reducing intraoperative blood loss and need for post-operative blood transfusion after total abdominal hysterectomy with/without salpingo-oophorectomy for symptomatic uterine leiomyomas.

Aim The aim of this study was to examine the effects of estradiol valerate (EV) on the thickness of clomiphene citrate (CC)-stimulated endometrium. Material and Methods Thirty-four normal ovulatory women were randomized double-blindly into two groups to receive CC 100 mg/day on day 2–6 of the treatment cycle, and either vitamin B (placebo) or EV 6 mg/day on day 10–14 of the cycle. The endometrial thickness, endometrial pattern, numbers of mature follicles, and maximal diameters of preovulatory follicles were evaluated by transvaginal sonographic examination. Results Thirty women completed both treatment cycles. Two other participants dropped out during the treatment due to side-effects (headache). The average endometrial thickness of the group treated with CC + placebo became slightly thinner when compared to the thickness at the baseline (9.04 vs 9.52 mm; P = 0.24). The CC + placebo and the CC + EV resulted in similar endometrial pattern, ovulation day, numbers of mature follicles, and sizes of the leading follicles before ovulation. However, an addition of EV into the CC cycle significantly increased the average endometrial thickness (10.7 mm vs 9.04 mm; P

Summary Solid surface vitrificaition (SSV) is a cryoperservative method that has been used in the cryopreservation of oocytes, and embryos. Here, we report an application of the SSV in the cryopreservation of human spermatozoa. We compared the SSV with a standard freezing method in terms of sperm motility, morphology, vitality and DNA integrity. Sperm motility was determined by computer assisted semen analysis, morphology and vitality were determined by eosin-methylene blue staining, and DNA integrity was determined by a TUNEL assay. We found that while both cryopreservative methods produced spermatozoa with comparable vitality and motility, the SSV gave slightly, but significantly fewer sperm with DNA damage, and loose tail. We concluded that, a cryopreservation of human spermatozoa by SSV is feasible and provides a quick and practical way to preserve human spermatozoa with a comparable, if not better, quality of the preserved spermatozoa to the standard freezing method.

Through the use of parabiosis in mice, aging-related deterioration of skeletal muscle and liver has been linked to a loss of systemic factors that support adult stem or progenitor cell activity. Since aging-related ovarian failure has recently been attributed, at least in part, to a loss of de-novo oocyte-containing follicle formation associated with declining oogonial stem cell activity, herein we tested in mice if aging-related changes in systemic factors influence the size of the ovarian follicle reserve. Ovaries of young (2-month-old) females parabiotically joined with young females for 5 weeks possess comparable numbers of healthy and degenerative (atretic) oocyte-containing follicles in their ovaries as those detected in non-parabiotic young females. Joining of young females with young males significantly increases follicle atresia without a net change healthy follicle numbers. Surprisingly, young females joined with aged (24-month-old) males exhibit a significant increase in the number of primordial follicles comprising the ovarian reserve, and this occurs without changes in follicle growth activation or atresia. Blood of aged males also induces ovarian expression of the germ cell-specific meiosis gene, Stimulated by retinoic acid gene 8 (Stra8), in ovaries of female parabionts, further supporting the conclusion that the observed changes in the follicle reserve of females joined with aged males reflect increased oocyte formation. Thus, factors in male blood exert dramatic effects on ovarian follicle dynamics, and aging males possess a beneficial systemic factor that significantly expands the ovarian follicle reserve in females through enhanced oogenesis.

The aim of this study was to examine the effects of estradiol valerate (EV) on the thickness of clomiphene citrate (CC)-stimulated endometrium.Thirty-four normal ovulatory women were randomized double-blindly into two groups to receive CC 100 mg/day on day 2-6 of the treatment cycle, and either vitamin B (placebo) or EV 6 mg/day on day 10-14 of the cycle. The endometrial thickness, endometrial pattern, numbers of mature follicles, and maximal diameters of preovulatory follicles were evaluated by transvaginal sonographic examination.Thirty women completed both treatment cycles. Two other participants dropped out during the treatment due to side-effects (headache). The average endometrial thickness of the group treated with CC + placebo became slightly thinner when compared to the thickness at the baseline (9.04 vs 9.52 mm; P = 0.24). The CC + placebo and the CC + EV resulted in similar endometrial pattern, ovulation day, numbers of mature follicles, and sizes of the leading follicles before ovulation. However, an addition of EV into the CC cycle significantly increased the average endometrial thickness (10.7 mm vs 9.04 mm; P0.001).We concluded that the addition of 6 mg/day EV following the CC treatment can prevent the endometrial thinning without perturbing folliculogenesis and ovulation.