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Transgene-free human HS5-SV.hiPS line was generated from human cesarean scar-derived fibroblasts using temperature-sensitive Sendai virus vectors carrying Oct4, Sox2, cMyc and Klf4 exogenous transcriptional factors. The viral constructs were eliminated from HS5-SV.hiPS line through heat treatment. Transgene-free HS5-SV.hiPS cells expressed pluripotent associated transcription factors Oct4, Nanog, Sox2, Rex1 and surface markers SSEA-4, TRA-1-60 and OCT4. HS5-SV.hiPS cells formed embryoid bodies and differentiated into three embryonic germ layers in vivo. HS5-SV.hiPS cells maintained their normal karyotype (46, XX) after culture for extended period. HS5-SV.hiPS displayed the similar pattern of DNA fingerprinting to the parenteral scar-derived fibroblasts. [ABSTRACT FROM AUTHOR]

Background: Mesenchymal stem cells (MSCs) can be derived from several tissues such as bone marrow, placenta, adipose tissue, or endometrial tissue. MSCs gain a lot of attention for cell-based therapy due to their characteristics including differentiation ability and immunomodulatory effect. Preclinical and clinical studies demonstrated that MSCs can be applied to treat female infertility by improving of the functions of ovary and uterus. This mini- review focuses on the current study of treatment of endometrial infertility by using MSCs., Methods: The present study performed a literature review focusing on the effect of MSCs for treatment of women infertility caused by endometrial dysfunction., Results: Bone marrow-, umbilical cord-, adipose-, amniotic-, and menstruation-derived MSCs enhance endometrial cell proliferation, injury repairs as well as reducing scar formation. The beneficial mechanism probably via immunomodulatory, cell differentiation, stimulates endometrial cell proliferation and down-regulation of fibrosis genes. The major advantage of using MSCs is to improve endometrial functions resulting in increased implantation and pregnancy., Conclusions: MSCs exhibit a potential for endometrial infertility treatment. Adipose- and menstruation-derived stem cells show advantages over other sources because the cells can be derived easily and do not causes graft rejection after autologous transplantation., Competing Interests: There are no conflicts of interest to declare., (© 2020 The Authors. Reproductive Medicine and Biology published by John Wiley & Sons Australia, Ltd on behalf of Japan Society for Reproductive Medicine.)

In a feeder-dependent culture system of human pluripotent stem cells (hPSCs), coculture with mouse embryonic fibroblasts may limit the clinical use of hPSCs. The aim of this study was to determine the feasibility of using human Caesarean scar fibroblasts (HSFs) as feeder cells for the culture of hPSCs. HSFs were isolated and characterised and cocultured with hPSCs, and the pluripotency, differentiation ability and karyotypic stability of hPSCs were determined. Inactivated HSFs expressed genes (including inhibin subunit beta A (INHBA), bone morphogenetic protein 4 (BMP4), fibroblast growth factor 2 (FGF2), transforming growth factor-β1 (TGFB1), collagen alpha-1(I) (COL1A1) and fibronectin-1 (FN1) that have been implicated in the maintenance of hPSC pluripotency. When HSFs were used as feeder cells, the pluripotency and karyotypic stability of hPSC lines did not change after prolonged coculture. Interestingly, exogenous FGF2 could be omitted from the culture medium when HSFs were used as feeder cells for hESCs but not hiPSCs. hESCs cocultured with HSF feeder cells in medium without FGF2 supplementation maintained their pluripotency (as confirmed by the expression of pluripotency markers and genes), differentiated invitro into embryonic germ layers and maintained their normal karyotype. The present study demonstrates that HSFs are a novel feeder cell type for culturing hPSCs and that supplementation of exogenous FGF2 is not necessary for the Chula2.hES line.

Because the diploid human embryonic stem cells (hESCs) can be successfully derived from tripronuclear zygotes thus, they can serve as an alternative source of derivation of normal karyotype hESC lines. The aim of the present study was to compare the pluripotency and trophoblast differentiation ability of hESCs derived from tripronuclear zygotes and diploid hESCs. In the present study, a total of 20 tripronuclear zygotes were cultured; 8 zygotes developed to the blastocyst stage and 1 hESC line was generated. Unlike the previous studies, chromosomal correction of tripronuclear zygotes during derivation of hESCs did not occur. The established line carries 3 sets of chromosomes and showed a numerical aberration. Although the cell line displayed an abnormal chromosome number, it was found the cell line has been shown to be pluripotent with the ability to differentiate into 3 embryonic germ layers both in vitro and in vivo. The expression of X inactive specific transcript (XIST) in mid-passage (passage 42) of undifferentiated triploid hESCs was detected, indicating X chromosome inactivation of the cell line. Moreover, when this cell line was induced to differentiate toward the trophoblast lineage, morphological and functional trophoblast cells were observed, similar to the diploid hESC line.

Although human pluripotent stem cells (hPSCs) can proliferate robustly on the feeder-free culture system, genetic instability of hPSCs has been reported in such environment. Alternatively, feeder cells enable hPSCs to maintain their pluripotency. The feeder cells are usually grown in a culture medium containing fetal bovine serum (FBS) prior to coculture with hPSCs. The use of FBS might limit the clinical application of hPSCs. Recently, human cord blood-derived serum (hUCS) showed a positive effect on culture of mesenchymal stem cells. It is interesting to test whether hUCS can be used for culture of feeder cells of hPSCs. This study was aimed to replace FBS with hUCS for culturing the human foreskin fibroblasts (HFFs) prior to feeder cell preparation. The results showed that HFFs cultured in hUCS-containing medium (HFF-hUCS) displayed fibroblastic features, high proliferation rates, short population doubling times, and normal karyotypes after prolonged culture. Inactivated HFF-hUCS expressed important genes, including Activin A, FGF2, and TGFβ1, which have been implicated in the maintenance of hPSC pluripotency. Moreover, hPSC lines maintained pluripotency, differentiation capacities, and karyotypic stability after being cocultured for extended period with inactivated HFF-hUCS. Therefore, the results demonstrated the benefit of hUCS for hPSCs culture system.

Human neural progenitor cells (hNPCs) are the starting material required for neuronal subtype differentiation. Proliferation of hNPCs allows researchers to study the mechanistic complexities and microenvironments present during neural differentiation and to explore potential applications for hNPCs in cell therapies. The use of enzymatic dissociation during hNPC proliferation causes dissociation-induced apoptosis; therefore, in the present study, we examined the effect of the p-160-Rho-associated coiled-coil kinase (ROCK) inhibitor Y-26732 on dissociation-induced apoptosis of hNPCs. We generated hNPCs via embryoid body formation using serum-free culture medium supplemented with noggin. The established hNPCs were characterized and the effect of the ROCK inhibitor on hNPC dissociation was studied. We demonstrated that supplementation of the culture media with 10 μM Y-26732 efficiently reduced apoptosis of dissociated hNPCs; this supplementation was effective when the inhibitor was applied either at (i) 24 h before dissociation of the cells and at 24 h after plating the cells or (ii) at 24 h after plating of the cells only. In addition to reducing apoptosis, both supplementation conditions with Y-26732 enhanced the proliferation of dissociated hNPCs. Our findings provide the optimal time window for ROCK treatment of hNPC dissociation in respect to apoptosis and cell proliferation., (© 2013 S. Karger AG, Basel.)

Human embryonic stem (hES) cells are considered to be a potential source for the therapy of human diseases, drug screening, and the study of developmental biology. In the present study, we successfully derived hES cell lines from blastocysts developed from frozen and fresh embryos. Seventeen- to eighteen-year-old frozen embryos were thawed, cultured to the blastocyst stage, and induced to form hES cells using human foreskin fibroblasts. The Chula2.hES cell line and the Chula4.hES and Chula5.hES cell lines were derived from blastocysts developed from frozen and fresh embryos, respectively. The cell lines expressed pluripotent markers, including alkaline phosphatase (AP), Oct3/4, stage-specific embryonic antigen (SSEA)-4, and tumor recognition antigen (TRA)-1-60 and TRA-1-81 as detected with immunocytochemistry. The real-time polymerase chain reaction (RT-PCR) results showed that the cell lines expressed pluripotent genes, including OCT3/4, SOX2, NANOG, UTF, LIN28, REX1, NODAL, and E-Cadherin. In addition, the telomerase activities of the cell lines were higher than in the fibroblast cells. Moreover, the cell lines differentiated into all three germ layers both in vitro and in vivo. The cell lines had distinct identities, as revealed with DNA fingerprinting, and maintained their normal karyotype after a long-term culture. This study is the first to report the successful derivation of hES cell lines in Thailand and that frozen embryos maintained their pluripotency similar to fresh embryos, as shown by the success of hES cell derivation, even after years of cryopreservation. Therefore, embryos from prolonged cryopreservation could be an alternative source for embryonic stem cell research.

Pluripotent stem cells would have great potential in cell therapies and drug development when genetically matched with the patient; thus, histocompatible cells could be used in transplantation therapy or as a source of patient-specific cells for drug testing. Pluripotent embryonic stem cells (ESCs)-generated via somatic cell nuclear transfer (SCNT) or parthenogenesis (pESC)-are potential sources of histocompatible cells and tissues for transplantation. Earlier studies used the piezoelectric microinjection (PEM) technique for nuclear transfer (NT) in mouse. No specific studies examined zona-free (ZF) NT as an alternative NT method to generate genetically matched ESCs of a nuclear donor. In this study, we compared the efficiency of nuclear transfer-derived ESC (ntESC) line establishment from ZF-NT, ZF-parthenogenetic (PGA), and ZF-fertilized embryos with that of the PEM-NT method. Different nuclei donor cells [cumulus, ESC, and mouse embryonic fibroblast (MEF)] were used and the efficiency of ntESC derivation was investigated, along with their in vitro characterization. The ZF-NT method's efficiency was higher than that of the PEM-NT using cumulus cells. When ESCs and cumulus cells were used as nuclear donor cells, they resulted in significantly higher ZF-NT-derived ntESC line establishment rates compared to MEF cells. In conclusion, the nuclear donor cell type significantly affected the efficiency of ntESC line establishment, and the ZF-NT method was efficient to establish pluripotent ntESC lines.

Objective: To establish human embryonic stem (hES) cells from human embryos., Design: Experimental study., Setting: Reproductive Medicine Unit, Department of Obstetrics and Gynecology, Faculty of Medicine, Chulalongkorn University., Material and Method: Abnormal and normal fertilization embryos were cultured in vitro until reaching blastocyst stage. Four different methods for isolation of ICMs were used. Immunosurgery, mechanical isolation, laser assists, and whole blastocyst culture were performed. The feeder layers used in the present study were fibroblasts, isolated from either mouse or human. Mechanical splitting of ICM outgrowths or hES-like cells was performed for propagation of cells. Characterization of hES-like cells was conducted by morphology, detection of immunostaining of Oct-4, and enzymatic activity of alkaline phosphatase (AP). HES-like cells were spontaneously differentiated through suspension culture of embryoid body (EB). Subsequent differentiation was done on gelatin-coated dishes., Main Outcome Measure: Establishment of hES cells., Results: By using abnormal fertilization embryos, 80.0% (8/10) of blastocysts were able to attach on the feeder layers, 50% (4/8) formed ICM outgrowths, but no hES-like cells were established. By using normal fertilization embryos, 84.6% (22/26) of blastocysts were able to attach on feeder layers, 18.2% (4/22) formed ICM outgrowths. One hES-like cell line was successfully established by using mechanical isolation of ICMs and human adult skin fibroblasts as feeder layers. This hES-like cells exhibited typical morphology of hES cells, positive staining for Oct-4 and AP. hES-like cells were able to form EB and differentiated into neural-like cells., Conclusion: This is the first report in Thailand that hES-like cells can be isolated from normal development human embryos at blastocysts-stage using mechanical isolation of ICM and culture with human adult skin fibroblast as feeder layers.

Objective: To determine the effect of leukemia inhibitory factor (LIF) on the quality of in vitro produced mouse blastocyst and the efficiency of embryonic stem (ES) cell derivation., Design: Experimental study, Setting: Reproductive Medicine Unit, Department of Obstetrics and Gynecology, Faculty of Medicine, Chulalongkorn University, Material and Method: In vivo fertilized zygotes were collected and subjected to in vitro culture in potassium simplex optimized medium (KSOM) containing 1,000 unit/ml LIF. The developmental ability of the zygote to blastocyst-stage and the cell numbers in blastocysts were evaluated Expanded blastocysts developed in different culture media were subsequently subjected to ES cell derivation. MAIN OUTCOME MEASURE (s): The influence of LIF on the quality of and the total cell numbers of blastocyst developed in vitro., Results: Supplementation of LIF in KSOM increased the rate of hatching blastocysts (63.8% vs. 53.7%; p < 0.05) and total cell numbers (91.4 +/- 15.0 vs. 85.1 +/- 7.7; p < 0.05) compared to KSOM alone. ES cells were obtained 66.7% from blastocysts developed in KSOM-LIF versus 41.7% in KSOM (p > 0.05). Established ES cell lines showed typical colony and characteristics of pluripotent murine ES cells., Conclusion: LIF improved the quality of in vitro produced blastocysts but not enhanced ES cell derivation.

Stem cell research has obtained more attention during the last decade because of its strong potential as a new tool to cure many chronic diseases. In addition, stem cell knowledge is an important basis for understanding pathophysiology at the cellular level and developing disease models for experimental research. There are different limitations on resources, budget, policy and regulation among countries. As a result, each country has particular advantages and disadvantages in stem cell research. This result in the establishment of international networks and collaborations to coordinate and promote stem cell research aimed at medical applications.

Embryonic stem cell is the promising novel therapeutic tool for various degenerative diseases and tissue injuries. With the concept of cell and tissue therapy, many chronic disorders will be curable. The present article provides basic knowledge of stem cell in areas of definition, classification and future clinical applications. In addition, stem cell application is not only focusing on regenerative purpose, but also concentrating on more understanding about the early human development and the pathophysiology of genetic diseases at the cellular level. However, there are some technical problems and ethical concern that should be resolved before applying stem cells into clinical practice.

A prospective randomized, double blind, single centre study was conducted to compare the efficacy, efficiency and clinical side effects of daily fixed dose regimen of either 100 IU or 200 IU of recombinant follicle stimulating hormone(rFSH) Follitropin beta in down-regulated women undergoing controlled ovarian hyperstimulation(COH) for either conventional in vitro fertilization(IVF) or intracytoplasmic sperm injection(ICSI). A total of sixty women were randomly allocated according to the criteria for the treatment by either 100 IU(n = 30) or 200 IU (n = 30) of FSH. Although more cycle cancellations due to low response were observed in the 100 IU group (n = 9 vs n = 2), two cases of mild and moderate ovarian hyperstimulation syndrome were noted in the higher dose group. Subjects in the group treated with 200 IU appeared to yield more follicles > 17 mm (4.4 vs 3.3, p = 0.05) and more oocytes compared to the group treated with 100 IU (9.2 versus 6.0 oocytes, NS). The total dosage required to develop at least three follicles according to the protocol was significantly lower in the group treated with 100 IU (1203.33 versus 2106. 67, P < 0.0001). In conclusion, a fixed daily dose of 200 IU of rFSH Follitropin beta compared to a fixed daily dose of 100 IU is more effective in terms of follicles > 17 mm development and the number of oocytes retrieved along with a lower cancellation rate, but less efficient as indicated by a higher total rFSH dose needed

To compare mouse blastocyst survival after cryopreservation with vitrification and the slow-freezing method, one-hundred and forty-eight in vitro mouse blastocysts were randomly frozen by the two methods: vitrification and conventional slow-freezing. After being thawed, the blastocysts were assessed for survival and hatching rate. The survival rates of blastocysts cryopreserved by vitrification and slow-freezing were 68.33 and 65.52 per cent (p = 0.89), whereas hatching rates were 51.22 and 44.74 per cent, respectively (p = 0.64). Therefore, vitrification of blastocyst-stage-embryos may be a useful, economic method for freezing the excess blastocysts in some centers where blastocysts are routinely transferred.

The determinant factors and the anxiety level of infertile couples during the treatment of in vitro fertilization and embryo transfer were studied in 60 infertile couples between 1 January to 31 May 2000. The instruments employed in the study were Personal and Health Data Questionnaire, the Cornell Medical Index, and the Determinant Factors of Anxiety. The average age was between 36-40 years old, holding a Bachelor's degree and working in private companies earning a monthly income between 10,000-20,000 Baht. Most infertile couples wanted to have a child in order to fulfill the meaning of being a "family" and were anxious about the treatment. The couples in general did not have any background of emotional disturbance. Women were found to have a slightly higher anxiety than men. The determinant factors of anxiety were found to be the side-effects of the infertility treatment, inadequate time to consult with the physician/nurse, the outcome of the infertility treatment, possibility to possibility of not succeeding/infertility cannot be treated and the process of the diagnostic procedures accordingly. The results of the study will serve as a guideline for improving better services and understanding between the physician and the patient regarding the expectations of the IVF treatment.

The objective of the study was to develop the somatic nuclear transfer technique by using rabbits as the model. The oocyte recipients aged 16 h post coitus were collected surgically from 20 superovulated rabbit doe with 28 and 40 mg Follicle Stimulating Hormone (FSH) after mating with a vasectomized male. The metaphase II plate and 1st polar body of oocyte was later aspirated by enucleated micropipette under an inverted microscope. A single donor cell; cumulus cell or cultured or frozen fibroblast cell from passage 1 to 9 were transferred to enucleated oocyte and fused with triple DC pulses, 3.2 kv, 20 micros. The fused embryos were cultivated in TCM 199 NaHCO3 + 10 per cent fetal calf serum (FCS) for 4 days. The cleavage rate (2-cell stage) was 37.2 per cent (32/86) from eight experiments, and 18.8 per cent (6/32) developed to the early morula stage. This study also indicated that the enucleation pipette and the somatic cell type influenced the success.

Alcohol and tobacco consumption are closely correlated and published results on their association with breast cancer have not always allowed adequately for confounding between these exposures. Over 80% of the relevant information worldwide on alcohol and tobacco consumption and breast cancer were collated, checked and analysed centrally. Analyses included 58,515 women with invasive breast cancer and 95,067 controls from 53 studies. Relative risks of breast cancer were estimated, after stratifying by study, age, parity and, where appropriate, women's age when their first child was born and consumption of alcohol and tobacco. The average consumption of alcohol reported by controls from developed countries was 6.0 g per day, i.e. about half a unit/drink of alcohol per day, and was greater in ever-smokers than never-smokers, (8.4 g per day and 5.0 g per day, respectively). Compared with women who reported drinking no alcohol, the relative risk of breast cancer was 1.32 (1.19-1.45, P<0.00001) for an intake of 35-44 g per day alcohol, and 1.46 (1.33-1.61, P<0.00001) for >/=45 g per day alcohol. The relative risk of breast cancer increased by 7.1% (95% CI 5.5-8.7%; P<0.00001) for each additional 10 g per day intake of alcohol, i.e. for each extra unit or drink of alcohol consumed on a daily basis. This increase was the same in ever-smokers and never-smokers (7.1% per 10 g per day, P<0.00001, in each group). By contrast, the relationship between smoking and breast cancer was substantially confounded by the effect of alcohol. When analyses were restricted to 22 255 women with breast cancer and 40 832 controls who reported drinking no alcohol, smoking was not associated with breast cancer (compared to never-smokers, relative risk for ever-smokers=1.03, 95% CI 0.98-1.07, and for current smokers=0.99, 0.92-1.05). The results for alcohol and for tobacco did not vary substantially across studies, study designs, or according to 15 personal characteristics of the women; nor were the findings materially confounded by any of these factors. If the observed relationship for alcohol is causal, these results suggest that about 4% of the breast cancers in developed countries are attributable to alcohol. In developing countries, where alcohol consumption among controls averaged only 0.4 g per day, alcohol would have a negligible effect on the incidence of breast cancer. In conclusion, smoking has little or no independent effect on the risk of developing breast cancer; the effect of alcohol on breast cancer needs to be interpreted in the context of its beneficial effects, in moderation, on cardiovascular disease and its harmful effects on cirrhosis and cancers of the mouth, larynx, oesophagus and liver.

Objective: To investigate the effect of growth hormone on the development of in vitro matured unstimulated human oocytes., Design: Randomized controlled study., Setting: Division of Reproductive Medicine, Department of Obstetrics and Gynecology, Faculty of Medicine, Chulalongkorn university., Material and Method: 108 germinal vesicle-stage oocytes were retrieved from 47 patients undergoing gynecologic surgery. They were aspirated either during gynecologic surgery or from excised ovaries. The oocytes were then cultured in vitro with or without growth hormone (1,000 ng/ml) in medium199 supplemented with sodium pyruvate, FSH, LH, antibiotic and synthetic serum. Incubation was done at 37 degree C with 5 per cent CO2 in air and nuclear stage was assessed after 18, 42, 66 and 90 h of incubation., Main Outcome Measure: Attainment of metaphase II and GVBD RESULTS: After in vitro culture, there were no significant differences in maturation and GVBD rate. 27 of 52 (51.9%) oocytes (GV) in growth hormone group matured to metaphase II compared with 25 of 53 (47.2%) GV in control group. GVBD rate for germinal vesicle-stage in growth hormone group was 76.9 per cent compared with 79.2 per cent in control group., Conclusion: Culture of immature oocytes in vitro with growth hormone results in similar maturation rate as that without GH.

This study was undertaken to evaluate the predictive value of the serum human chorionic gonadotropin (hCG) in pregnancies achieved by assisted reproductive techonology (ART). Two hundred and eighty-six pregnancies were studied retrospectively from September 1989 to June 1998. The serum hCG samples at 2-6 weeks after embryo transfer (ET) were analysed by fluoroimmunoassay. Pregnancy status was followed by ultrasonography. There were 100 nonviable pregnancies (NP), 140 viable single pregnancies (VSP) and 46 viable multiple pregnancies (VMP). The sensitivity, specificity, positive and negative predictive value of the D14 hCG (<160 mIU/ml) in distinguishing NP from VSP were 79 per cent, 75 per cent, 68 per cent and 84 per cent, respectively and of the D14 hCG (>350 mIU/ml) in distinguishing VMP from VSP were 82 per cent, 75 per cent, 56 per cent and 91 per cent, respectively. In conclusion, the serum hCG may be used to predict the outcome of early pregnancy achieved by ART.

The aim of this study was to compare day-3 serum inhibin B levels of poor and good ovarian responders in the IVF-program. The study group consisted of 20 poor ovarian responders. The control group composed of 40 good ovarian responders who had similar demographic characteristics. The serum inhibin B levels were analysed by two-site-immunosorbent assay or sandwich ELISA (Enzyme Linked ImmunoSorbent Assay). The mean inhibin B level of the study group was 113.18 +/- 57.96 picogram per millilitre and of the control group was 94.05 +/- 61.81 picogram per millilitre. There was no statistically significant difference. The results might be useful as base-line data for further study.

The Executive Board of the Medical Council of Thailand has set up an ad hoc committee to establish the regulations of practising of assisted reproductive technology. The committee assigned the Royal Thai College of Obstetricians and Gynecologists to supervise and take charge of the administration and formulate an annual report in accordance with the Thai Medical Council Declaration. The regulation was finally approved on October 9, 1997. It was announced in the Royal Gazette on December 26, 1997 and since then the prescription of standard measures for ART practice has been effected.

Objective: To evaluate the efficacy of zidovudine (ZDV) administered during labor and to the infants in the first 6 weeks of life in reduction of perinatal HIV-1 transmission., Design: Open label clinical trial. SITE: King Chulalongkorn Memorial Hospital, Bangkok, Thailand., Material and Method: One hundred asymptomatic, antiretroviral naive HIV-1 infected pregnant women who had either late or no prenatal care were recruited from the obstetric service of King Chulalongkorn Memorial Hospital, Bangkok, Thailand. They were given ZDV 300 mg orally every 3 hours during the intrapartum period until delivery. ZDV syrup 2 mg/kg orally every 6 hours were given to the infants immediately after birth for 6 weeks. Breast feeding was not allowed. Infant's blood for HIV-1 PCR test was obtained at age 1 day, and 1, 3 and 6 months. HIV-antibody test was determined at age 18 months. Infants with at least one positive HIV-1 PCR test performed at or after 1 month of age or positive HIV-antibody test at age 18 months were classified as HIV-1 infected infants., Results: There were 100 healthy infants delivered without complication. Fourteen infants were excluded due to; 13 lost to follow-up and 1 drug intolerance. Of the remaining 86 infants who were followed-up, 27 infants (31.4%) did not receive intrapartum ZDV treatment and 9 infants were HIV-1 infected. The perinatal transmission rate was 10.5 per cent, (95% CI 3.9, 17.1)., Conclusion: The result of this study suggests that intrapartum oral ZDV treatment in asymptomatic HIV-1 infected mothers together with ZDV treatment in the neonates for 6 weeks can reduce the rate of perinatal HIV-1 transmission. This regimen may be an alternative treatment for prevention of HIV-1 infection in infants born to HIV-1 seropositive mothers who have had either late or no prenatal care.

A multicenter randomized, double blind, placebo-controlled clinical trial was conducted to evaluate the effectiveness of a short course of oral zidovudine (ZDV) treatment in HIV-1 infected pregnant women, starting at 38 weeks of gestation plus ZDV infusion during labor until delivery, to reduce HIV-1 vertical transmission in non-breast fed infants. One hundred and eighty two asymptomatic antiretroviral naïve HIV-1 infected pregnant women were enrolled. Each patient was randomly allocated into either the ZDV or placebo group. The ZDV group received 250 mg ZDV orally twice a day initiated at 38 weeks' gestation until the onset of labor. During the intrapartum period, ZDV infusion at the rate of 2 mg/kg was administered within the first hour and then continuously infused at the rate of 1 mg/kg/h until delivery. The placebo group received an identical capsule during pregnancy and normal saline infusion during labor until delivery. HIV-1 transmission was documented by nested polymerase chain reaction in infants at birth and at 1, 3 and, 6 months of age. The estimated HIV-1 vertical transmission rate was 14.9 per cent (95% CI = 11.1 to 18.7) and 16.3 per cent (95% CI = 12.3 to 20.9) in ZDV and placebo group, respectively (p > 0.05). The short course ZDV in antiretroviral naïve pregnant women initiated at 38 weeks' gestation plus intrapartum ZDV infusion without treatment in the infants was not effective to prevent HIV-1 vertical transmission.

The mechanism of infertility associated with endometriosis is poorly understood. There is evidence supporting that women with ovarian endometriosis have a lower pregnancy rate than women with peritoneal lesions only. This study aimed to evaluate the effects of endometriotic contents contamination while retrieving oocytes on oocytes' quality. Thirty-eight infertile patients with endometriotic cysts from January 1993 to June 2000 were enrolled in this study. There were no statistically significant differences among the quality of oocytes and embryos from the contaminated, non-contaminated, and control group. However, the fertilization rate and pregnancy rate were impaired by the contamination of endometriotic contents. We conclude that ovarian endometriosis should be treated before starting in vitro fertilization program in order to increase the pregnancy outcome.

Objectives: To evaluate the contraceptive reliability and clinical effects of a monophasic oral contraceptive containing 20 microg ethinylestradiol and 150 microg desogestrel (Mercilon) in Thai women., Method: The study was carried out at the Family Planning Clinic of King Chulalongkorn Memorial Hospital, Bangkok, Thailand. One hundred and forty six healthy women of fertile age were enrolled and treated with the study oral contraceptives for 12 cycles. Clinical data of vaginal bleeding, side effects, blood pressure and body weight were assessed periodically., Results: One thousand four hundred and twenty five cycles were evaluated. No conception occurred. The cycles were almost regular. The incidence of irregular bleeding was highest in the first cycle, after the third cycle, the value was below 4 per cent. Side effects were very few. There was no change in blood pressure. A slight decrease in mean body weight was observed., Conclusion: The oral contraceptive containing 20 microg ethinylestradiol and 150 microg desogestrel has high contraceptive efficacy, good cycle control and minimal side effects.

This prospective study was undertaken to establish the correlation between the follicular diameter, oocyte recovery, metaphase II (MII) oocyte, and fertilization rate in intracytoplasmic sperm injection (ICSI) programs. Thirty one patients undergoing 31 ICSI cycles from August 1998 to January 1999 at King Chulalongkorn Memorial Hospital were studied. The patients were stimulated with either short or long GnRHa portocol. During transvaginal aspiration of follicles, follicular diameters were measured. The maturation stage of oocytes and fertilization rate were evaluated. Four hundred and seventy eight follicles were measured and classified into 3 groups, group A (<10 mm), group B (10-14 mm) and group C (>14 mm). The oocyte recovery rate and MII oocytes were different between groups A, B, and C but the fertilization rate was not different. In conclusion, the follicular diameter correlated with oocyte recovery and oocyte maturation but not with the fertilization rate in ICSI programs. The quality of embryos was lower in oocytes from small follicles.

A double uterus with a unilaterally obstructed hemivagina is a rare condition, usually associated with ipsilateral renal agenesis. Herein, we report two cases, the first case presenting with abdominal pain and pelvic mass. Hemihysterectomy was performed leaving the contralateral uterus intact. The second case presented with chronic foul smelling vaginal discharge. The diagnosis was a double uterus and pyocolpos of the left vagina. Excision of the left vaginal septum and drainage were performed. The postoperative course of both cases was uneventful and the patients were well at the six-week follow-up. An accurate diagnosis, appropriate management and the prevention of future fertility problems are discussed.

Objective: To compare the efficacy and safety of HyCoSy with chromolaparoscopy for the diagnosis of tubal occlusion and uterine abnormalities., Methods: Sixty infertile women in whom the cause of infertility was thought to be tubal occlusion or uterine abnormalities and who satisfied the inclusion and exclusion criteria as specified in the study protocol were included. HyCoSy was performed during the first half of the menstrual cycle at least 24 hours prior to chromolaparoscopy. The results from both HyCoSy and chromolaparoscopy were compared in assessing tubal occlusion and uterine pathology., Results: For the evaluation of fallopian tubes, we found corresponding results between HyCoSy and chromolaparoscopy in 80.0%. The agreement between both procedures in assessing the uterine pathology was 80.4%. Twenty-two women experienced adverse events. The most common complaint was pelvic pain. Other events encountered were: nausea (3 women) and vaginal bleeding (2 women). All events were thought to be not related to the study drug., Conclusion: HyCoSy showed good diagnostic performance in the evaluation of the fallopian tubes and uterus in infertile women. The adverse events reported in this study are minor and procedure-related (catheter insertion) rather than the trial substance.

The aim of this study was to investigate the efficacy of two different embryo biopsy techniques, direct aspiration and partial zona dissection (PZD)-push, on subsequent in vitro and in vivo development of 8-cell stage mouse embryos. It was found that the rates of normal blastocyst formation and hatching blastocysts of direct aspiration, PZD-push, solution control and control embryos were not significantly different (80.8%, 81.6%, 84.5%, 86.7% and 71.9%, 72.3% and 74.6%) respectively. There was, however, a significant reduction in rate of complete hatching blastocysts (P < 0.1) (72.9% aspiration versus 85.2 per cent solution control and 86.4% control) and rate of live-born fetuses (24.2% aspiration versus 43.3% solution control and 41.2% control) (P < 0.05) in the direct aspiration group but no significant difference in the PZD-push group (80.3% of complete hatched blastocysts and 33.8% of live-born fetuses). These findings indicated that embryo biopsy with PZD-push was superior to the direct aspiration method. This mouse embryo biopsy model was useful in advancing development of biopsy technique for human preimplantation genetic diagnosis.

The clinical efficacy and side effects of oral gestrinone, 2.5 mg twice weekly, were evaluated in this prospective study involving 22 patients with laparoscopically confirmed endometriosis. All patients came to the hospital with infertility problem. After 6 months of treatment, 81 per cent of patients had amenorrhea. Dysmenorrhea and pelvic pain were reduced from 90.9 per cent to 14.3 per cent and 81.8 per cent to 9.5 per cent respectively. Pelvic tenderness and induration were improved from 55.6 per cent and 50.0 per cent to 15.8 per cent and 10.5 per cent respectively. Androgenic effects such as acne was founded in 18.2 per cent of the patients. Return of fertility was observed in 25 per cent (5 patients) after 30-254 days post treatment. No serious side effect was detected during the treatment. The results suggest that gestrinone may be considered an option for the treatment of endometriosis related infertility.

Fifteen infertile women who required tubal passage evaluation by chromolaparoscopy were recruited. Those who had lower genital tract infection or abnormal uterine bleeding were excluded from the study. Transvaginal HyCoSy was performed during the first half of the menstrual cycle at least 24 hours prior to chromolaparoscopy. The results from both transvaginal HyCoSy and chromolaparoscopy were compared in assessing tubal patency and uterine pathology. A high correlation was noted regarding uterine examination using transvaginal HyCoSy compared with chromolaparoscopy (sensitivity, specificity, PPV and NPV were 91.7%, 33.3%, 84.6% and 50%, respectively). The correlation of the outcome between the two procedures in assessing tubal patency, when combining both tubes, was also high (sensitivity, specificity, PPV and NPV were 100%, 55.6%, 80% and 100%, respectively). The most common adverse event was only mild pelvic pain which did not necessitate any treatment. These preliminary results reveal the potential value of transvaginal HyCoSy as an alternative in infertility investigations. It seems to be as effective but less invasive than conventional chromolaparoscopy. The adverse events reported in this study relate more to the procedure (catheter insertion) rather than the trial substance.

In men with obstructive azoospermia, bypass surgery would obviously be the most acceptable form of treatment as it gives the couple an opportunity to conceive naturally. However, when this has failed to restore patency or when surgery is not feasible (congenital absence of vas), fertility treatment using spermatozoa aspirated from the epididymis should be considered. Percutaneous epididymal sperm aspiration (PESA) is more acceptable to patients than micro epididymal sperm aspiration (MESA) because it eliminates the requirement for a general anaesthetic, post-operative pain, and the risk of haematoma formation, thus allowing a rapid return to normal activity of the husband. To our knowledge, this is the first reported case in Thailand where a pregnancy resulted in a couple whose infertility was due to azoospermia from the congenital absence of vas deferens. Spermatozoa collected through PESA were used in ICSI to achieve fertilization. The PESA technique, due to its simplicity is the choice of treatment for obstructive azoospermia.

Objective: To assess the reliability of sex determination in mouse preimplantation embryos using the two-step polymerase chain reaction method., Setting: Division of Immunology, Department of Microbiology and Division of Reproductive Medicine, Department of OB/GYN., Methods: The Sry and Zfy genes, known to be present in the sex-determining region of mouse Y chromosome, were selected for Y-specific target sequences and DXNds 3 locus located on mouse X chromosome was served as the internal control sequence. DNAs extracted from heart blood of male and female mice were used to test the correctness and specificity of the selected primers using the two-step PCR method. The same experimental conditions were then used to amplify the single copy genes in single mouse blastomeres with two pairs of primers for each of the target sequences. The sex-determined embryos were transferred to the uteri of pseudopregnant recipients to test the consistency of the assay system., Results: All male and female blood DNA sample results confirmed the correct sex identification of the origin (100%). Nineteen of 20 single blastomeres showed the accurate diagnosis when compared with theirs 7/8 embryos. The sex of 36 of 37 mouse pups born from biopsied male and female embryos agreed with the predicted sex., Conclusion: The reliable genetic analysis of sex chromosome- specific sequences in single cell is possible by the two-step PCR method and could be applied for diagnosis of defective genes of human preimplantation embryos derived from the in vitro fertilization program.

Serum FSH level may be an indicator of the functional potential of the ovary. The basic control of LH and FSH is by a negative feedback system involving the hypothalamus, anterior pituitary, and ovary. Basal serum FSH on day 3 of previous cycles in 36 patients underwent ovarian stimulation with a combination of GnRH agonist and FSH and hMG in an IVF program were evaluated. The results in this study show that a high basal serum FSH level is associated with a higher cancellation rate, a lower peak serum E2 level, lower number of oocytes retrieved and a lower clinical pregnancy rate.

Cervical cancer is probably caused by a sexually transmitted agent. A case-control study was conducted in three hospitals in Thailand to investigate further the role of male sexual behavior, particularly regarding sexual contacts with prostitutes, in the development of this disease. Data were obtained from interviews with 225 married women with invasive squamous cell cervical carcinoma and 791 hospitalized controls, all of whom reported having only one sexual partner, and from interviews with their husbands. Risk of cervical cancer was strongly related to the women's husbands having visited prostitutes without using a condom when the husbands were less than 30 years old. A strong increasing trend in risk in relation to decreasing frequency of the husbands' condom use with prostitutes was observed, and a weaker increasing trend in risk with husbands' estimated lifetime total number of visits to prostitutes was found. The average latent period between the women's likely initial exposure to a sexually transmitted oncogenic agent and her diagnosis of invasive cervical cancer was about a quarter of a century. Regular use of condoms by customers of prostitutes could reduce the number of invasive cervical cancer cases in the general population of Thailand by at least one fourth.

We have presented a case of bilateral tubal pregnancy following in vitro fertilization and embryo transfer. The most likely cause, in our opinion, is 'spray effect' which was discussed. The diagnosis of ectopic pregnancy must always be kept in mind in IVF&ET program, due to its high incidence of occurrence compared to natural conception. Although the incidence of bilateral tubal pregnancy is not high, one should identify the opposite adnexa when tubal pregnancy is diagnosed in an IVF&ET patient. Early diagnosis is essential for prevention of significant maternal morbidity and mortality.

A cross-sectional study was designed to determine trabecular bone density in 75 long-term depot-medroxyprogesterone acetate (DMPA) users (> 3 yr) matched with non-DMPA users by age, body mass index (18-25), limitation of age (< 45 yr), and body weight (< 60 kg). The long-term DMPA cases were divided into 3 groups according to duration of injectable contraceptive use. Neither cases nor controls had a smoking or chronic alcohol consumption history. Cases and controls were matched by age. Trabecular bone of the femoral neck were assessed by X-ray and interpreted by a single-blinded radiologist. Trabecular bone patterns were graded according to Singh's Index. Blood collection for determination of estradiol, prolactin, calcium, phosphorus, and medroxyprogesterone acetate were performed in cases and controls. Venous blood was taken at twelfth week of injection of DMPA and within 5 days after menstrual bleeding cessation in the controls. Mean trabecular bone in the cases was 5.5 +/- 0.6 (range 4-6). It was not statistically different from that in the controls (mean 5.5 +/- 0.6, range 2-6). No statistically significant difference of serum, calcium, phosphorus, prolactin, and estradiol was seen in the cases when compared to controls at mid follicular phase of normal menstrual cycle. Serum MPA of individual case at twelfth week of injection was 4.1 +/- 1.1 nmol/l. In conclusion, trabecular bone density in long-term DMPA users were not statistically different from normal menstruating women who have not received injectable DMPA.

This is a preliminary report on the first 24 babies born during the period of 1 year follow-up after IVF-ET in Chulalongkorn Hospital. Based on this study, our data suggests that these children have a normal range of growth and development. There was also no evidence of major congenital anomalies identified in spite of some minor ones. However, a need for more detail and on-going long-term follow-up at least until school age is being planned.

Endometrial bleeding and alteration in blood coagulation, fibrinolysis, ovarian function and endometrial morphology were studied in twelve normally menstruating women who received the injectable contraceptive norethisterone enanthate (NET-EN) 200mg at 60-day intervals. Levels of clotting factor VII declined significantly during episodes of irregular bleeding compared to those during both normal pretreatment menstruation and the bleeding-free period during treatment. Antithrombin III and fibrinolytic activity, expressed by euglobulin lysis time, showed no marked change. The average level of progesterone during the bleeding-free period was slightly but significantly lower than that during the bleeding period. There were no significant alterations in the mean levels of estradiol and the ratio of estradiol to progesterone. The endometrial biopsies showed considerable individual variation and seem to be independent of the effects of NET-EN on ovarian function.