Can biomarkers identified from the uterine fluid transcriptome be used to establish a noninvasive endometrial receptivity prediction tool? A proof-of-concept study.

BACKGROUND: Embryo implantation in a receptive endometrium is crucial for successful pregnancy. Endometrial receptivity (ER) prediction tools based on endometrial transcriptome biomarkers by endometrial biopsy have been used to guide successful embryo implantation in in vitro fertilization (IVF) patients. However, no reliable noninvasive ER prediction method has been established, and one is greatly needed. We aimed to identify biomarkers from uterine fluid transcriptomic sequencing data for establishing noninvasive ER prediction tool and to evaluate its clinical application potential in patients undergoing IVF. METHODS: The non-invasive RNA-seq based endometrial receptivity test (nirsERT) was established by analyzing transcriptomic profile of 144 uterine fluid specimens (LH + 5, LH + 7, and LH + 9) at three different receptive status from 48 IVF patients with normal ER in combination with random forest algorithm. Subsequently, 22 IVF patients who underwent frozen-thaw blastocyst transfer were recruited and analyzed the correlation between the predicted results of nirsERT and pregnancy outcomes. RESULTS: A total of 864 ER-associated differentially expressed genes (DEGs) involved in biological processes associated with endometrium-embryo crosstalk, including protein binding, signal reception and transduction, biomacromolecule transport and cell-cell adherens junctions, were selected. Subsequently, a nirsERT model consisting of 87 markers and 3 hub genes was established using a random forest algorithm. 10-fold cross-validation resulted in a mean accuracy of 93.0%. A small cohort (n = 22) retrospective observation shows that 77.8% (14/18) of IVF patients predicted with a normal WOI had successful intrauterine pregnancies, while none of the 3 patients with a displaced WOI had successful pregnancies. One patient failed due to poor sequencing data quality. CONCLUSIONS: NirsERT based on uterine fluid transcriptome biomarkers can predict the WOI period relatively accurately and may serve as a noninvasive, reliable and same cycle test for ER in reproductive clinics. TRIAL REGISTRATION: Chinese Clinical Trial Registry: ChiCTR-DDD-17013375. Registered 14 November 2017, http://www.chictr.org.cn/index.aspx .

An endometrium of type C along with an endometrial thickness of < 8 mm are risk factors for ectopic pregnancy after stimulated cycles with fresh embryo transfer.

BACKGROUND: The study investigated whether specific ultrasonographically observed endometrial features (including endometrium type and thickness) were linked to ectopic pregnancy after stimulated cycles with fresh embryo transfer. METHOD: Of 6246 pregnancy cycles after fresh embryo transfer, 6076 resulted in intrauterine pregnancy and 170 in ectopic pregnancy. The primary outcome of the study was ectopic pregnancy, with the main variables being endometrium type and endometrial thickness. Univariate and subsequent multiple-stepwise logistic regression analyses were used to identify the risk factors of ectopic pregnancy. RESULTS: 1. Compared with patients with an endometrial thickness ≥ 8 mm, the adjusted odds ratio for those with an endometrial thickness < 8 mm was 3.368 (P < 0.001). The adjusted odds ratio for women with a type-C endometrium was 1.897 (P = 0.019) compared with non-type C. 2. A larger dose of gonadotropin used during controlled ovarian hyperstimulation was a protective factor against ectopic pregnancy (P = 0.008). 3. The GnRH antagonist protocol (P = 0.007) was a risk factor for ectopic pregnancy, compared with the use of GnRH agonists. CONCLUSION: (1) An endometrial thickness < 8 mm coupled with a type C endometrium significantly increased the risk of ectopic pregnancy after fresh embryo transfer. (2) A thin endometrial thickness and a type C endometrium could be further related to an abnormal endometrial receptivity/peristaltic wave. (3) Patients at a high risk of ectopic pregnancy should therefore be given special attention, with early diagnosis during the peri-transplantation period may assist in the prevention of ectopic pregnancy.

Endometrial fluid aspiration immediately prior to embryo transfer does not affect IVF/vitrified-warmed embryo transfer outcomes - a prospective matched cohort study.

RESEARCH QUESTION: Does the endometrial aspiration of ultrasound-invisible fluid immediately preceding embryo transfer affect IVF/vitrified-warmed embryo transfer outcomes? DESIGN: A prospective matched cohort study was conducted in 96 women and 96 control participants to assess the effect on pregnancy outcomes of endometrial aspiration performed immediately before embryo transfer. This study was carried out at a university-affiliated assisted reproductive medical centre between January 2019 and December 2019. Patients were divided into two groups. The EA group had cycles with endometrial aspiration of ultrasound-invisible fluid performed before embryo transfer and the non-EA group featured cycles without endometrial aspiration. The EA group was matched by propensity score with the non-EA group in a 1:1 ratio. The EA group consisted of 99 participants before and 96 participants after propensity score matching. There were 203 and 96 participants in the non-EA group before and after propensity score matching. RESULTS: No significant differences were detected in the baseline characteristics and cycle characteristics of the EA and non-EA groups. No significant between-group differences were found in reproductive outcomes in the overall population. Subgroup analysis of blastocyst transfer cycles showed the implantation rate was significantly higher in the EA group (61 women per group, 57.1% versus 40.8%, relative risk 1.40, 95% confidence interval 1.04-1.88; P = 0.022). Live birth rate, clinical pregnancy rate, ongoing pregnancy rate and multiple pregnancy rate were not different among the groups. CONCLUSIONS: Endometrial aspiration immediately preceding embryo transfer does not affect IVF/vitrified-warmed embryo transfer outcomes. Interestingly, it might improve the vitrified-warmed blastocyst implantation rate. Randomized controlled trials are needed to confirm this result.

Associations between dyslipidaemia and pregnancy outcomes in the first complete cycle of IVF/ICSI: a real-world analysis.

RESEARCH QUESTION: Are there associations between dyslipidaemia and pregnancy outcomes in the first complete cycle of IVF/intracytoplasmic sperm injection (ICSI)? DESIGN: This long-term, retrospective real-world analysis involved 5030 infertile women who underwent a first complete IVF/ICSI cycle between January 2015 and October 2020. They were categorized into dyslipidaemia (n = 1903) and control (n = 3127) groups according to serum lipid concentrations before ovarian stimulation. Propensity score matching and multivariable logistic regression were used to control for confounding variables. RESULTS: In the raw cohort, women with dyslipidaemia had a significantly increased late miscarriage rate (P = 0.039), decreased term birth rate (P = 0.002) and decreased live birth rate (P = 0.005) compared with non-dyslipidaemic women. In the propensity score-matched cohort, the term birth rate (P = 0.038) and live birth rate (P = 0.044) were significantly lower in the dyslipidaemia group (n = 1686) than the controls (n = 1686). Multivariable logistic regression indicated that infertile women with dyslipidaemia (P = 0.026) and elevated serum total cholesterol concentrations (total cholesterol ≥5.20 mmol/l; P = 0.028) were significantly less likely to have a live birth. Rates of late miscarriage (P = 0.027), term birth (P = 0.003) and live birth (P = 0.010) differed significantly among women with normal, borderline increased and increased serum lipid concentrations. Compared with controls, women with increased serum lipid concentrations had a significantly higher late miscarriage rate, lower term birth rate and lower live birth rate. Women with increased serum lipid concentrations were significantly less likely than controls to have a live birth. CONCLUSIONS: Dyslipidaemia, total cholesterol ≥5.20 mmol/l and degrees of elevated serum lipid concentrations are negatively associated with live birth rate in the first complete IVF/ICSI cycle in infertile women.

Chromosomal analysis of blastocysts from balanced chromosomal rearrangement carriers.

Balanced chromosomal rearrangements (CRs) are among the most common genetic abnormalities in humans. In the present study, we have investigated the degree of consistency between the chromosomal composition of the blastocyst inner cell mass (ICM) and trophectoderm (TE) in carriers with balanced CR, which has not been previously addressed. As a secondary aim, we have also evaluated the validity of cleavage-stage preimplantation genetic diagnosis (PGD) based on fluorescence in situ hybridization (FISH) of blastocysts from CR carriers. Blastocyst ICM and TE were screened for chromosomal aneuploidy and imbalance of CR-associated chromosomes based on whole-genome copy number variation analysis by low-coverage next-generation sequencing (NGS) following single-cell whole-genome amplification by multiple annealing and looping-based amplification cycling. The NGS results were analyzed without knowledge of cleavage-stage FISH results. NGS results for blastocyst ICM and TE from CR carriers were 86.49% (32/37) consistent. Of the 1702 (37 × 46) chromosomes examined, 99.47% (1693/1702) showed consistency. However, only 40.0% (18/45) of all embryos had consistent results for chromosomes involved in CR, as determined by blastocyst NGS and cleavage-stage FISH. Of the 85 CR-affected chromosomes analyzed by FISH, 37.65% (32/85) were incongruous with NGS results, with 87.5% (28/32) showing imbalanced composition by FISH but balanced composition by NGS. These results indicate that chromosomal composition of blastocyst ICM and TE in balanced CR carriers is highly consistent, and that PGD based on cleavage-stage FISH is inaccurate; therefore, using blastocyst TE biopsies for NGS-based PGD is recommended for identifying chromosomal imbalance in embryos from balanced CR carriers.

Morphometric analysis and developmental comparison of embryos from carriers with balanced chromosomal rearrangements in preimplantation genetic diagnosis cycles.

The morphological parameters of embryos from 22 carriers with balanced chromosomal rearrangements (CRs) were quantified and evaluated to determine their possible link to chromosomal composition. The morphometric characteristics of 168 embryos diagnosed by fluorescence in situ hybridisation were measured using an imaging tool and then analysed retrospectively. The mean zygotic diameter of normal-balanced embryos was significantly smaller compared with that of abnormal embryos (P=0.015). In addition, the reduction in total cytoplasmic volume for Day-3 embryos was significantly lower in normal or balanced embryos than in abnormal embryos (P=0.027). Moreover, the pronuclear volumes of embryos that failed to reach the blastocyst stage were significantly smaller compared with those of blastocysts (P=0.016). These findings indicate that morphometric characteristics are correlated with developmental outcomes as well as with chromosomal composition in embryos from balanced CR carriers. However, an effective indicator of developmental outcomes may not accurately reflect chromosomal composition. Combining morphometric and traditional qualitative assessment may increase the precision and standardisation of embryo evaluation as well as contributing to improved efficiency of preimplantation genetic diagnosis by selecting embryos with high developmental potential and preferentially testing embryos predicted to have a low risk of chromosomal imbalance.

[Influence of chromosomal polymorphisms on the clinical outcome of patients undergoing in vitro fertilization embryo transfer].

OBJECTIVE: To explore the influence of chromosome polymorphisms on the outcome of in vitro fertilization embryo transfer (IVF-ET). METHODS: Patients who completed the first cycle of in vitro fertilization fresh embryo transfer were retrospective studied. Patients with the chromosome polymorphisms were classified as to the study group (200 treatment cycles), all patients with normal chromosomes at the same period were classified as the control group (4777 treatment cycles). RESULTS: No significant difference was found between the chromosome polymorphisms and the control groups in terms of clinical pregnancy rate (44.50% vs. 39.85%, P=0.750), early abortion rate (15.73% vs. 10.79%, P=0.163) and live birth rate per cycle (34.5% vs. 30.73%, P=0.437) except for fertilization rate (60.94% vs. 64.08%, P=0.001), cleavage rate (95.01% vs. 97.09%, P=0.000) and good quality embryo rate (53.8% vs. 58.2%, P=0.001). CONCLUSION: Chromosomal polymorphisms appeared to have no adverse influence on the outcome of IVF-ET treatment.

Homeostatic Model Assessment for Insulin Resistance Is Associated With Late Miscarriage in Non-Dyslipidemic Women Undergoing Fresh IVF/ICSI Embryo Transfer.

Objective: To evaluate the associations between homeostatic model assessment for insulin resistance (HOMA-IR) and pregnancy outcomes in non-dyslipidemic infertile women undergoing in vitro fertilization/intracytoplasmic sperm injection-embryo transfer (IVF/ICSI-ET). Materials and Methods: This is a retrospective study involving 3,615 non-dyslipidemic infertile women who attend to the Reproductive Medicine Center of Xiangya Hospital, Central South University (CSU) between January 2014 and October 2021. Eligible participants were divided into three groups according to the quartiles of HOMA-IR: Group 1 (HOMA-IR <1.46), Group 2 (1.46 to <2.71) and Group 3 (HOMA-IR ≥2.71). Baseline data, clinical characteristics during the assisted reproductive technology (ART) procedure, pregnancy, and neonatal outcomes were compared among the three groups. Subgroup analysis based on presence or absence of the polycystic ovary syndrome (PCOS) status was also performed to analyze the effects of HOMA-IR among non-PCOS populations. Results: The late miscarriage rate and percentage of macrosomia increased with the HOMA-IR group (for late miscarriage rate: 2.23% vs. 3.04% vs. 7.35%, P<0.001; for macrosomia: 0.21% vs. 1.70% vs. 3.23%, P=0.002). Increased HOMA-IR (HOMA-IR≥2.71) was positively associated with late miscarriage (crude OR 3.50, 95% CI 1.64-7.47, P=0.001; adjusted OR 3.56, 95% CI 1.56-8.15, P=0.003). In the subgroup analysis, there were 3,165 participants in the non-PCOS group and 450 were assigned to the PCOS group. Late miscarriage rate increased with the HOMA-IR group among non-PCOS populations (2.20% vs. 3.03% vs. 7.67%, P<0.001). Late miscarriage rate of PCOS women were comparable among the three HOMA-IR groups (2.50% vs. 3.06% vs. 5.71%, P=0.634). Among non-PCOS women, increased HOMA-IR (HOMA-IR≥2.71) was positively associated with late miscarriage (crude OR 3.71, 95% CI 1.66-8.30, P=0.001; adjusted OR 3.82, 95% CI 1.59-9.17, P=0.003). Conclusions: Late miscarriage rate and prevalence of macrosomia increased with the HOMA-IR index. Preconception HOMA-IR is an independent risk factor for late miscarriage in normolipidemic women undergoing IVF/ICSI-ET. Controlling insulin resistance before ART might prevent the occurrence of late miscarriage and macrosomia.

An Endometrial Thickness < 8 mm Was Associated With a Significantly Increased Risk of EP After Freeze-Thaw Transfer: An Analysis of 5,960 Pregnancy Cycles.

Introduction: Endometrium characteristics that are most likely to induce ectopic pregnancy were investigated on the basis of the data of 5,960 pregnant freeze-thaw cycles. Methods: A total of 5,960 pregnancy cycles after freeze-thaw embryos transfer were included, with the number of intrauterine and ectopic pregnancies being 5,777 and 183, respectively. Ectopic pregnancy was the primary outcome. Endometrial thickness was the main measured variable. The risk factors of ectopic pregnancy were eventually determined based on univariate analysis and subsequent multiple-stepwise logistic regression analysis. Results: 1. After adjusting for confounders, endometrial thickness could independently predict ectopic pregnancy. The adjusted odd ratios for women with endometrial thickness in the ranges of < 8 mm, 8-9.9 mm, and 10-11.9 mm were 3.270 [95% confidence interval (CI), 1.113-9.605, P = 0.031], 2.758 (95% CI, 0.987-7.707, P = 0.053), and 1.456 (95% CI, 0.502-4.225, P = 0.489), respectively, when compared with those having an endometrial thickness of 12-13.9 mm. 2. Endometrial type and preparation protocol were however not identified as risk factors for ectopic pregnancy. Discussion: 1. After freeze-thaw embryo transfer, risks of ectopic pregnancy were significantly higher when the endometrial thickness was < 8 mm. 2. A thin endometrial thickness could be linked with abnormal endometrial peristaltic waves or abnormal endometrial receptivity. 3. Adequate attention should therefore be paid to patients with a thin endometrial thickness to prevent EP or to achieve early diagnosis during the peri-transplantation period.

[Impact of sperm source and parameters on the outcome of intracytoplasmic sperm injection].

OBJECTIVE: To evaluate the impact of sperm source and sperm parameters on the outcome of intracytoplasmic sperm injection (ICSI). METHODS: This retrospective study included 433 ICSI cycles from June 2005 to December 2008 in Reproductive Medical Center of Xiangya Hospital. The patients were divided into 2 major groups according to the source of spermatozoa used for ICSI: ejaculated (group A, n=336) and epididymal (group B, n=97). Group A was divided into 3 subgroups according to the sperm parameters: normal (Group A1, n=95), single parameter defect (Group A2, n=119), and multiple parameter defect (Group A3, n=122). RESULTS: The basic characteristics among the 4 groups had no statistic difference (P>0.05), and the difference in the fertilization rate, normal fertilization rate, cleaving embryo rate,good quality embryo rate, implanted rate, clinical pregnancy rate, and early abortion rate among the 4 groups were not significant (P>0.05). CONCLUSION: The outcome is similar no matter whether the spermatozoa is from ejaculated sperm or epididymis. ICSI can treat male infertility of various factors, and the outcome is the same with one or multiple sperm parameter abnormality. ICSI with epididymal spermatozoa through percutaneous epididymal sperm aspiration is effective for infertility due to obstructive azoospermia.

[Sperm-oocyte interaction and in vitro fertilization clinical outcomes in patients with unexplained infertility].

OBJECTIVE: To determine the sperm-zona pellucida (ZP) binding and ZP-induced acrosome reaction in patients with unexplained infertility, and to discuss the relationship between ZP-induced acrosome reaction and fertilization rate. METHODS: We compared the fertilization rate and good embryo rate in patients with unexplained infertility after fertilization in 2 ways. Based on the causes of infertility, patients were divided into an unexplained infertility group (Group A) and a pure female tubal factor group (Group B). Oocytes which were obtained by super ovulation from 25 patients with unexplained infertility were randomly divided into 2 groups with conventional in vitro fertilization (IVF) (Group A1) and intracytoplasmic sperm injection (ICSI) fertilization (Group A2). The pure female tubal factor group (Group B) had conventional IVF. We conducted sperm-ZP binding and ZP-induced acrosome reaction experiments with 2 groups of men's sperms separately. We compared the number of sperm-egg binding and ZP-induced acrosome reaction rate and discussed the relationship between the ZP-induced acrosome reaction and fertilization rate, and also the fertilization rate, good embryo rate and pregnancy rate in patients with unexplained infertility after fertilization in 2 ways. RESULTS: The average number of sperm-egg binding (78.29 ± 16.31) and the ZP-induced acrosome reaction rate (55.87 ± 27.69) % in Group A were lower than those of Group B [94.63 ± 6.72, (82.53 ± 17.99)%]. The difference between the average number of sperm-egg binding and the ZP-induced acrosome reaction was significant (P <0.01). The fertilization rate of Group A1 was significantly lower than that of Group B and Group A2 (P <0.01). But there was no significant difference in the good embryo rate among the 3 groups. There was no significant difference between Group A2 and B in fertilization rate and good embryo rate (P <0.05). There was no significant difference in pregnancy rate between Group A and B (P <0.05). Fertilization rate and the rate of acrosome reaction had marked positive correlation with statistical significance (r =0.932, P <0.01). CONCLUSION: ZP binding and ZP-induced acrosome reaction are very important experiments in sperm function test for patients with unexplained infertility. It can not only effectively avoid no embryo transferring due to complete failure of fertilization but also get a desirable outcome of pregnancy using half-ICSI fertilization in patients with unexplained infertility.

Early prediction of live birth for assisted reproductive technology patients: a convenient and practical prediction model.

Live birth is the most important concern for assisted reproductive technology (ART) patients. Therefore, in the medical reproductive centre, obstetricians often need to answer the following question: "What are the chances that I will have a healthy baby after ART treatment?" To date, our obstetricians have no reference on which to base the answer to this question. Our research aimed to solve this problem by establishing prediction models of live birth for ART patients. Between January 1, 2010, and May 1, 2017, we conducted a retrospective cohort study of women undergoing ART treatment at the Reproductive Medicine Centre, Xiangya Hospital of Central South University, Hunan, China. The birth of at least one live-born baby per initiated cycle or embryo transfer procedure was defined as a live birth, and all other pregnancy outcomes were classified as no live birth. A live birth prediction model was established by stepwise multivariate logistic regression. All eligible subjects were randomly allocated to two groups: group 1 (80% of subjects) for the establishment of the prediction models and group 2 (20% of subjects) for the validation of the established prediction models. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of each prediction model at different cut-off values were calculated. The prediction model of live birth included nine variables. The area under the ROC curve was 0.743 in the validation group. The sensitivity, specificity, PPV, and NPV of the established model ranged from 97.9-24.8%, 7.2-96.3%, 44.8-83.8% and 81.7-62.5%, respectively, at different cut-off values. A stable, reliable, convenient, and satisfactory prediction model for live birth by ART patients was established and validated, and this model could be a useful tool for obstetricians to predict the live rate of ART patients. Meanwhile, it is also a reference for obstetricians to create good conditions for infertility patients in preparation for pregnancy.

Uterine size and volume are associated with higher live birth rate in patients undergoing assisted reproduction technology: A prospective cohort study.

To investigate how uterine size and volume are associated with live birth rate in patients undergoing assisted reproduction technology.This prospective cohort study was conducted at the Reproductive Medicine Centre from January 2010 to May 2017. Multivariate binary logistic regression was used to evaluate the relations between uterine size, total volume, and live birth outcomes, after they were adjusted for the main influencing factors.A total of 7320 women of clinical pregnancy were enrolled. Compared with uterine lengths of 50 to 59 mm (referent), women with uterine lengths ≥60 mm had a lower live birth rate (RR = 1.541). Compared with uterine widths of ≥50 mm (referent), women with uterine widths <30 mm had a lower live birth rate (RR = 1.430). Compared with uterine anteroposterior diameters of <30 mm (referent), women with uterine anteroposterior diameters ≥50 mm had a lower live birth rate (RR = 1.636). Compared with uterine volumes of 30 to 49 mL (referent), women with volumes <30 mL and ≥70 mL had lower live birth rates (RR = 1.368 and 1.742, respectively).Our findings indicate that uterine sizes and volumes that were too large or too small reduced the live birth rate.

Uterine size and volume are associated with a higher clinical pregnancy rate in patients undergoing assisted reproduction technology: A longitudinal study (A STROBE-compliant article).

The aim of this study was to investigate the relationships between uterine size and volume and clinical pregnancy rate.This longitudinal study was conducted among patients undergoing assisted reproduction technology (ART) treatment at the Reproductive Medicine Center from January 2010 to May 2017, all of whom provided informed consent to participate in the study. The uterine size, for all patients, was measured by transvaginal ultrasonography before ovarian stimulation. Clinical pregnancy was diagnosed by ultrasound confirmation of at least an intrauterine gestational sac and fetal cardiac activity 4 weeks after embryo transfer.A total of 11,924 patients were enrolled in this study. Compared to patients with uterine lengths of 50 to 59 mm (referent), patients with uterine lengths ≥60 mm had a lower clinical pregnancy rate. Compared to patients with uterine widths of 30 to 39 mm (referent), patients with uterine widths of 40 to 49 mm and those with uterine widths of ≥50 mm had a lower clinical pregnancy rate. Compared with those with a uterine anteroposterior diameter of <30 mm (referent), patients with uterine anteroposterior diameters of ≥50 mm had a lower clinical pregnancy rate. Compared with those with a uterine volume of 30 to 49 mL (referent), patients with a uterine volume ≥70 mL had a lower clinical pregnancy rate.The patients with an optimal uterine length, width, anteroposterior diameter, and volume had a higher clinical pregnancy rate than those with suboptimal uterine measurements. Uterine sizes and volumes that were too large reduced the clinical pregnancy rate.

Uterine dimensions in gravida 0 phase according to age, body mass index, and height in Chinese infertile women.

The aim of this study was to describe the size and the shape of gravida-0 uteri in infertile Chinese Han women according to age, height, and body mass index (BMI).Registered data obtained from the Department of Reproductive Medicine, Xiangya Hospital of Central South University, were collected and analyzed. The length, width, and anteroposterior diameter of the uteri of nonpregnant women aged 20 to 45 years were measured by transvaginal ultrasonography. Statistical analyses among different populations were conducted using a 1-way analysis of variance analysis or a Kruskal-Wallis H test.A total of 5726 primary infertile women were enrolled. The mean age of the sample group was 29.18 ±â€Š4.22 years, and the mean BMI and the mean height of them were 21.51 ±â€Š2.91 kg/m and 158.13 ±â€Š4.71 cm, respectively. The mean uterine length, width, anteroposterior diameter, and L/W ratio were 49.33 ±â€Š7.00 mm, 39.94 ±â€Š7.23 mm, 44.95 ±â€Š8.11 mm, and 1.2662 ±â€Š0.2465, respectively. There were a statistically significant positive correlations between uterine length, width, anteroposterior diameter, and age in infertile women (all P < .001). Uterine L/W ratio gradually decreased with age, which was statistically significant (P < .001). The correlations between uterine length, width, anteroposterior diameter, and height were also considered statistically significant (all P < .001), while there was no correlation between L/W ratio and height. The results showed that uterine size and BMI had no statistical significance.The uterine length, width, and anteroposterior diameter gradually increased with increased age and height, but the increasing extents was different, and the uterine shape became rounder with age and had not changed with height in primary infertile women.

[Preimplantation genetic diagnosis of Duchenne muscular dystrophy by single cell triplex PCR].

OBJECTIVE: To detect two exons of Duchenne muscular dystrophy (DMD) gene and a gender discrimination locus amelogenin gene by single cell triplex PCR, and to evaluate the possibility of this technique for preimplantation genetic diagnosis (PGD) in DMD family with DMD deletion mutation. METHODS: Single lymphocytes from a normal male, a normal female, two DMD patients (exon 8 and 47 deleted, respectively) and single blastomeres from the couples treated by the in vitro fertilization pre-embryo transfer (IVF-ET) and without family history of DMD were obtained. Exons 8 and 47 of DMD gene were amplified by a triplex PCR assay, the amelogenin gene on X and Y chromosomes were co-amplified to analyze the correlation between embryo gender and deletion status. RESULTS: In the normal single lymphocytes, the amplification rate of exons 8 and 47 of DMD and amelogenin gene were 93.8%, 93.8%, and 95.3% respectively. The false positive rate was 3.3%. In the exon 8 deleted DMD patient, the amplification rate of exon 47 of DMD and amelogenin gene was 95.8%, and the false positive rate was 3.3%. In the exon 47 deleted DMD patient, the amplification rate of exon 8 of DMD and amelogenin gene was 95.8%, and the false positive rate was 0. In the single blastomeres, the amplification rate of exons 8 and 47 of DMD and amelogenin gene was 82.5%, 80.0% and 77.5%, respectively, and the false positive rate was 0. CONCLUSION: The single cell triplex PCR protocol for the detection of DMD and amelogenin gene is highly sensitive, specific and reliable, and can be used for PGD in those DMD families with DMD deletion mutation.