Jihui Ai

Serum progesterone level effects on the outcome of in vitro fertilization in patients with different ovarian response: an analysis of more than 10,000 cycles

OBJECTIVE To investigate the relationship between serum P levels on the day of hCG administration and pregnancy outcomes in different responders undergoing IVF. DESIGN Retrospective study. SETTING Teaching hospital. PATIENT(S) A total of 11,055 women who underwent their first IVF/intracytoplasmic sperm injection cycles and a subgroup of 4,021 women undergoing frozen-embryo transfer (FET) cycles. INTERVENTION(S) Patients underwent IVF-ET with the long GnRH agonist protocol. The ovarian response was classified as high (≥ 20 oocytes; n = 2,023), poor (≤ 4 oocytes; n = 827), or intermediate (remaining cases; n = 8,205) according to the number of oocytes retrieved. Clinical outcomes of IVF-ET and FET cycles were analyzed according to plasma P levels. MAIN OUTCOME MEASURE(S) Ongoing pregnancy rates (PRs). RESULT(S) Ongoing PRs in fresh cycle were inversely associated with serum P levels on the day of hCG administration for all patients. Different P threshold concentrations were determined according to different ovarian response: We proposed a serum P level of 1.5 ng/mL as the threshold for poor responders, 1.75 ng/mL for intermediate responders, and 2.25 ng/mL for high responders. Our study does not show negative results for elevated P levels on oocyte performance in terms of fertilization, cleavage rate, or PR of FET cycles within different ovarian responses, offering no evidence for a detrimental effect of high P on oocyte quality. CONCLUSION(S) Elevated P levels on the day of hCG administration negatively influence PR regardless of different ovarian responses, although increased P threshold concentration is associated with better ovarian responses. The detrimental effect of P elevation on PR seems to be unrelated to oocyte quality in all responders.

Effect of Dehydroepiandrosterone Administration in Patients with Poor Ovarian Response According to the Bologna Criteria

Background Dehydroepiandrosterone (DHEA) is now widely used as an adjuvant to IVF treatment protocols in poor responders. However, clinical evidence for DHEA on improvement of ovarian response and IVF outcome is still limited, the validity of the results of the earlier studies, especially the varied inclusion criteria, is a subject of debate. Recently, the ESHRE Working Group developed a new definition, the Bologna criteria. The aim of the current study was to investigate the potential effect of DHEA treatment on in vitro fertilization (IVF) outcome of poor ovarian responders that fulfill the Bologna criteria. Methods This study investigated 386 poor ovarian responders that fulfill the Bologna criteria. Patients underwent IVF-ET treatment with the GnRH antagonist protocol. The study group contained 189 patients, who received 75 mg of DHEA daily (25 mg three times daily) before the IVF cycle. The control group was composed of 197 patients who received infertility treatment, but did not receive DHEA. The IVF outcome parameters in each group were compared. Results The study and control groups did not show statistically significant differences in terms of patient demographics characteristics, mean numbers of oocytes retrieved, mature oocytes, fertilization rate, cleavage rate, or embryo availability. While the DHEA group demonstrated significantly higher implantation rates (18.7% vs. 10.1%; P<0.01) and ongoing PRs (26.7% vs. 15.8%; P<0.05) as compared with the control. Conclusions DHEA pre-treatment does not significantly increase oocyte yield. However, the ongoing PRs in this subgroup of women are significantly higher after DHEA administration, suggesting that DHEA may increase IVF results by improving oocyte and embryo quality.

Administration of calcitonin promotes blastocyst implantation in mice by up-regulating integrin β3 expression in endometrial epithelial cells

STUDY QUESTION Does exogenous calcitonin improve the efficiency of implantation in mice by increasing uterine receptivity? SUMMARY ANSWER The administration of calcitonin could improve the efficiency of implantation by increasing the expression of several receptivity-related genes in endometrial epithelial cells (EECs). WHAT IS KNOWN ALREADY Calcitonin is one of the biomarkers of uterine receptivity, which is transiently produced in the uterine epithelia during the period of implantation both in humans and mouse. STUDY DESIGN, SIZE, DURATION Hormone-replaced mice were used for in vivo experiments. To evaluate the effect of calcitonin on uterine receptivity, the expression of endometrial genes was analyzed 36 h after i.p. injection of 0.5 IU calcitonin in a treatment group versus saline in the control. To evaluate the effect of calcitonin on implantation efficiency in vivo, two groups received 0.5 IU or 2 IU calcitonin (i.p.) 24 h before embryo transfer, and a control group received saline (i.p.) (n = 18 mice per group). Implantation sites were counted 7 days after embryo transfer. The RL95-2 human endometrial carcinoma cell line was used to study the mechanisms underlying the effect of calcitonin on gene expression in the endometria. Using an in vitro model of endometrium-trophoblast interaction, established with RL95-2 cells and JAR (human choriocarcinoma cell line) trophoblast, endometrial receptivity was evaluated by comparing attachment and outgrowth of JAR spheroids in control and treatment groups. PARTICIPANTS/MATERIALS, SETTING, METHODS Uterine receptivity in ovariectomized mice was induced by injection of estradiol and progesterone. Expression of eight genes in murine endometrium and RL95-2 cells was analyzed by real-time RT-PCR, western blot, immunohistochemical analysis, flow cytometry and enzyme-linked immunosorbent assay. We tested the effects of a protein kinase C inhibitor, matrigel and an antibody against integrin αvβ3 using RL95-2 cells and performed attachment and outgrowth assays using the in vitro model of endometrium-trophoblast interaction. Implantation efficiency was evaluated by counting the implantation sites after embryo transfer. MAIN RESULTS AND THE ROLE OF CHANCE Calcitonin up-regulated αvβ3 in RL95 cells, which in turn resulted in increased levels of the leukemia inhibitory factor (LIF) and heparin binding-epidermal growth factor (HB-EGF) mRNA (both P < 0.01 versus control) and protein (both P < 0.05 versus control). The attachment and expansion of JAR spheroids was promoted by pretreatment of EECs with calcitonin (P < 0.05 versus control) together with significantly increased expression of αvβ3, LIF and HB-EGF. Moreover, the injection of calcitonin in the preimplantation phase increased the total number of implantation sites in treatment groups (55 in control versus 78 and 85 in 0.5 and 2 IU groups, respectively). Compared with the control group (3.11 ± 2.14), the average number of implantation sites in the 2 IU calcitonin treatment group increased (4.72 ± 1.87, P = 0.022). LIMITATIONS, REASONS FOR CAUTION Experiments were performed in mice and human cell lines but not in primary cultures of human endometrial cells. WIDER IMPLICATIONS OF THE FINDINGS The findings presented here have important implications, in that calcitonin administration (currently used for treatment of hypercalcemia or osteoporosis) may have clinical benefits in assisted reproduction programs, by facilitating endometrial receptivity and embryo implantation. However, further studies are required to confirm these findings. STUDY FUNDING/COMPETING INTEREST(S) This work was supported by National Science Foundation of China (No. 81170619). There are no financial or commercial conflicts in this study.

Elevated Progesterone Levels on the Day of Oocyte Maturation May Affect Top Quality Embryo IVF Cycles.

In contrast to the impact of elevated progesterone on endometrial receptivity, the data on whether increased progesterone levels affects the quality of embryos is still limited. This study retrospectively enrolled 4,236 fresh in vitro fertilization (IVF) cycles and sought to determine whether increased progesterone is associated with adverse outcomes with regard to top quality embryos (TQE). The results showed that the TQE rate significantly correlated with progesterone levels on the day of human chorionic gonadotropin (hCG) trigger (P = 0.009). Multivariate linear regression analysis of factors related to the TQE rate, in conventional IVF cycles, showed that the TQE rate was negatively associated with progesterone concentration on the day of hCG (OR was -1.658, 95% CI: -2.806 to -0.510, P = 0.005). When the serum progesterone level was within the interval 2.0–2.5 ng/ml, the TQE rate was significantly lower (P <0.05) than when the progesterone level was < 1.0 ng/ml; similar results were obtained for serum progesterone levels >2.5 ng/ml. Then, we choose a progesterone level at 1.5ng/ml, 2.0 ng/ml and 2.5 ng/ml as cut-off points to verify this result. We found that the TQE rate was significantly different (P <0.05) between serum progesterone levels < 2.0 ng/ml and >2.0 ng/ml. In conclusion, the results of this study clearly demonstrated a negative effect of elevated progesterone levels on the day of hCG trigger, on TQE rate, regardless of the basal FSH, the total gonadotropin, the age of the woman, or the time of ovarian stimulation. These data demonstrate that elevated progesterone levels (>2.0 ng/ml) before oocyte maturation were consistently detrimental to the oocyte.

Blastocyst culture and cryopreservation to optimize clinical outcomes of warming cycles

Surplus embryos available for cryopreservation in fresh cycles are considered as having good potential for future use. However, the optimal stage of embryo cryopreservation remains unclear. In this study, 1190 patients with surplus embryos on day 3 were divided into two groups: cleavage-stage embryo cryopreservation (control group) and blastocyst cryopreservation (blastocyst group). The clinical outcomes of the subsequent warming cycles were evaluated. The proportion of cycles with blastocyst formation was 73.8% in the blastocyst group. Although in the blastocyst group, the cancellation rate of blastocyst transfer was increased due to lack of blastocysts available for cryopreservation, the blastocyst group achieved significantly higher rates of clinical pregnancy/cycle (43.2% versus 34.9%; P=0.003), pregnancy/transfer (59.5% versus 35.4%; P<0.001) and implantation (46.5% versus 22.2%; P<0.001) from the first warming cycle compared with the control group. In an embryo-number classified analysis, the clinical pregnancy rate was also higher in the blastocyst group. However, the cumulative pregnancy was similar between the two groups. Blastocyst culture as an embryo selection tool will not improve embryo viability but it will help patients to achieve pregnancy more quickly. Extended culture of surplus embryos to the blastocyst stage for cryopreservation optimizes the clinical outcomes.

Correlation between physical status of human papilloma virus and cervical carcinogenesis

The prevalence of human papilloma virus (HPV)-16 in patients with cervical cancer, the physical status of HPV-16 in patients with cervical lesions, and the role of HPV-16 integration in cervical carcinogenesis were investigated. HPV genotyping was performed by using PCR approach with the primer GP5+/GP6+ and type-specific primer on biopsy specimens taken operatively from 198 women. Multiple PCR was done to detect physical status of HPV-16 in a series of cervical liquid-based cytology samples and biopsy specimens obtained from different cervical lesions with HPV-16 infection, including 112 specimens with cervical cancer, 151 specimens with CIN I, 246 specimens with CIN and 120 specimens with CINIII. The results showed that there were 112 cervical cancer samples (56.57% of total cervical cancer patients) with HPV-16 infection. The frequency of HPV-16 pure integration was 65.18% (73/112), 56.57% (47/120), 23.58% (58/246) and 7.95% (12/151) in cervical cancer, CINIII, CINII and CINI patients respectively. In situ hybridization was performed on some paraffin-embedded sections of CINII, CINIII and cervical cancer to verify the physical status of HPV-16 infection. Significant difference was observed between cervical cancer and CIN I, CINII, CINIII in the frequency of HPV-16 integration (P<0.01). It is suggested that HPV-16 is the most prevalent type and is associated with cervical cancer. In the case of HPV-16 infection there are close associations between the severity of cervical lesions and the frequency of HPV-16 integration. The application of testing HPV genotyping and physical status based on detection of HC-II HPV DNA would be in favor of predicting the prognosis of cervical precancerosis and enhancing the screening accuracy of cervical cancer.SummaryThe prevalence of human papilloma virus (HPV)-16 in patients with cervical cancer, the physical status of HPV-16 in patients with cervical lesions, and the role of HPV-16 integration in cervical carcinogenesis were investigated. HPV genotyping was performed by using PCR approach with the primer GP5+/GP6+ and type-specific primer on biopsy specimens taken operatively from 198 women. Multiple PCR was done to detect physical status of HPV-16 in a series of cervical liquid-based cytology samples and biopsy specimens obtained from different cervical lesions with HPV-16 infection, including 112 specimens with cervical cancer, 151 specimens with CIN I, 246 specimens with CIN and 120 specimens with CINIII. The results showed that there were 112 cervical cancer samples (56.57% of total cervical cancer patients) with HPV-16 infection. The frequency of HPV-16 pure integration was 65.18% (73/112), 56.57% (47/120), 23.58% (58/246) and 7.95% (12/151) in cervical cancer, CINIII, CINII and CINI patients respectively. In situ hybridization was performed on some paraffin-embedded sections of CINII, CINIII and cervical cancer to verify the physical status of HPV-16 infection. Significant difference was observed between cervical cancer and CIN I, CINII, CINIII in the frequency of HPV-16 integration (P<0.01). It is suggested that HPV-16 is the most prevalent type and is associated with cervical cancer. In the case of HPV-16 infection there are close associations between the severity of cervical lesions and the frequency of HPV-16 integration. The application of testing HPV genotyping and physical status based on detection of HC-II HPV DNA would be in favor of predicting the prognosis of cervical precancerosis and enhancing the screening accuracy of cervical cancer.

The artificial cycle method improves the pregnancy outcome in frozen–thawed embryo transfer: a retrospective cohort study

Abstract The aim of this retrospective cohort study was to investigate which preparation method is optimal for frozen–thawed embryo transfer (FET) treatment. Analyses were performed on 3160 FET cycles, including 654 cycles with a natural cycle (NC) protocol and 2506 cycles with an artificial cycle (AC) protocol. The primary outcome measures were the clinical pregnancy rate (CPR) and the live birth rate (LBR) per transfer. The Student’s t-test, chi-square test and multiple logistic regression were used for statistical analysis. The CPR per transfer was 49.4% in the NC group and 58.6% in the AC group (OR = 1.270, 95% CI: 1.037–1.554). The LBR per transfer was 42.2% and 50.8% in the NC and AC groups, respectively (OR = 1.269, 95% CI: 1.037–1.552). Dividing the patients according to the type of transferred embryos, the CPR (67.3% versus 57.0%, p < 0.01) and LBR (58.8% versus 49.7%, p < 0.01) were higher after the AC protocol than after NC protocol in patients with blastocyst transfer. The NC and AC protocols yielded comparable CPR and LBR in the patients with cleavage embryo transfer. Our data indicate better pregnancy outcomes after the AC protocol than after the NC protocol. The AC protocol should be recommended in patients who were counseled before receiving FET treatment. Further studies are needed to confirm this finding. Chinese abstract 这项回顾性队列研究的目的是探讨冷冻解冻胚胎移植（frozen-thawed embryo transfer，FET）的最佳准备方法。对3160例FET周期进行分析，其中自然周期（natural cycle，NC）方案组654例，人工周期（artificial cycle，AC）方案组2506例。以每个周期的临床妊娠率（clinical pregnancy rate，CPR）和活产率（live birth rate，LBR）作为评价指标。使用t检验、卡方检验和多重logistic回归进行统计学分析。NC组每周期的CPR为49.4%，AC组每周期的CPR为58.6%（OR=1.270，95% CI：1.037-1,554）。NC组和AC组每周期的LBR分别为42.2%和50.8%（OR=1.269,95% CI: 1.037-1.552）。将患者按照移植的胚胎类型来分，其中行囊胚移植的患者，AC方案的CPR（67.3% Vs 57.0, p<0.01）和LBR(58.8% Vs 49.7%, p<0.01)均高于NC方案。而行卵裂期胚胎移植的患者，NC方案和AC方案具有相似的CPR和LBR。我们的数据表明AC方案的妊娠结局优于NC方案。对于接受FET治疗的患者推荐使用AC方案。这一结论需要进一步的研究来证实。

Association of controlled ovarian hyperstimulation treatment with down-regulation of key regulators involved in embryonic implantation in mice

The debate exists whether or not gonadotropin-releasing hormone (GnRH) analogs used in controlled ovarian hyperstimulation (COH) impair endometrial receptivity. Homeobox A11 (Hoxa11), Meis homeobox 1 (Meis1), cadherin 1 (Cdh1), and catenin beta 1 (Ctnnb1) are well known to be involved in successful implantation. In this study, the endometrial expression of Hoxa11, Meis1, Cdh1, and Ctnnb1 during the peri-implantation period was investigated in an in vitro fertilization (IVF) mouse model by real-time RT-PCR and Western blot to evaluate the relationship between Hoxa11, Meis1, Cdh1, and Ctnnb1 expression and the impact of the COH on endometrial receptivity. The mimic COH protocols included GnRH agonist plus human menopausal gonadotropin (HMG) (GnRH agonist group), GnRH antagonist plus HMG (GnRH antagonist group), and HMG alone (HMG group). The expression levels of Hoxa11, Meis1, Cdh1, and Ctnnb1 mRNA and protein were decreased in all of the COH groups. The expression levels of Hoxa11 and Ctnnb1 were the lowest in the GnRH agonist group, and those of Meis1 and Cdh1 were lower in the GnRH analog groups than the HMG group. There were positive correlations between the expression of Hoxa11 and Ctnnb1, as well as the expression of Meis1 and Cdh1 among all the groups. In conclusion, the COH protocols, particularly with GnRH analogs, suppressed Hoxa11, Meis1, Ctnnb1 and Cdh1 expression, in mouse endometrium during the peri-implantation period. Our data reveal a novel molecular mechanism by which the COH protocols might impair endometrial receptivity.SummaryThe debate exists whether or not gonadotropin-releasing hormone (GnRH) analogs used in controlled ovarian hyperstimulation (COH) impair endometrial receptivity. Homeobox A11 (Hoxa11), Meis homeobox 1 (Meis1), cadherin 1 (Cdh1), and catenin beta 1 (Ctnnb1) are well known to be involved in successful implantation. In this study, the endometrial expression of Hoxa11, Meis1, Cdh1, and Ctnnb1 during the peri-implantation period was investigated in an in vitro fertilization (IVF) mouse model by real-time RT-PCR and Western blot to evaluate the relationship between Hoxa11, Meis1, Cdh1, and Ctnnb1 expression and the impact of the COH on endometrial receptivity. The mimic COH protocols included GnRH agonist plus human menopausal gonadotropin (HMG) (GnRH agonist group), GnRH antagonist plus HMG (GnRH antagonist group), and HMG alone (HMG group). The expression levels of Hoxa11, Meis1, Cdh1, and Ctnnb1 mRNA and protein were decreased in all of the COH groups. The expression levels of Hoxa11 and Ctnnb1 were the lowest in the GnRH agonist group, and those of Meis1 and Cdh1 were lower in the GnRH analog groups than the HMG group. There were positive correlations between the expression of Hoxa11 and Ctnnb1, as well as the expression of Meis1 and Cdh1 among all the groups. In conclusion, the COH protocols, particularly with GnRH analogs, suppressed Hoxa11, Meis1, Ctnnb1 and Cdh1 expression, in mouse endometrium during the peri-implantation period. Our data reveal a novel molecular mechanism by which the COH protocols might impair endometrial receptivity.

Fertility outcome analysis after modified laparoscopic microsurgical tubal anastomosis

Modified laparoscopic microsurgical tubal anastomosis is an alternative for microsurgical anastomosis via laparotomy to reverse sterilization in women with renewed child wish. The current study aims to evaluate the fertility outcome after modified laparoscopic microsurgical tubal anastomosis. A retrospective study was performed. Fifty-eight women who underwent modified laparoscopic microsurgical tubal anastomosis were monitored to investigate the fertility outcome and characteristics of this new technology. Of the 58 patients, the cumulative pregnancy rate (PR) in the 42 patients with follow-up data was 23.8% (10/42), 57.1% (24/42), 66.7% (28/42), and 73.8% (31/42) within 6, 12, 24, and 36 months after surgery, respectively. The intrauterine PR was 69.0% (29/42). Two patients (4.8%) had ectopic pregnancies that occurred within 24 months of surgery; three cases ended in spontaneous abortion. The delivery rate was 83.9% (26/31). The length of operating time was 1.2±0.3 h, with a range of 1.0–2.5 h (60–145 min), and the mean time was approximately 75 min. The blood loss was relatively small, between 10 and 50 ml with an average amount of 22 ml. Thus, the modified laparoscopic tubal anastomosis is a highly successful procedure and a viable alternative to open abdominal microsurgical approaches. Compared with the traditional laparoscopic tubal sterilization reversal, this modified approach has three advantages: (1) less invasive approach via a trocar reduction; (2) remodeling of tube is better performing tied together after 3–4 sutures; and (3) faster operating time.

Propensity score-matched study and meta-analysis of cumulative outcomes of day 2/3 versus day 5/6 embryo transfers

The superiority of the cumulative outcomes of day 5/6 embryo transfer to those of day 2/3 embryo transfer in infertile couples has been debated. This retrospective study included data collected from 1051 patients from July 2011 to June 2014. Multiple maternal baseline covariates were subjected to propensity score matching analysis, and each day 5/6 group woman was matched to one day 2/3 group woman. A systematic meta-analysis was conducted to validate the results. After matching was completed, 217 patients on the day 2/3 group were matched with those on the day 5/6 group, and no significant differences in the baseline characteristics were observed between the two groups. The cumulative pregnancy rate (57.14% vs. 53.46%, OR 1.16, 95% CI 0.79–1.70) and cumulative live birth rate (53.00% vs. 49.77%, OR 1.14, 95% CI 0.78–1.66) of day 5/6 embryo transfers were higher than those of day 2/3 embryo transfers, but this difference was not significant. The mean cycles per live birth and mean days per live birth in the day 5/6 group were significantly lower than those in the day 2/3 group. This study demonstrated that day 5/6 embryo transfer is a more cost-effective and time-efficient policy than day 2/3 embryo transfer to produce a live baby.