Novel pathogenic splicing variants in helicase for meiosis 1 (HFM1) are associated with diminished ovarian reserve and poor pregnancy outcomes.

PURPOSE: Diminished ovarian reserve (DOR) is associated with compromised fertility that affects approximately 10% of couples. Gene mutations are implicated in the pathogenesis of DOR. Here, we aimed to assess the clinical and genetic characteristics of two sisters with impaired fertility history. The two sisters showed DOR and suffered from recurrent pregnancy loss (RPL) in natural pregnancy and in vitro fertilization-embryo transfer (IVF-ET). METHODS: Whole exome sequencing (WES) was performed for the proband and pathogenic variants detected were validated by Sanger sequencing in all available family members. Minigene assay was performed to evaluate the impact of sequence variants on splicing effect. RESULTS: Two novel heterozygous variants on the HFM1 gene, c.1978-2A > C and c.2680 + 3_2680 + 4delAT, were observed in the two patients. The genotype of their parents was all heterozygous, while the unaffected sister and brother did not carry the variants. Both variants could produce alternative transcripts compared to wild-type counterparts, which might result in protein dysfunction. CONCLUSION: Our results demonstrated that the pathogenic splicing variants in HFM1 are associated with DOR in these two sisters. Mutations in HFM1 may contribute to RPL and poor IVF-ET outcomes because of descending quality and quantity of oocytes. The study enriched the genetic defect spectrum of DOR and understanding of the roles of HFM1 in female reproduction.

Abdominal ectopic pregnancy following a frozen embryo transfer cycle: a case report.

BACKGROUND: Abdominal ectopic pregnancy (AEP) is a rare form of ectopic pregnancy. As the number of in-vitro fertilization (IVF) procedures continues to increase, the incidence of AEP will also rise. However, the rarity and atypical presentation of AEP make early diagnosis challenging. CASE PRESENTATION: Herein, we report an AEP following frozen-thawed embryo transfer (FET) in an artificial cycle. The patient was misdiagnosed with implantation failure when the serum human chorionic gonadotropin (hCG) level was detected as 2.59mIU/ml at fourteenth day after embryo transfer. Therefore, she was suggested to stop luteal phase support. However, a ruptured AEP was developed 33 days following embryo transfer, which was diagnosed by laparoscopic surgery. CONCLUSIONS: The case highlighted the delayed serum ß-hCG and massive intraperitoneal hemorrhage may be clues to make early diagnosis of AEP. Clinicians must attach great importance to close monitoring and bear in mind the possibility of abdominal pregnancy.

Predictive value of anti-Müllerian hormone on pregnancy outcomes in in-vitro fertilization/intracytoplasmic single sperm injection patients at different ages.

PURPOSE: To investigate the predictive value of AMH level for pregnancy outcomes in different age groups of IVF/ICSI patients. METHODS: The study was a cohort study that included 11,484 patients that had their first IVF/ICSI procedure between 2016 and 2019. All patients who met the inclusion and exclusion criteria were divided into 6 groups according to 5-year age intervals, namely, Group 1: 20-24 years (n = 725); Group 2: 25-29 years (n = 4019); Group 3: 30-34 years (n = 3600); Group 4: 35-39 years (n = 1915); Group 5: 40-44 years (n = 1006); and Group 6: ≥ 45 years (n = 219). RESULTS: Receiver operating characteristic (ROC) curve analysis revealed that AMH level could only predict the outcome of live birth in Group 3 and Group 4 (p < 0.05). The area under the curve (AUC) of Group 3 was 0.536 (95% CI 0.510-0.561, p = 0.006), and that of Group 4 was 0.562 (95% CI 0.527-0.598, p = 0.001). The cutoff values of AMH for predicting live birth in Group 3 and Group 4 were 1.84 ng/ml and 1.86 ng/ml, respectively. Further logistic regression analysis showed that only the cutoff values of AMH and age could predict live birth in Groups 3 and 4. CONCLUSIONS: AMH level could predict live birth in IVF/ICSI patients at the age of 30-39. However, it could not be used to predict live birth in patients < 30 years or ≥ 40 years.

[Relationship between letrozole administration during the luteal phase after oocyte retrieval and the early-stage ovarian hyperstimulation syndrome oocurrence].

OBJECTIVE: To investigate the effect of letrozole in decreasing the early-stage ovarian hyperstimulation syndrome (OHSS) occurrence during the luteal phase for patients of OHSS high-risk after oocyte retrieval. METHODS: A total of 176 high-risk OHSS patients were randomly divided into two groups after oocyte retrieval. Patients in experiment group (n = 86) received 5 mg letrozole per day from the retrieval day and last for 5 days. Others in control group (n = 90) received placebo. The serum concentration of FSH, LH, estradiol (E2), progesterone (P) and vascular endothelial growth factor (VEGF) from the day of hCG injection to days after injection (5 days, 8 days, 10 days) were measured. And the incidence of moderate and severe OHSS was observed. RESULTS: The concentration of E2 on the indicated days (5 days, 8 days, 10 days after hCG injection) in experiment group and control group were (5 727±2 089) versus (11 826±4 281) pmol/L, (1 613±879) versus (7 925±3 507) pmol/L, (193± 90) versus (1 628±888) pmol/L; the concentration of VEGF on the indicated days in the two groups were (80±14) versus (108±19) ng/L, (66±11) versus (126±14) ng/L, (48±7) versus (148±14) ng/L; the concentration of E2 and VEGF were lower than those in control group (all P < 0.01). The FSH concentration in experiment group were (2.1±1.1) and (3.5±1.3) U/L on the day of fifth and eighth day after hCG injection, which were significantly higher than (0.7±0.3) and (0.7±0.4) U/L in control group (P < 0.05); the LH concentration in experiment group were (0.26±0.19) and (0.72±0.60) U/L on the day of fifth and eighth day after hCG injection, which were significantly higher than (0.11±0.03) and (0.14±0.08) U/L in control group (P < 0.05). The incidence of moderate and severe OHSS was signicantly decreased after letrozole treatment compared with control group [2% (2/86) versus 12% (11/90), P < 0.05]. CONCLUSION: Administration of 5 mg/d letrozole for 5 days during the luteal phase can reduce the E2 and VEGF levels for the high-risk OHSS patients who needed cryopreserve all embryos, and also reduce the occurrence of early OHSS.

Effect of anti-Müllerian hormone on early pregnancy loss in hormone replacement therapy-frozen-thawed embryo transfer cycles: An analysis including 6597 pregnant patients undergoing their first in vitro fertilization/intracytoplasmic sperm injection cycle.

OBJECTIVE: To investigate the relationship between anti-Müllerian hormone (AMH) level and early pregnancy loss in patients who underwent their first embryo transfer by hormone replacement therapy-frozen-thawed embryo transfer (HRT-FET) and analyze the threshold effect. METHODS: A retrospective cohort analysis was performed on pregnant women undergoing HRT-FET at the Reproductive Medical Center of Henan Provincial People's Hospital from January 2016 to December 2021. The patients were divided into four groups based on AMH concentration according to the Poseidon criteria: group A (≤1 µg/L), group B (1-≤2 µg/L), group C (2-≤6 µg/L), and group D (>6 µg/L). Univariate analysis, multivariate logistic regression analysis, smooth curve fitting, and threshold effect analysis were applied to investigate the influence of AMH on the outcome of early pregnancy loss in in vitro fertilization/intracytoplasmic sperm injection and HRT-FET cycles. RESULTS: Of the 6597 pregnant women, early pregnancy loss occurred in 893, giving an early pregnancy loss rate of 13.54%. Univariate regression analysis demonstrated that age, female body mass index, AMH, antral follicle count, endometrial thickness at endometrial transformation day, total retrieved oocyte number, number of pregnancies, duration of infertility, type of infertility, and the number of embryos transferred were all factors influencing the early pregnancy loss rate (P < 0.050). Multivariate logistic regression analysis, after adjusting for confounders, further stratified the analysis of patients of different ages. With group A as the control group, the results showed that when age was younger than 35 years, the pregnancy loss rates in groups B, C, and D were lower than that in group A, with statistical significance (P < 0.050); when age was 35 years or older, there was no statistically significant difference in outcome indicators between the groups (P > 0.050). A threshold effect analysis revealed that the AMH threshold was 2.83 µg/L. When the AMH concentration was less than 2.83 µg/L, the early pregnancy loss rate decreased significantly with increasing AMH concentration; the early pregnancy loss rate decreased by 21% for each unit increase in AMH (odds ratio 0.79; 95% confidence interval 0.71-0.88; P < 0.001); when the AMH concentration was 2.83 µg/L or more, there was no statistical difference in the change in early pregnancy loss rate (odds ratio 1.01; 95% confidence interval 0.99-1.03; P = 0.383). CONCLUSION: For pregnant women after their first embryo transfer, there is a curvilinear relationship between the influences of AMH levels on early pregnancy loss rates in patients younger than 35 years. When the AMH level was less than 2.83 µg/L, the early pregnancy loss rate declined significantly with increasing AMH levels.

Association with pre-pregnancy BMI on early pregnancy loss in infertility patients in first HRT-FET cycles: A retrospective cohort study.

OBJECTIVE: To investigate the association between pre-pregnancy body mass index (BMI) and the early pregnancy loss rate in patients in first hormone replacement therapy-frozen-thawed embryo transfer (HRT-FET) cycles and find the threshold. METHODS: A retrospective cohort study was conducted using a total of 14030 HRT-FET cycles at the Reproductive Center from January 2017 to December 2021. The association of pre-pregnancy BMI on early pregnancy loss rate in patients in HRT-FET cycles was assessed by performing univariate analysis, multivariable logistic regression, curve fitting and threshold effect analysis. RESULTS: There were 2076 cycles of early pregnancy loss, and the pregnancy loss rate was 14.80%. After adjusting for confounding factors, the early pregnancy loss rate of the obese group was significantly higher than that of the normal weight group (P < 0.05). The threshold effect analysis showed that as the pre-pregnancy BMI ranged from 21.2 to 25.8 kg/m2, the early pregnancy loss rate came to the plateau phase at the low level. In addition, when the BMI was ≥ 25.8 kg/m2, the early pregnancy loss rate increased by 3% (aOR = 1.03, P = 0.01) with each 1 kg/m2 increment of BMI. CONCLUSION: The early pregnancy loss rate might achieve a low level when the pre-pregnancy BMI was within the range of 21.2- 25.8 kg/m2. The early pregnancy loss rate would increase when pre-pregnancy BMI is more than 25.8 kg/m2. For patients in HRT-FET cycles, adjusting their pre-pregnancy BMI to the optimal level by following a healthy diet and daily exercise may help to reduce the early pregnancy loss.

Destination image, nostalgic feeling, flow experience and agritourism: An empirical study of Yunling Tea Estate in Anxi, China.

This study introduces destination image, nostalgic feeling, and flow experience into tea estate tourism and constructs a theoretical model that includes destination image, nostalgic feeling, flow experience, cultural identity, and tourists' behavioral intention. Then, an empirical study is conducted with tourists at Yunling Tea Estate in Anxi, China. The results show that all hypotheses are supported except the hypothesis pertaining to the significance of the influence of flow experience on behavioral intention, which is not supported. The model includes eight mediating effects and one moderating effect that is influenced by cultural memory.

Post-hoc evaluation of peripheral blood natural killer cell cytotoxicity in predicting the risk of recurrent pregnancy losses and repeated implantation failures.

Peripheral blood NK cytotoxicity assay (NKC) is one of the commonly utilized diagnostic tools for recurrent pregnancy losses (RPL) and repeated implantation failures (RIF). In this retrospective cohort study, we aimed to assess the cutoff values of NKC for RPL and RIF. A total of 883 women were included in this study; 24 nonpregnant fertile women, 604 nonpregnant women with three or more RPL, 163 nonpregnant women with two or more of RIF, 48 normal pregnant women, and 44 pregnant women with a history of RPL. Peripheral blood NKC assay was performed by flow cytometry. The differences between groups were analyzed using Student's t-test, a logistic regression analysis, and the area under the receiver operating characteristic curve analysis. Both nonpregnant fertile and normal pregnant women had significantly lower NKC at an effector to target cell ratio (E:T) of 50:1 (13.5 ± 1.1% and 12.9 ± 1.0%, respectively) when compared to women with RPL and RIF, and pregnant women with a history of RPL (23.6 ± 0.3%, 23.9 ± 0.5%, and 23.7 ± 1.0%, P < 0.0001 respectively). In addition, the area under the receiver operating characteristics curve for RPL and RIF using pre-conception NKC was 0.863 (P < 0.0001) and 0.879 (P < 0.0001), respectively, and for RPL using post-conception NKC was 0.736 (P = 0.001). These findings suggest that NKC significantly distinguishes nonpregnant women with RPL and RIF from fertile controls and pregnant RPLwomen from normal pregnant controls.

Does thyroid autoimmunity affect the reproductive outcome in women with thyroid autoimmunity undergoing assisted reproductive technology?

PROBLEM: Our study aims to investigate whether the anti-thyroperoxidase antibody (TPO-Ab) and TSH level in euthyroid women have any association with reproductive outcomes after the ART cycle. METHODS OF STUDY: A total of 1107 patients who were enrolled in the study were divided into four groups based on serum TSH level and TPO-Ab status: group A, 0.3 ≤ TSH < 2.5 mIU/L and TPO-Ab- ; group B, 0.3 ≤ TSH < 2.5 mIU/L and TPO-Ab+ ; group C, 2.5 ≤ TSH < 4.2 mIU/L, and TPO-Ab- ; and group D, 2.5 ≤ TSH < 4.2 mIU/L, TPO-Ab+ . The differences in ART cycles and pregnancy outcomes were analyzed between study groups. RESULTS: The fertilization rate in group D (73%) was significantly lower than that in groups A (83% P < .001), B (84% P = .001), and C (82% P = .002). The biochemical pregnancy rates of groups B (7%) and D (12%) were significantly higher than those of group A (2%) (P = .028 and P = .017, respectively). TPO-Ab was related to a higher biochemical pregnancy rate (P = .002, OR = 5.311, 95% CI 1.859-15.169) and TSH over 2.5 mIU/L was related to higher ICSI rate (P = .001, OR = 1.759, 95% CI 1.250-2.476) by logistic regression analysis. The receiver operating characteristic (ROC) also verified the results. CONCLUSION: The impacts of TSH ≥ 2.5 mIU/L on the intracytoplasmic sperm injection (ICSI) rate, TSH ≥ 2.5 mIU/L and TPO-Ab+ on the fertilization rate, and TPO-Ab+ on the biochemical pregnancy rate, rather than the effect on abortion, clinical pregnancy, and live birth, were emphasized.

Decreased HLA-C1 alleles in couples of KIR2DL2 positive women with recurrent pregnancy loss.

Specific killer cell immunoglobulin-like receptor (KIR) in women with recurrent pregnancy loss (RPL) and HLA ligands in couples invoke a susceptibility to RPL. However, the relationship between KIR2DL2 and its cognate ligand HLA-C1 has not been explored. In this prospective cohort study, 160 Caucasian women with RPL and 99 partners were included. KIR/HLA-C typing, NK assay, Th1/Th2 intracellular cytokine ratios, 25-(OH)-vitamin D level, and the presence of autoantibodies were analyzed. KIR2DL2 positive women (P = 0.023) and their partners (P = 0.017) had lower allele frequencies of HLA-C1 than those of KIR2DL2 negative women. KIR2DL2 positive women had significantly lower genotype frequency of HLA-C1C1 as compared to the North American Caucasian population controls (P < 0.05). In the partners of KIR2DL2 positive women, there was a substantially higher frequency of HLA-C2C2 than controls (P = 0.016). Besides, KIR2DL2 negative women had a higher prevalence of anti-ssDNA antibody as compared with that of KIR2DL2 positive women (P = 0.043). There were no differences in the distribution of HLA-C genotypes based on KIR2DL2, regardless of pregnancy outcome in women with RPL and their partners while on immunomodulation treatment. In conclusion, decreased ligands for inhibitory KIRs (inhKIR) could lead to insufficient inhibition of maternal uterine NK cells toward the trophoblast, thereby contributing to the pathogenesis of RPL. Specific KIR and HLA-C genotyping may predict the reproductive outcome of women with RPL.

PD-1 and PD-L1 expression on T-cell subsets in women with unexplained recurrent pregnancy losses.

PROBLEM: Does programmed death-1 (PD-1)/programmed death ligand-1 (PD-L1) expression on the T-cell subsets such as T helper (Th) 1, Th17, and Treg cells differentiate women with recurrent pregnancy losses (RPL) from normal fertile women? METHOD OF STUDY: The study was designed as a prospective cohort study. Forty-five women with two or more RPL of unknown etiology and twenty fertile women who had at least one or more live-born infants were enrolled prospectively from Jan 2017 to Jul 2019. PD-1 and PD-L1 expression on T-cell subsets were measured by flow cytometric analysis. RESULTS: The proportions of PD-1+ Th1 (CD4+ /IFN-γ+ /CD279+ and CD4+ /TNF-α+ /CD279+ ) and PD-1+ Th17 cells (CD4+ /IL17+ /CD279+ ) were significantly lower in RPL group than those of controls (P < .05, respectively). The proportion of PD-1+ Tregs (CD4+ /CD25+ /CD127dim/- /CD279+ ) in RPL group was not different from that of controls. The proportion of PD-L1+ Th17 cells (CD4+ IL17+ CD274+ ) was significantly lower as compared with that of /controls (P < .05). However, the proportions of PD-L1+ Th1 (CD4+ /IFN-γ+ /CD274+ and CD4+ /TNF-α+ /CD274+ ) and PD-L1+ Treg (CD4+ /CD25+ /CD127dim/- /CD274+ ) cells were not different between the RPL group and controls (P > .05, respectively). In Th1, Th17 and Treg cells, the proportions of PD-L1+ (CD274+ ) cells were significantly higher than those of PD-1+ (CD279+ ) cells in both RPL group and controls (P < .05, respectively). CONCLUSION: PD-1 and PD-L1 expressions on Th17 cells as well as PD-1 expression on Th1 cells were significantly downregulated in women with RPL, which may lead to increased Th1 and Th17 immunity, and imbalance between Th17, Th1, and Treg cells in women with RPL.

COVID-19 and immunomodulation treatment for women with reproductive failures.

COVID-19 pandemic is affecting various areas of health care, including human reproduction. Many women with reproductive failures, during the peri-implantation period and pregnancy, are on the immunotherapy using immune modulators and immunosuppressant due to underlying autoimmune diseases, cellular immune dysfunction, and rheumatic conditions. Many questions have been raised for women with immunotherapy during the COVID-19 pandemic, including infection susceptibility, how to manage women with an increased risk of and active COVID-19 infection. SARS-CoV-2 is a novel virus, and not enough information exists. Yet, we aim to review the data from previous coronavirus outbreaks and current COVID-19 and provide interim guidelines for immunotherapy in women with reproductive failures.

[Cryopreservation by cryoloop damages human immature oocytes].

OBJECTIVE: To examine the influence of cryoloop on the spindle and chromosome configurations of human oocytes cryopreserved in the germinal vesicle (GV) and metaphase II (M II) stages, as well as on the survival rate and potential for in vitro maturation (IVM). METHODS: GV oocytes were randomly assigned into a control group (matured in vitro into the M II stage), a GV cryopreserved group (cryopreserved in the GV stage and then matured in vitro), and an M II cryopreserved group (matured in vitro and cryopreserved in the M II stage). After cryopreservation and IVM, immunostaining of the tubulin and chromatin was performed followed by visualization using laser scanning confocal microscopy (LSCM). RESULTS: A significantly higher survival rate was observed in the GV cryopreserved group than in the M II , but the maturation rate showed no significant difference between the GV cryopreserved group and the control (P > 0.05). Compared with the control group, there was a statistically significant decrease in the rates of normal meiotic spindles and chromosomes in the GV cryopreserved group (P < 0.05). A significantly lower rate of normal spindles was noted in the M II cryopreserved group than in the control, but no statistical difference was shown in the rate of normal meiotic chromosomes between the two groups (P > 0.05). CONCLUSION: Cryopreservation by cryoloop has a damaging effect on the spindle and chromosome of human oocytes in the GV and M II stages.

Clinical outcomes of frozen-thawed single blastocyst transfer in patients requiring whole embryo freezing.

We compared clinical outcomes amongst frozen-thawed cleavage-stage embryo, double and single blastocyst transfers in patients requiring whole embryo freezing. Data of infertile patients undergoing in-vitro fertilization and embryo transfer (IVF-ET) in our Reproductive Medicine Center from January 2010 to December 2012 were retrospectively analyzed. According to patients' wishes, patients were divided into cleavage-stage embryo transfer groups (group A, n = 456), double blastocyst transfer group (group B, n = 106), and single blastocyst transfer group (group C, n = 402). We found that the number of frozen embryos was significantly less in groups B and C than in group A (all p < 0.05), but the implantation rate was significantly higher in groups B and C as compared to group A (all p < 0.05). The clinical pregnancy rate and pregnancy rate per included cycle were significantly higher in group B than in groups A and C (all p < 0.05). The multiple pregnancy rate was significantly lower in group C than in groups A and B (all p < 0.05). The rate of early abortion was significantly lower in group C as compared to group A (p < 0.05). The data support the view that it may be the best clinical strategy for patients who require whole embryo freezing and have four or more Day 3 embryos available, to incubate Day 3 embryos into blastocysts, which are then vitrified for elective single blastocyst transfer.

Effects of different doses of letrozole on the incidence of early-onset ovarian hyperstimulation syndrome after oocyte retrieval.

We explored the effects of different doses of letrozole on the incidence of ovarian hyperstimulation syndrome (OHSS) after oocyte retrieval during in vitro fertilization (IVF) in patients with high-risk OHSS. A total of 88 patients were randomly divided into a control group, and groups treated with 2.5 mg, 5 mg, or 7.5 mg of letrozole. We found that from the fifth day after human chorionic gonadotrophin (hCG) treatment, the E2 level decreased and there were statistical differences between the four groups (p < 0.05). From the eighth day after hCG treatment, the luteinizing hormone (LH) level increased, but the progesterone (P) level decreased. There were statistical differences between groups (p < 0.05). From the fifth day after hCG treatment, the level of vascular endothelial growth factor (VEGF) increased in the control, but decreased in the letrozole groups in a dose-dependent manner. There were statistically significant differences between groups (p < 0.001). The incidence of moderate and severe OHSS was lower in the 7.5 mg group than in the control group (p < 0.05). In the patients with high-risk OHSS undergoing whole embryo frozen transfer, treatment with 7.5 mg letrozole may be useful to limit OHSS.