miR-188-3p-targeted regulation of ATG7 affects cell autophagy in patients with nonobstructive azoospermia.

BACKGROUND: Nonobstructive azoospermia (NOA) is one of the most difficult forms of male infertility to treat, and its pathogenesis is still unclear. miRNAs can regulate autophagy by affecting their target gene expression. Our previous study found that miR-188-3p expression in NOA patients was low. There are potential binding sites between the autophagy gene ATG7 and miR-188-3p. This study aimed to verify the binding site between miR-188-3p and ATG7 and whether miR-188-3p affects autophagy and participates in NOA by regulating ATG7 to influence the autophagy marker genes LC3 and Beclin-1. METHODS: Testicular tissue from 16 NOA patients and 16 patients with normal spermatogenesis and 5 cases in each group of pathological sections were collected. High-throughput sequencing was performed to detect mRNA expression differences. Quantitative real-time polymerase chain reaction (qRT-PCR), Western blotting, immunohistochemical staining and immunofluorescence were used to detect protein localization and expression. Autophagosome changes were detected by electron microscopy. The targeting relationship between miR-188-3p and ATG7 was confirmed by a luciferase assay. RESULTS: ATG7 protein was localized in the cytoplasm of spermatogenic cells at all levels, and the ATG7 gene (p = 0.019) and protein (p = 0.000) were more highly expressed in the NOA group. ATG7 expression after overexpression/inhibition of miR-188-3p was significantly lower (p = 0.029)/higher (p = 0.021) than in the control group. After overexpression of miR-188-3p, the ATG7 3'UTR-WT luciferase activity was impeded (p = 0.004), while the ATG7 3'UTR-MUT luciferase activity showed no significant difference (p = 0.46). LC3 (p = 0.023) and Beclin-1 (p = 0.041) expression in the NOA group was significantly higher. LC3 and Beclin-1 gene expression after miR-188-3p overexpression/inhibition was significantly lower (p = 0.010 and 0.024, respectively) and higher (p = 0.024 and 0.049, respectively). LC3 punctate aggregation in the cytoplasm decreased after overexpression of miR-188-3p, while the LC3 punctate aggregation in the miR-188-3p inhibitor group was higher. The number of autophagosomes in the miR-188-3p mimic group was lower than the number of autophagosomes in the mimic NC group. CONCLUSIONS: LC3 and Beclin-1 were more highly expressed in NOA testes and negatively correlated with the expression of miR-188-3p, suggesting that miR-188-3p may be involved in the process of autophagy in NOA. miR-188-3p may regulate its target gene ATG7 to participate in autophagy anDual luciferase experiment d affect the development of NOA.

Abnormal BMI in Male and/or Female Partners Are Deleterious for Embryonic Development and Pregnancy Outcome During ART Process: A Retrospective Study.

Objective: To investigate the effect of BMI in male and/or female partners on embryo development and clinical pregnancy outcome during ART. Methods: Data of 11,130 cycles between January 2018 and December 2020 were retrospectively analyzed. They were divided into Group A, B, C, and D based on couples' BMI values, also were divided into Group I, II, III and IV in IVF cycles and Group i, ii, iii, and iv in ICSI cycles. After grouping, inter-group indicators comparisons and logistic regression analysis were performed. Results: In IVF cycles, CPR in Group I and Group III were higher than Group IV. In Group III, it was higher than Group II. The AR in Group IV was higher, but the LBR was lower than Group I, Group II, and Group III. Logistic regression analysis results suggested that AR in Group IV was higher than that in Group I in IVF cycles, whereas LBR was lower.In ICSI cycles, high-quality embryo rate in Group i and Group ii were both higher than that in Group iii and Group iv. The CPR in Group i was higher than Group ii and Group iv, and in Group iii was higher than Group ii and Group iv. The AR in Group i was lower than Group iii and Group iv, and AR in Group ii was lower than Group iv. LBR, in Group I it was higher than Group ii, Group iii, and Group iv. Logistic regression analysis results suggested CPR in Group ii was significantly lower than that in Group i. AR in Group iii was considerably higher than that in Group i. LBR in Group ii and Group iv were significantly lower than that in Group i. Conclusion: Female higher BMI was not conducive to the formation of high-quality embryos in ICSI cycle. Female and/or male BMI affected AR and LBR more than CPR not only in IVF cycles, but also in ICSI cycles.

Reproductive Outcomes of Single Embryo Transfer in Women with Previous Cesarean Section.

The aim of this study is to evaluate the reproductive outcomes of patients with previous cesarean section (CS) undergoing single-embryo transfer (SET). A total of 5479 patients previously underwent CS or vaginal delivery (VD) were included. The patients with previous CS included single/double cleavage (SCT/DCT), single/double blastocyst stage embryo transfer (SBT/DBT). The comparison of reproductive outcomes between CS and VD, SET, and double-embryo transfer of CS was conducted. The main outcome measures included clinical pregnancy rate (CPR), multiple pregnancy rate (MPR), live birth rate (LBR), and cumulative live birth rate (CLBR). There were no differences in CPR and LBR between patients previously underwent CS and VD, and the preterm birth rate of multiple births in patients underwent CS were significantly higher than underwent VD. As for patients underwent CS, the CPR, MPR, and LBR were lower in patients treated with SCT than DCT in IVF/ICSI cycles. The CPR and LBR of patients treated with SCT were not statistically different compared with DCT and SBT when patients' age were younger than 35 years in FET cycles, and the MPR of patients treated with SCT was lower. The CPR and LBR of patients treated with SCT were lower than DCT/SBT when patients' age were 35 years or older in FET cycles. The CPR, LBR, and CLBR of patients treated with SBT were not lower than DCT in IVF/ICSI-ET, FET, and complete cycles, but the MPR was lower. Reducing the number of embryos transferred should be considered seriously for CS. SCT under the 35 years of FET cycles is feasible. SBT is an effective strategy to reduce MPR of IVF/ICSI, FET, and complete cycles regardless of overall or age stratification.

Follicle-stimulating hormone may predict sperm retrieval rate and guide surgical approach in patients with non-obstructive azoospermia.

Testicular sperm aspiration- (TESA) or micro-dissection testicular sperm extraction- (MD-TESE) combined intracytoplasmic sperm injection (ICSI) was the only option for non-obstructive azoospermia (NOA) patients to have a biological offspring and they had different success rates in sperm retrieval. Our study aimed to find predictor(s) for predicting the sperm retrieval rate (SRR) in NOAs and guide clinicians in choosing different surgical approaches, TESA or MD-TESE for NOAs. 294 NOAs who had undergone TESA or MD-TESE were divided into TESA group and MD-TESE group. Depending on sperm retrieval, each group was divided into two subgroups: successful subgroups and failure subgroups. They respectively were 24 cases and 131 cases, 53 cases and 86 cases. Clinical data, including body mass index (BMI), testicular volume, and serum hormone levels, were analyzed in a retrospective manner. The results showed that follicle-stimulating hormone (FSH) and luteinizing hormone (LH) levels and SRR were lower in TESA group as compared to these in MD-TESE group, while testicular volume was higher (P < 0.05). The surgical approach of sperm retrieval significantly affected the SRR (P < 0.05). In TESA subgroups, testicular volume, FSH and LH differed significantly (P < 0.05). In MD-TESE subgroups, the level of FSH and LH differed significantly between both groups (P < 0.05). Using logistics regression, we found a negative correlation (ß=-0.083) between FSH and the SRR in TESA group but a positive correlation (ß = 0.064) in MD-TESE group (P < 0.05). In conclusion, serum FSH level can predict the SRR of NOAs and guide the clinicians while selecting the suitable surgery approach for NOAs.

Factors predicting clinical pregnancy rate of in vitro fertilization-embryo transfer (a STROBE-compliant article).

The aim of this study was to investigate the factors predicting clinical pregnancy rate of in vitro fertilization-embryo transfer (IVF-ET).The data of 9960 patients receiving IVF-ET fresh cycle at our Reproductive Center from January 2009 to December 2017 were first divided into pregnant group and non-pregnant group to find the clinical pregnancy rate-related factors. According to the serum HCG levels at 36 hours and 12 hours after HCG trigger, all patients were divided into 4 groups including <50 mIU/ml, ≥50 and <100 mIU/ml, ≥100 and <200 mIU/ml, and ≥200 mIU/ml groups to know whether the HCG levels at 36 hours and 12 hours affect the pregnancy rate. According to the serum HCG ratio at 36 hours to 12 hours (36 h/12 h) after HCG trigger, all patients were divided into three groups including <0.88, 0.88-1.06 and >1.06 groups to observe whether the serum HCG ratio (36 h/12 h) affects the clinical pregnancy rate. According to different assisted pregnancy modes, all patients were divided into 3 groups including IVF, ICSI, and IVF/ICSI groups to observe whether the assisted pregnancy mode affects the clinical pregnancy rate. The correlation of the clinical pregnancy rate with pregnancy rate-related factors obtained above was analyzed using logistic regression analysis model.The clinical pregnancy rate significantly increased (P < .01) in the HCG ratio (36 h/12 h) >1.06 group as compared with the HCG ratio (36 h/12 h) < 0.88 and 0.88-1.06 groups. The serum estrogen (E2) level at 36 hours was significantly lower and the number of retrieved oocytes was significantly higher in the HCG ratio (36 h/12 h) >1.06 group than in the HCG ratio (36 h/12 h) <0.88 and 0.88-1.06 groups (P = .000).The serum HCG ratio (36 h/12 h) may be used as a predictor of IVF-ET clinical pregnancy rate. High clinical pregnancy rate is probably associated with E2 down-regulation in the HCG ratio (36 h/12 h) >1.06 group.

Clinical analysis of spontaneous pregnancy reduction in the patients with multiple pregnancies undergoing in vitro fertilization/intracytoplasmic sperm injection-embryo transfer.

OBJECTIVE: To investigate the spontaneous pregnancy reduction (SPR) rate, SPR-related factors and the effects of SPR on pregnancy outcomes in the patients with multiple pregnancies undergoing in vitro fertilization/intracytoplasmic sperm injection-embryo transfer (IVF/ICSI-ET). METHODS: Between January 1998 and December 2010, 3957 patients undergoing fresh/frozen-thawed cycles (IVF/ICSI-ET) and their 5106 neonates were enrolled in this study. According to spontaneous pregnancy reduction (SPR), this study included singleton originating from twins [(2→1) group] or from triplets [(3→1) group], and twins originating from triplets [(3→2) group]. According to SPR time, this study included ≤8 week, 8-18 week and ≥18 week's groups. Outcome measures were SPR rate, preterm rate, mean birth weight and the rates of low birth weight and very low birth weight. RESULTS: SPR rate was higher in triplets group than in twins group, in frozen-thawed cycles than in fresh cycles, in the patients ≥35 years than in the patients <35 years (all P<0.05). Compared with ≤8 week group, preterm rate was significantly increased in 8-18 week group (P<0.05). Pregnancy outcomes were better in (2→1) group than in twins group, in (3→1) group than in triplets group (all P<0.05). After multi-fetal pregnancy reduction (MFPR), the mean birth weight was higher and low birth weight was lower in SPR group than in only MFPR group (all P<0.05). CONCLUSION: SPR rate is related to age and the initial number of gestational sacs. Both SPR and MFPR can improve pregnancy outcomes. The later the SPR occurs, the worse the neonatal outcomes are. Due to the possibility of SPR, it is necessary to appropriately delay MFPR until 8 gestational weeks.

Effects of the distance between small intramural uterine fibroids and the endometrium on the pregnancy outcomes of in vitro fertilization-embryo transfer.

AIM: To explore the effects of the distance between small intramural uterine fibroids (≤4 cm) and the endometrium on the outcomes of in vitro fertilization-embryo transfer (IVF-ET). METHODS: We prospectively analyzed pregnancy outcomes in 117 infertile women with small intramural uterine fibroids and 117 infertile women without uterine fibroids who all underwent IVF-ET. The size and number of small intramural uterine fibroids and the shortest distance between the small intramural uterine fibroids and the endometrium were measured by transvaginal three-dimensional ultrasound. The endometrial and subendometrial blood flow parameters, implantation rate, clinical pregnancy rate, abortion rate and live birth rate were compared between the women with and without small uterine fibroids and among the different shortest distances (≤1, 1-3 and >3 mm). The effects of the size and number of small intramural uterine fibroids on IVF-ET outcomes were observed. RESULTS: The endometrial volume on ET day, the implantation rate and the live birth rate were significantly lower, but the abortion rate was significantly higher, in the women with small intramural uterine fibroids than in those without uterine fibroids (p < 0.05). The endometrial flow index was higher in the shortest distance ≤1-mm group than in the groups with 1-3 and >3 mm, and the implantation rate was higher in ≤1-mm group than in the >3-mm group (p < 0.05). There were no significant differences in clinical outcomes between different sizes and numbers of small intramural uterine fibroids. CONCLUSION: Small intramural uterine fibroids can affect IVF-ET outcomes. Compared with other shortest distances (1-3 and >3 mm), the shortest distance of ≤1 mm has a higher implantation rate.

An research on the isolation methods of frozen-thawed human ovarian preantral follicles.

OBJECTIVE: To explore the effective isolation method for preantral follicles from human frozen-thawed ovarian tissue. METHODS: The ovarian cortical tissue was frozen by direct cover vitrification (DCV). The frozen-thawed ovarian tissue was used for isolation of preantral follicles with collagenase combined with mechanical method and mechanical method alone, respectively. RESULTS: 1. There was no statistical difference in the survival rates of follicles in various stages between before and after freezing (P > 0.05). 2. The survival rate of secondary follicles was higher, but the survival rate of primordial follicles was lower in mechanical method alone than in collagenase combined with mechanical method (all P < 0.05). 3. The diameters of follicles were larger and E2 levels were higher in mecha-nical method alone than that in collagenase combined with mechanical method (all P < 0.05). CONCLUSION: After the frozen-thawed ovarian tissue was cultured for 6 days, compared with collagenase combined with mechanical method, mechanical method alone can obtain higher survival rate of secondary follicles, greater follicular diameter and higher E2 level, which are conducive to follicular subsequent development.

Comparative study of pregnancy outcomes between day 3 embryo transfer and day 5 blastocyst transfer in patients with progesterone elevation.

OBJECTIVE: To explore the effects of progesterone (P) elevation on pregnancy outcomes of day 3 embryo and day 5 blastocyst transfer. METHODS: Clinical outcomes (pregnancy and ectopic pregnancy rates) following day 3 embryo and day 5 blastocyst transfer cycles were retrospectively analysed. Day 3 embryo and day 5 blastocyst transfer cycles were divided into normal P level (P ≤ 1.5 ng/ml) and P elevation group (P > 1.5 ng/ml), based on the serum P level on the day of human chorionic gonadotropin (hCG) administration. RESULTS: A total of 2868 cycles were analysed. In day 3 embryo transfer cycles (n = 2345), the clinical pregnancy rate was significantly higher in the normal P level group compared with the P elevation group (55.4% versus 46.7%, respectively) and the ectopic pregnancy rate was significantly lower in the normal P level group compared with the P elevation group (2.8% versus 7.9%, respectively). In day 5 blastocyst transfer cycles (n = 523), there were no significant differences in the clinical pregnancy and ectopic pregnancy rates between the two groups, based on the P level. CONCLUSION: These preliminary findings suggest that day 5 blastocyst transfer should be adopted for patients with P elevation on the day of hCG administration.

Follow-up of children born after intracytoplasmic sperm injection with epididymal and testicular spermatozoa.

BACKGROUND: To evaluate the safety of intracytoplasmic sperm injection (ICSI) with epididymal or testicular sperm, this study compared children born after ICSI treatment with epididymal or testicular sperm with children conceived after ICSI with ejaculated sperm. METHODS: This retrospective study included 317 children born after ICSI with percutaneous epididymal sperm aspiration (PESA), 103 children born after ICSI with testicular sperm aspiration (TESA), and a control group of 1008 children born after ICSI with ejaculated sperm. All of the patients received their assisted reproductive treatment in the Reproductive Medicine Center of the First Affiliated Hospital of Zhengzhou University from January 2004 to December 2011. Data, such as the rate of stillbirths, perinatal mortality, gestational age, birth weight, and the rate of congenital malformations of the three groups, were compared. RESULTS: PESA and TESA children were not different from ICSI children in the rate of stillbirths, perinatal mortality, infant mortality rate, gestational age, the rate of prematurity, and the rate of malformations (P > 0.05). A slight increase in birth defects was reported in the TESA group compared with those in the control group, but there was no significant difference between the groups (P > 0.05). CONCLUSION: ICSI with epididymal or testicular sperm does not lead to more stillbirths or congenital malformations compared with ICSI using ejaculated sperm.

Ectopic pregnancy in frozen-thawed embryo transfer: a retrospective analysis of 4,034 cycles and related factors.

We analyzed the incidence of ectopic pregnancy in frozen-thawed embryo transfer (FET) as a function of natural and hormone replacement cycles and ectopic pregnancy-related factors. In this study, there were 4,034 FET cycles performed in our center between January 2005 and December 2010, and the rates of ectopic pregnancy were compared between natural and hormone replacement cycles. The analysis of ectopic pregnancy-related factors in FET was performed with 1:4 age-matched chi-square tests. The rate of ectopic pregnancy was lower in natural FET cycles (1.46%) than in hormone replacement FET cycles (3.31%) with a statistical significance (P < 0.05). Many factors were associated with ectopic pregnancy in FET, but only treatment protocols were considered as a controllable factor. We conclude that the incidence of ectopic pregnancy is significantly lower in natural FET cycles than in hormone replacement FET cycles. The application of exogenous sex hormones in assisted reproductive cycles may be an important factor to cause ectopic pregnancy in FET. This suggests that care should be taken when selecting the treatment protocol in order to avoid ectopic pregnancy.

Molecular karyotype single nucleotide polymorphism analysis of early fetal demise.

We explored the application of single nucleotide polymorphism microarray (SNP array) in molecular karyotype analysis for early spontaneous abortion detection in assisted reproductive technology (ART). SNP array was performed in 81 cases. Of the 81 cases, 16 experienced natural conception (NC) and 65 were pregnant by ART. Of the 65 cases, 4 underwent artificial insemination (AI), 32 fresh in vitro fertilization-embryo transfer (IVF-ET), 9 fresh intracytoplasmic sperm injection (ICSI), and 20 thawed embryo transfer. In the 81 cases examined 69.1% displayed an abnormal molecular karyotype. In the subjects greater than 35 years of age, the abnormal molecular karyotype rate was 87.5% higher compared to 61.4% in younger individuals (P < 0.05). There was no significant difference in the abnormal molecular karyotype rate or type between ART (64.6%) and NC (87.5%). Compared with traditional cytogenetic diagnosis, the SNP array can identify a greater number of abnormal karyotypes.

[Study on the incidence and influences on ectopic pregnancy from embryo transfer of fresh cycles and frozen-thawed cycles].

OBJECTIVE: To investigate ectopic pregnancy from embryo transfer (ET)of in-vitro fertilization (IVF) cycle and intracytoplasmic sperm injection (ICSI) cycle and frozen-thawed (FET) cycle. METHODS: From Jan.2005 to Dec. 2010, a total of 9037 IVF-ET or ICSI-ET cycles and 4034 FET cycles were performed in our reproductive medicine center, Affiliated First Hospital of Zhengzhou University. The incidence of ectopic pregnancy rate was studied in fresh cycles IVF-ET (5998) and ICSI-ET (3039) cycles, and natural FET (2198) and hormone replacement (E-P) FET (1836) cycles. RESULTS: Of 4034 FET cycles, 1090 clinical pregnancies and 26 ectopic pregnancies were observed, the incidence of ectopic pregnancy was 2.38% (26/1090). Of 9037 fresh cycles, 3602 cycles were clinical pregnancy, and 133 cycles were ectopic pregnancy, and the incidence of ectopic pregnancy was 3.69% (133/3602). The ectopic pregnancy rate in FET cycles was lower than in fresh cycles significantly (P < 0.05). Of 3039 fresh ICSI-ET cycles, the incidence of ectopic pregnancy was 2.62% (34/1298) in 1298 clinical pregnancies. Of 5998 IVF-ET cycles, 2304 clinical pregnancies were observed, the incidence of ectopic pregnancy was 4.30% (99/2304). Ectopic pregnancy rate in the fresh ICSI-ET cycles was lower than that of IVF-ET group significantly (P < 0.01). The ectopic pregnancy rate in the natural FET cycles was 1.46% (8/547), which was significantly lower than 3.31% (18/543) in E-P group (P < 0.05). CONCLUSIONS: The incidence of ectopic pregnancy of FET cycles was significantly lower than that of fresh embryo transfer cycles. The application of exogenous sex hormones in assisted reproductive cycles might increase occurrence of ectopic pregnancy.

Effect of coincubation time of sperm-oocytes on fertilization, embryonic development, and subsequent pregnancy outcome.

Several studies have reported improved IVF by shortening the time of sperm-oocyte coincubation from 16-18 hours to 1-4 hours. The objective of this study was to examine the advantages and disadvantages of a shortened sperm-oocyte coincubation time in order to assess the effects of this insemination method for clinical IVF practice. Two insemination methods, the shortened method (4 hours) and the standard method (16-18 hours) of coincubation of sperm-oocytes for two groups of patients based on the quality of sperm were compared. Group I, was composed of couples without male factor; Group II, involved couples with mild male factor. Fertilization, good quality embryos, clinical pregnancy, and implantation rates were compared by two different insemination methods. In Group I, fertilization, clinical pregnancy, and implantation rates were not different between the two insemination methods. However, the polyspermy rate was significantly higher (P < 0.05) in the shortened (7.3%) than in the standard (4.1%) insemination method. In Group II, the fertilization rate was significantly lower (P < 0.05) using the shortened insemination method (62.6%) compared to the standard insemination method (68.7%). When fertilization failed with the shortened insemination method, the clinical pregnancy and implantation rates were 34.7% and 24.1%, respectively, from the rescue intracytoplasmic sperm injection (ICSI). The live birth rate from the rescue ICSI was 32.0% with normal infants. The duration of sperm-oocyte coincubation does not affect fertilization, embryo quality, clinical pregnancy, and implantation rates. However, fertilization rates will decrease with the shortened insemination method when the sperm parameters are poor. From the results of the present study we suggest that the combination of the shortened sperm-oocyte coincubation and rescue ICSI method may be an efficient method for IVF treatment in order to prevent fertilization failure when sperm parameters were poor as mild male factor.

Comparative study on the pregnancy outcomes of in vitro fertilization-embryo transfer between long-acting gonadotropin-releasing hormone agonist combined with transvaginal ultrasound-guided cyst aspiration and long-acting gonadotropin-releasing hormone agonist alone.

OBJECTIVE: To retrospectively analyze the effects of long-acting gonadotropin-releasing hormone agonist (GnRH-a) combined with transvaginal ultrasound-guided cyst aspiration on the pregnancy outcomes of in vitro fertilization-embryo transfer (IVF-ET) in the infertile patients with ovarian endometriosis. METHODS: The 134 patients with ovarian endometriosis who underwent GnRH-a combined with transvaginal ultrasound-guided cyst aspiration and IVF-ET were served as experimental group. The 102 patients with ovarian endometriosis who underwent GnRH-a and IVF-ET were served as control group. After treatment, the cyst size, follicle stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E2), number of ovum pick-up, fertility rate, cleavage rate, high-quality embryo rate, implantation rate, clinical pregnancy rate and abortion rate were compared between the two groups. RESULTS: In the 134 patients of experimental group, 138 cysts disappeared after GnRH-a combined with three times of transvaginal ultrasound-guided cyst aspiration. In the 102 patients of control group, of the 114 cysts, 34 disappeared after GnRH-a, 67 were decreased and 13 were unchanged. The abortion rate was significantly lower in experimental group than in control group. The level of serum E2 on HCG day, the number of ovarian follicles with 14 mm or more, the number of retrieved oocytes, high-quality embryo rate, implantation rate and clinical pregnancy rate were higher in experimental group than in control group (all P<0.05). CONCLUSION: GnRH-a combined with transvaginal ultrasound-guided cyst aspiration can obtain better therapeutic effects and pregnancy outcomes in infertile patients with ovarian endometriosis who underwent IVF-ET.

Effect of modified single cell fixation method on cell-nuclear areas and fluorescence in-situ hybridization signals.

BACKGROUND: Fluorescence in situ hybridization (FISH) is an irreplaceable method in pre-implantation genetic diagnosis. We explored the effects of a modified single cell fixation method on the cell-nuclear area and FISH signal. METHODS: From January 2006 to March 2008, the blastomeres with marked nuclei from D3 embryos were selected. Cells were fixed with three different methods. The effects of the three methods on the cell-nuclear areas and FISH signals were then analyzed. RESULTS: The cell fixation rate was higher in conventional (Group B, 94.85%) and modified (Group C, 95.79%) Tween-20/HCl + methanol/glacial acetic acid methods than in the methanol/glacial acetic acid method (Group A, 86.73%) with p < 0.05. The complete signal rates in group A, B, and C were 95.3%, 93.5%, and 93.4%, respectively, with p > 0.05. The mean cell-nuclear areas in groups A, B, and C were 55.3, 46.2, and 49.5 microm3, respectively, with p < 0.05 in group A compared with group B or C, but with p > 0.05 between Group B and C. There was no significant difference in signal overlap and splitting rates between the three groups. CONCLUSIONS: Modified Tween-20/HCl + methanol/glacial acetic acid method fails to increase FISH signal overlap and splitting rates. It is simple and its fixation time is short. It can be widely used in clinical practice.

Relationship between processed total motile sperm count of husband or donor semen and pregnancy outcome following intrauterine insemination.

We retrospectively analyzed 6,360 artificial insemination cycles of husband's semen through intrauterine insemination (AIH-IUI) or artificial insemination with donor semen through intrauterine insemination (AID-IUI) in patients with infertility between August, 1998 and August, 2010. The relationship between processed total motile sperm count (PTMS) and pregnancy outcome was determined. The study was divided into 6 groups according to PTMS. Group 1: ≤ 2.0 million, Group 2: 2.1-4.0 million, Group 3: 4.1-6.0 million, Group 4: 6.1-8.0 million, Group 5: 8.1-10.0 million, and Group 6: >10.0 million. There was no statistically significant difference in age, duration of infertility, unilateral tubal patency, induced ovulation, and single IUI or double IUI between the 6 groups in both AIH-IUI and AID-IUI. The total clinical pregnancy rate of AIH-IUI was 10.81 % and AID-IUI was 27.52 %. Among the 6 groups, the clinical pregnancy rate was the lowest in Group 1 (P < 0.05) in both AIH-IUI and AID-IUI. With the increased PTMS, the clinical pregnancy rate of IUI was improved. However, a statistical difference between groups was only observed for Group 1. When PTMS is ≤ 2 × 10(6) the clinical pregnancy rate of IUI is significantly decreased. In this case in vitro fertilization (IVF) should be adopted.

Clinical outcome of emergency egg vitrification for women when sperm extraction from the testicular tissues of the male partner is not successful.

The development of an effective oocyte cryopreservation system will have a significant impact on the clinical practice of reproductive medicine. However, the important option of emergency oocyte cryopreservation has yet to be well documented. In this report, we review the cases of 15 women with male partners who were diagnosed with nonobstructive azoospermia and for whom testicular sperm extraction on the day of oocyte retrieval failed. Emergency oocyte vitrification was performed and after two months, the vitrified oocytes were warmed and the surviving oocytes inseminated with frozen-thawed donor sperm by intracytoplasmic sperm injection (ICSI). A total of 117 mature oocytes from the 15 women were vitrified and warmed. The post-warming survival rate was 84.6% (99/117), and the fertilization rate following ICSI was 83.8% (83/99). We selected 30 embryos for transfer to 15 patients, 8 of whom became pregnant. The clinical pregnancy rate was 53.3% (8/15) and the implantation rate was 30.0% (9/30). Nine healthy live births resulted from 8 pregnancies. These results indicate that emergency oocyte vitrification is an effective rescue technique that can be applied clinically with acceptable pregnancy and live birth rates when testicular sperm extraction from the male partner failed on the day of oocyte retrieval. These results also highlight another important option for oocyte cryopreservation through the use of vitrification technology.

[Effects of cryopreservation time and thawing method of human oocyte vitrification on the outcome of assisted reproduction].

OBJECTIVE: To evaluate the effects on pregnancy outcome of freezing time from oocyte retrieval and thawing method for metaphaseII human oocytes vitrification. METHODS: From Mar 2007 to Mar 2009, the clinical outcome of 30 infertile women undergoing vitrified-thawing oocytes of in vitro fertilization-embryo transfer (IVF-ET) in the Reproductive Medical Center of the First Affiliated Hospital of Zhengzhou University was studied retrospectively, including 21 women with double fallopian tube obstruction and 9 women's husband azoospermia. All infertile women were divided into three groups, including 5 cases in group A (freezing between 4 and 5 hours from oocyte retrieval and conventional thawing method), 9 cases in group B (freezing within 2 hours from retrieval and conventional thawing method) and 16 cases in group C (freezing within 2 hours from retrieval and improved thawing method). The vitrified oocytes were preserved for 2 months to 1 year and thawed for Intracytoplasmic sperm injection (ICSI) and embryo transfer. The outcome of IVF and pregnancy were recorded. RESULTS: (1) The rates of oocyte survival was (65 ± 33)% in group B and (72 ± 23)% in group C and the rate of transfer cycle was 9/9 in group B and 16/16 in group C, which were all significantly higher than (16 ± 17)% of oocyte survival and 1/5 of transfer cycle in group A (P = 0.001, 0.021). However, the rate of oocyte survival and transfer cycle between group B and group C did not reach statistical difference (P > 0.05). The rate of implantation and clinical pregnancy of (33 ± 38)% and 9/16 in group C were significantly higher (4 ± 11)% and 1/9 in group B (P = 0.033, 0.040). (2) The mean age of women in group C were (28.6 ± 2.1) in oneself oocyte, (28.0 ± 4.6) in donor oocyte and (28.1 ± 3.4) in donor sperm. The rate of oocyte survival was (73 ± 25)%, (88 ± 10)% and (66 ± 25)%. The rate of fertilization rate was (84.6 ± 0.9)%, (79.3 ± 2.0)% and (82.8 ± 15.0)%. The rate of implantation was (20.0 ± 44.7)%, (33.0 ± 0.1)%, (41.6 ± 41.7)%. The rate of clinical pregnancy was 1/5 in oneself cycles, 3/3 in donor oocyte cycles, 5/8 banked donor sperm cycles in group C. All above clinical parameters were not statistically different (P > 0.05). (3) In group A, one women underwent IVF-ET and no clinical pregnancy was observed. One women pregnancy was terminated at two months in group B. The clinical pregnancies rate of group C was 9/16, late abortion occurred in 1 woman, the other 8 women underwent term pregnancy, including 5 male infants and 4 female infants. All of infants showed normal Karyotype. Live-birth rates per warmed oocyte was 5.9%(8/135). The mean gestational weeks and birth weight of the infants were (39.4 ± 0.9) weeks and (3574 ± 569) g, respectively. CONCLUSIONS: Embryo quality and clinical outcome of thawing cycles could be significantly improved when oocyte vitrification was performed within 2 hours from oocyte retrieval and improved thawing method.