Evaluation of Ovarian Reserve Tests and Age in the Prediction of Poor Ovarian Response to Controlled Ovarian Stimulation-A Real-World Data Analysis of 89,002 Patients.

Aims: This study aimed to explore the value of ovarian reserve tests (ORTs) for predicting poor ovary response (POR) and whether an age cutoff could improve this forecasting, so as to facilitate clinical decision-making for women undergoing in vitro fertilization (IVF). Methods: A retrospective cohort study was conducted on poor ovary response (POR) patients using real-world data from five reproductive centers of university-affiliated hospitals or large academic hospitals in China. A total of 89,002 women with infertility undergoing their first traditional ovarian stimulation cycle for in vitro fertilization from January 2013 to December 2019 were included. The receiver operating characteristic (ROC) curve was performed to estimate the prediction value of POR by the following ORTs: anti-Mullerian hormone (AMH), antral follicle count (AFC), basal FSH (bFSH), as well as patient age. Results: In this retrospective cohort, the frequency of POR in the first IVF cycle was 14.8%. Age, AFC, AMH, and bFSH were used as predicting factors for POR, of which AMH and AFC were the best indicators when using a single factor for prediction (AUC 0.862 and 0.842, respectively). The predictive values of the multivariate model included age and AMH (AUC 0.865), age and AFC (AUC 0.850), age and all three ORTs (AUC 0.873). Compared with using a single factor alone, the combinations of ORTs and female age can increase the predictive value of POR. Adding age to single AMH model improved the prediction accuracy compared with AMH alone (AUC 0.865 vs. 0.862), but the improvement was not significant. The AFC with age model significantly improved the prediction accuracy of the single AFC model (AUC 0.846 vs. 0.837). To reach 90% specificity for POR prediction, the cutoff point for age was 38 years old with a sensitivity of 40.7%, 5 for AFC with a sensitivity of 55.9%, and 1.18 ng/ml for AMH with a sensitivity of 63.3%. Conclusion: AFC and AMH demonstrated a high accuracy when using ROC regression to predict POR. When testing is reliable, AMH can be used alone to forecast POR. When AFC is used as a prediction parameter, age is suggested to be considered as well. Based on the results of the cutoff threshold analysis, AFC ≤ 5 and AMH ≤ 1.18 ng/ml should be recommended to predict POR more accurately in IVF/ICSI patients.

[Study on the factors associated with clinical pregnancy rate of in-vitro fertilization in endometriosis related infertility].

OBJECTIVE: To evaluate the factors associated with clinical pregnancy rate of in-vitro fertilization (IVF) in endometriosis related infertility. METHODS: Total of 326 patients with endometriosis related infertility undergoing IVF between January 2007 and December 2011 were studied in Department of Reproductive Medicine, First Affiliated Hospital, Nanjing Medical University, retrospectively, which were divided into 141 cases in clinical pregnancy group and 185 cases in non-pregnancy group. Those factors including age, body mass index (BMI), basic FSH, antral follicle count (AFC), CA125 and CA199, endometriotic stage and history of surgery, stimulation scheme were analyzed by bivariate analysis and multivariable logistic regression. RESULTS: (1) Pregnancy rate:total of 141 pregnant cases and 185 non-pregnant cases treated by IVF were observed, pregnancy rate was 43.2% (141/326). (2) Basic parameters: there was no statistical difference in age, BMI, basic FSH, AFC, CA125 and CA199 between clinical pregnancy group and non-pregnancy group (P > 0.05). (3) Bivariate analysis: clinical pregnancy rate of 50.0% (87/174) among patients with infertility year less than five years was significantly higher than 35.5% (54/152) in patients with more than five years. Pregnancy rate of 56.1% (46/82) in stage I-II was significantly higher than 42.5% (79/186) in stage III-IV. Pregnancy rate of 46.6% (125/268) with history of surgery was significantly higher than 27.6% (16/58) with no history of surgery (P < 0.05). Pregnancy rate of 48.2% (79/164) in long-term scheme was higher than 38.3% (62/162) in short-term scheme, but there was no significant difference (P = 0.075). (4) Multivariable logistic regression: clinical pregnancy rate of infertility year with less than 5 years, stage I-II, history of surgery proved stage I-II and stage III-IV was significantly higher compared with infertility year more than 5 years, stage III-IV and no history of surgery respectively (adjusted OR and 95%CI: 2.003, 1.263 - 3.175; 1.899, 1.110 - 3.248; 3.769, 1.802 - 7.887, P < 0.05). CONCLUSION: Factors affecting clinical pregnancy rate of IVF in endometriosis related infertility were infertility year, stage and surgery.

Oocyte vitrification technology has made egg-sharing donation easier in China.

When infertile women undergoing IVF or intracytoplasmic sperm injection (ICSI) have more than 20 mature oocytes retrieved, at least 15 oocytes are inseminated by their husband's spermatozoa. The extra oocytes are cryopreserved by vitrification. If the patients became pregnant and have healthy live births, the patients are encouraged to donate their remaining cryopreserved oocytes. Forty-seven egg-sharing donors were recruited after having normal deliveries and they donated their remaining oocytes, totalling 395 cryopreserved oocytes, to 75 recipients. The survival rate of vitrified-warmed oocytes was 83.0%. Following insemination by ICSI, the fertilization and cleavage rates were 83.8% and 89.8%, respectively. Out of 75 recipients, 71 recipients completed the treatment cycles and 30 of them became pregnant with clinical pregnancy and implantation rates of 42.3% and 25.5%, respectively. The birthweight of the new-born infants (22 from singleton and two from one set of twins) were 3344.5 ± 669.1g and 2425.0 ± 742.5 g, respectively. No birth defects were observed for the live births. These results indicate that oocyte vitrification is an effective methodology for an egg-sharing donation programme, with acceptable pregnancy and implantation rates.

[Sperm chromatin structure assay predicts the outcome of intrauterine insemination].

OBJECTIVE: Sperm chromatin structure assay (SCSA), as a clinically practical technique for the analysis of DNA damage, is rarely reported in China. This study focuses on the correlation of DNA damage with the pregnancy rate of intrauterine insemination (IUI). METHODS: We performed semen analysis for 482 couples undergoing IUI, calculated the DNA fragmentation index (DFI) by SCSA, and observed the relationship between DFI and the pregnancy rate of IUI. RESULTS: Clinical pregnancy was achieved in 5 (5.26%) of the 95 cases with DFI > 25%, and in 59 (15.25%) of the 387 cases with DFI < or = 25%. Those with sperm DFI >25% had significantly lower rates of biochemical pregnancy and clinical pregnancy than those with DFI < or = 25% (OR: 0.37, 95% CI: 0.14 - 0.96 and OR: 0.38, 95% CI: 0.16 - 0.97). No significant differences were found in the DFI of 54 cases between the first and the second cycle ([15.05 +/- 7.98]% vs [17.25 +/- 12.18]%, P > 0.05). Sperm DFI was significantly negatively correlated with sperm concentration, sperm motility and total progressively motile sperm count (P < 0.01). CONCLUSION: The pregnancy rate of IUI is significantly lower in couples with DFI >25% than in those with DFI < or = 25%. Sperm DFI obtained from SCSA is partly correlated with sperm concentration and motility, and it is a robust predictor of the IUI outcome.

[Y-chromosome microdeletions do not affect the outcomes of ICSI for infertile males].

OBJECTIVE: To compare the outcomes of intracytoplasmic sperm injection (ICSI) for infertile males with Y-chromosome microdeletions and for those with azoospermia or severe oligospermia but without Y-chromosome microdeletions. METHODS: We retrospectively analyzed 56 cycles of ICSI for 48 infertile cases with Y microdeletions (Group A) and 94 cycles for 90 cases with azoospermia or severe oligospermia but without Y-chromosome microdeletions (Group B) during the same period. We compared the two groups in the females' age, duration of infertility, males' age, number of oocytes retrieved, number of ICSI oocytes, fertilization rate, good embryo rate, number of embryos transferred, implantation rate, clinical pregnancy rate, abortion rate, live birth rate and babies' sexes. RESULTS: There were no significant differences between Groups A and B in the females' age, duration of infertility, males' age, number of oocytes retrieved, number of ICSI oocytes and number of embryos transferred (P > 0.05), nor in the rates of fertilization (69.0% vs 73.2%), good embryos (53.3% vs 48.7%), implantation (24.0% vs 30.3%), biochemical pregnancy (41.1% vs 44.7%), clinical pregnancy (37.5% vs 35.1%), early abortion (4.8% vs 6.1%) and live birth (35.7% vs 29.2%) (P > 0.05). CONCLUSION: Y-chromosome microdeletions do not affect the outcomes of ICSI. The affected couples should be informed of the necessity of prenatal genetic diagnosis before embryo implantation and the inevitability of vertical transmission to male offspring.

Effect of estradiol on proliferation and differentiation of side population stem/progenitor cells from murine endometrium.

BACKGROUND: In our previous study, endometrium side population cells (SP cells) were isolated from postpartum murine uterus, and characterized by a heterogeneous population of stem/progenitor cells. In this study, we investigated the effect of estrogen on the proliferation and differentiation of SP cells. METHODS: SP and non-SP cells of postpartum murine endometrium were isolated by DNA dye Hoechst 33342. The expression of estrogen receptor 1 (ESR1) was measured by reverse transcription polymerase chain reaction (RT-PCR), Real-time PCR, Western blot, immunofluorescence and immunohistochemistry. The proliferation and differentiation of SP cells treated with different concentrations [10(-8) M-10(-6) M] of estradiol (E2) and E2+ ICI182780 (Faslodex, inhibitor of ESR1) were measured by 3-(4, 5-dimethylthiazoly1-2)-2,5-diphenyltetrazolium bromide(MTT) and clonogenic assays. RESULTS: (1) SP cells expressed ESR1 at a higher level than non-SP cells. (2) The level of E2 in the serum and the expression of ESR1 in the uterus of postpartum murine changed in the same manner with the ratio of SP cells to total uterus cells at a different postpartum time point. ESR1, as ABCG2 is also predominantly located in the stroma and the glandular epithelium of the uterus. (3) 10(-6) M E2 notably promoted the proliferation of SP cells after treatment for 24 h. This effect could be inhibited by ICI182780. E2 at the concentration of 10(-7) M or 10(-8) M was sent to impair the large cloning efficiency (CE) of SP cells. CONCLUSIONS: The effect of estrogen on the proliferation and differentiation of endometrium SP cells via ESR1 was observed and it was in a concentration dependent fashion. Clearly, more work is needed to understand the in vivo effect of E2 at the physiological concentration on the differentiation of SP cells.

The role of ezrin-associated protein network in human sperm capacitation.

Membrane modifications in sperm cells represent a key step in sperm capacitation; however, the molecular basis of these modifications is not fully understood. Ezrin is the best-studied member of the ezrin/radixin/merlin family. As a cross-linker between the cortical cytoskeleton and plasma membrane proteins, ezrin contributes to remodeling of the membrane surface structure. Furthermore, activated ezrin and the Rho dissociation inhibitor, RhoGDI, promote the formation of cortical cytoskeleton-polymerized actin through Rho activation. Thus, ezrin, actin, RhoGDI, Rho and plasma membrane proteins form a complicated network in vivo, which contributes to the assembly of the structure of the membrane surface. Previously, we showed that ezrin and RhoGDI1 are expressed in human testes. Thus, we sought to determine whether the ezrin-RhoGDI1-actin-membrane protein network has a role in human sperm capacitation. Our results by Western blot indicate that ezrin is activated by phosphorylation of the threonine567 residue during capacitation. Co-immunoprecipitation studies revealed that, during sperm capacitation, the interaction between ezrin and RhoGDI1 increases, and phosphostaining of two dimensional electrophoresis gels showed that RhoGDI1 is phosphorylated, suggesting that RhoGDI1 dissociates from RhoA and leads to actin polymerization on the sperm head. We speculate that activated ezrin interacts with polymerized actin and the glycosylated membrane protein cd44 after capacitation. Blocking sperm capacitation using ezrin- or actin-specific monoclonal antibodies decreases their acrosome reaction (AR) rate, but has no effect on the AR alone. Taken together, our results show that a network consisting of ezrin, RhoGDI1, RhoA, F-actin and membrane proteins functions to influence the modifications that occur on the membrane of the sperm head during human sperm capacitation.

Isolation and characterization of side population cells in the postpartum murine endometrium.

Endometrium is a highly active organ that is periodically remodeled during the life span. Previous studies have indicated the presence of an adult stem or progenitor cell population in this tissue. In this study, side population (SP) cells were isolated from the endometrium of postpartum murine uterus but not from the endometrium of a uterus undergoing a normal estrus cycle. Phenotype analysis showed that SP cells were negative for hematopoietic, endothelial, and mesenchymal stem cell (MSC) markers, but they expressed stem cell antigen 1 (Sca-1) and c-kit at various degrees. Side population cell is a heterogeneous population of endometrial stem/progenitor cells that have colony-forming capacity. They were found to reside in quiescence in the stroma but not in the luminal epithelium. These data suggest that, like other tissues and organs, the murine endometrium also contains SP cells. Their specific role in the regeneration of the endometrium warrants further study.

AF-2364 is a prospective spermicide candidate.

AbstractInhibition of sperm motility has recently become a promising target for male contraceptive development. AF-2364, an analogue of Lonidamine (LND), had a contraceptive effect when orally administered to adult Sprague-Dawley rats. LND can also target mitochondria to inhibit oxygen consumption and block energy metabolism in tumour cells. However, there are no reports of the effects of AF-2364 on human sperm function. Herein we describe the action of AF-2364 on human sperm in vitro, as well as the mechanisms involved. AF-2364 specifically blocked human sperm motility in vitro. Further experiments revealed that AF-2364 can target sperm mitochondrial permeability transition (MPT) pores to induce the loss of sperm mitochondrial membrane potential (DeltaPsim) and decrease ATP generation; however, no significant changes in the cytoskeletal network or the human sperm proteome were detected after exposure to AF-2364. Incubation of AF-2364 with other human or mouse cell lines indicated that the spermicidal effect at the lower concentration was specific. In summary, the spermicidal effect of AF-2364 involves direct action on sperm MPT pores, and this compound should be further investigated as a new spermicide candidate.

[Correlation between serum progesterone level at the day with human chorionic gonadotrophin administration and the outcome of pregnancy in in-vitro fertilization].

OBJECTIVE: To investigate the relationship between serum progesterone level at the day with human chorionic gonadotrophin (hCG) administration and pregnant outcome from in in-vitro fertilization-embryo transfer (IVF-ET). METHODS: From Mar. 2002 to Apr. 2007, 786 cycles with serum progesterone measurement on the day of hCG administration for final oocyte maturation in IVF were analyzed retrospectively in Reproductive Medicine Center in First Affiliated Hospital of Nanjing Medical University. All stimulations were down-regulated with gronadotrophin release hormone agonist (GnRH-a) in both long protocols and short protocols before gonadotrophin stimulation. When the thresholds of serum progesterone were set at 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5 and 9.0 nmol/L, respectively. If the level of progesterone was less than the thresholds, those patients were in lower progesterone group, on the contrary, more than the threshold value, those patients were in higher progesterone group. The laboratory results and the clinical outcomes between all patients at lower and higher progesterone group at different thresholds value were analyzed. RESULTS: The rate of normal fertilization, quality embryos, successful implantation, chemical pregnancy, clinical pregnancy and live birth did not exhibit remarkable difference between patients with higher and lower serum progesterone level at multiple thresholds on the day of hCG administration in the 786 cycles (P > 0.05). However, when the thresholds of serum progesterone were at 8.5 and 9.0 nmol/L, early abortion rates of 27.3% (3/11) and 3/7 in higher progesterone group were significantly higher than 8.8% (26/297) and 8.6% (26/301) in lower progesterone group (P < 0.05). And the total abortion rates of 3/7 in higher progesterone group were significantly higher than 11.0% (34/301) in lower progesterone group when the thresholds of serum progesterone were 9.0 nmol/L (P < 0.05). CONCLUSIONS: This study did not prove the correlationship between progesterone level at the day with hCG administration and the probability of clinical pregnancy or live birth. However, early abortion rates or the total abortion rates were associated with higher progesterone level when the thresholds of serum progesterone were at 8.5 nmol/L or 9.0 nmol/L.

[Clinical value of artificial insemination by donor].

OBJECTIVE: To investigate the clinical value of artificial insemination by donor (AID). METHODS: We retrospectively analyzed 480 cycles of AID among 258 infertile couples, who were divided according to the women's age into a < or = 30 yr group and a > or = 31 yr group. RESULTS: A total of 120 pregnancies were achieved in 480 AID cycles, with a cycle pregnancy rate of 25.00% and a cumulative pregnancy rate of 46.51%. In the natural cycles, the cycle pregnancy rate was 29.65% and the cumulative pregnancy rate was 51.00% in the < or = 30 yr group, significantly higher than 13.33% and 25.00% in the > or = 31 yr group (P < 0.05). In the ovulation induction cycles, no significant differences were found in the cycle and cumulative pregnancy rates between the two groups (24.02 and 48.86% versus 23.81 and 43.48% , P > 0.05). The cycle and cumulative pregnancy rates decreased with the increase of infertility duration and the women's age, but had no significant differences. In the first four cycles, the cycle pregnancy rates were 24.03, 24.94, 24.69 and 25.00% (P > 0.05), and the cumulative pregnancy rates were 24.03, 39.53, 45.74 and 46.51%, with significant differences between the first cycle and the other three (P < 0.01). CONCLUSION: Ovulation induction is superior to natural cycle in AID for older women. IVF/ICSI can be resorted to only after AID has failed three or four times.

[Clinical effects of oocyte cryopreservation in assisted reproduction technology].

OBJECTIVE: To investigate the clinical effects of oocyte cryopreservation in assisted reproduction technology (ART). METHODS: 258 patients undergoing retrieval of more than 20 oocytes during in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) were divided into 2 groups:Group A, undergoing surplus oocytes cryopreservation (84 cycles) and Group B undergoing embryo cryopreservation (174 cycles) according to the patients' choices. Fertilization rate and clinical pregnancy rate of fresh embryo transfer cycle were compared between these two groups. Twenty-three infertile couples' frozen oocytes were thawed for further ART treatment. Among them, fifteen couples received embryo transfer using their own frozen-thawed oocytes, and other four couples used donated frozen-thawed oocytes. The survival rate, fertilization rate, cleavage rate, implantation rate, and clinical pregnancy rate of these 19 cycles were compared to the outcome of 56 frozen-thawed embryo transfer cycles. RESULTS: The fertilization rate of Group A who underwent IVF was 65.9%, not significantly different from that of Group B who received IVF (66.9%, P > 0.05), and the fertilization rate of Group A who underwent ICSI was 71.6%, not significantly different from that of Group B who received ICSI (64.1%, P > 0.05). The clinical pregnancy rate (per embryo transfer cycle) of Group A who received IVF was 52.9%, not significantly different from that of Group B who received IVF (42.3%, P > 0.05), and the clinical pregnancy rate (per embryo transfer cycle) of Group A who received ICSI was 35.5%, not significantly different from that of Group B who received ICSI (34.4%, P > 0.05). The clinical pregnancy rate of frozen-thawed oocyte group (per embryo transfer cycle) was 47.4% (9/19). Four couples used donated frozen-thawed oocytes, two of them got clinical pregnancy and one of them had term delivery. CONCLUSION: For women who undergo retrieval of more than 20 oocytes in IVF/ICSI, the clinical outcome of fresh embryo transfer cycle, such as fertilization rate and clinical pregnancy rate, are not influenced by oocyte cryopreservation and embryo cryopreservation. There is no significant difference in the clinical pregnancy rate (per embryo transfer cycle) between frozen-thawed oocyte group and frozen-thawed embryo group. Compared with embryo cryopreservation, oocyte cryopreservation has obvious advantages in fertility preservation and oocyte donation.

[Influence of nuclear factor-kappaB decoy oligonucleotides on RANTES expression and monocyte chemotactic activity in stromal cells of ectopic endometrium].

OBJECTIVE: To study the inhibitory effect on the expression of regulated upon activation, normal T cell expressed and secreted (RANTES) and monocyte chemotactic activity of ectopic endometrial stromal cells by nuclear factor (NF)-kappaB decoy oligonucleotides (ODN). METHODS: The stromal cells of ectopic endometrium were divided into 3 groups. Two groups were cultured with or without 10 microg/L of interleukin (IL)-1beta. Another group was transfected with NF-kappaB decoy ODN with the aid of a lipofectamine reagent. After 4 h of transfection, 10 microg/L of IL-1beta was added to induce the stromal cells to secrete RANTES. Concentration of RANTES in the supernatant at 4, 8, 12, 24 and 36 h was measured with the sandwich enzyme linked immunosorbent assay (ELISA). U937 monocyte chemotactic activity was assayed in Boyden chambers. The specific RANTES-neutralizing monoclonal antibodies at serial doses (0.5, 1, 2, 4 and 8 mg/L) were added into IL-1beta induced medium of 24 h to detect the monocyte chemotactic activity of RANTES in supernatant. RESULTS: The concentration of RANTES secreted by stromal cells was respectively (58 +/- 10), (150 +/- 35), (360 +/- 46) and (586 +/- 42) ng/L after IL-1beta stimulation for 8, 12, 24 and 36 h, significantly higher than that of stromal cells cultured without IL-1beta. The concentrations of RANTES were respectively (86 +/- 16), (128 +/- 28) and (183 +/- 32) ng/L after IL-1beta stimulation for 12, 24 and 36 h in stromal cells transfected with NF-kappaB decoy ODN, evidently lower than that of stromal cells stimulated with IL-1beta alone. The monocyte chemotactic index of 12, 24, 36 h in conditioned medium of stromal cells transfected with NF-kappaB decoy ODN was respectively 10.3 +/- 0.9, 13.7 +/- 1.1, 18.6 +/- 1.2, which was evidently lower than that of stromal cells stimulated with IL-1beta alone. The anti-RANTES antibody at 0.5, 1, 2, 4 and 8 mg/L inhibited respectively 5%, 23%, 40%, 62% and 61% of the chemotactic activity in 12 h medium treated with IL-1beta. CONCLUSIONS: RANTES accounts for the majority of the monocyte chemotactic activity in IL-1beta induced medium of 24 h. NF-kappaB decoy ODN may influence the feed-forward inflammatory loop whereby IL-1beta from activated macrophages can lead to RANTES production by ectopic implants and further monocyte chemotaxis.

Proteomic analysis of human ovaries from normal and polycystic ovarian syndrome.

Polycystic ovary syndrome (PCOS) is the most common cause of anovulatory infertility, affecting 5-10% of females of reproductive age. Currently, little is known about the changes in whole proteins between PCOS and normal ovaries. In the present study, a proteomic approach comprised two-dimensional gel electrophoresis (2DE) analysis and mass spectroscopy was used to identify proteins and examine expression patterns in three PCOS and normal ovaries. One hundred and ten protein spots were separated and showed different intensities between PCOS and normal ovaries. Sixty-nine proteins associated with cellular metabolism and physiological process were identified from 72 spots. Fifty-four proteins were up-regulated in PCOS ovaries and 15 other proteins were up-regulated in normal ovaries. These data demonstrate, for the first time, the complexity in the regulation of ovarian protein expression in human PCOS, and will provide important insight for a better understanding of the pathogenetic mechanisms underlying this clinical disorder.

Delivery after transfer of frozen-thawed embryos from in vitro-matured oocytes in a woman at risk for ovarian hyperstimulation syndrome.

The present report describes the birth of a healthy infant after cryopreservation of embryos produced from in vitro-matured oocytes retrieved from a woman at risk of developing ovarian hyperstimulation syndrome (OHSS) during conventional in vitro fertilization (IVF) cycles. A conventional long protocol including gonadotropin-releasing hormone agonist (GnRHa) and gonadotropins induced a risk of OHSS. Oocyte retrieval was performed on day 11 of the cycle, and 27 immature oocytes were obtained. Following incubation for 24 h in maturation medium, 74.1% (20/27) of the oocytes were at the metaphase II stage. Fourteen oocytes (14/20, 70.0%) were fertilized after intracytoplasmic sperm injection (ICSI) with her husband's spermatozoa and cultured for 3 days. On day 4 following oocyte retrieval, three embryos at the 8-16 cell stage were transferred into the woman's uterus, and five spare embryos were frozen. Since the fresh embryo transfer failed to result in pregnancy, three post-thaw embryos were transferred into the woman three months later. Transfer of the frozen embryos resulted in pregnancy with delivery of a healthy infant girl.

Pregnancies and births resulting from in vitro matured oocytes fertilized with testicular spermatozoa.

PURPOSE: In vitro maturation (IVM) of immature human oocytes is an attractive option for the treatment of infertility. Similarly, intracytoplasmic sperm injection (ICSI) followed by testicular fine needle aspiration (TEFNA) is an important treatment for primarily male-factor infertility. This report highlights the combination of these two advanced assisted reproduction techniques, namely IVM and fertilization with TEFNA-retrieved spermatozoa by ICSI to overcome both of male and female infertility problems. METHODS: Before immature oocyte retrieval (IOR), gonadotropin stimulation was given for 3 or 5 days. Following IVM, and mature oocytes were inseminated by ICSI followed by TEFNA. RESULTS: Four couples with five completed treatment cycles were performed, and total of 36 immature oocytes were retrieved. Following 36 to 48 h of culture, 32 (88.89%, 32/36) oocytes became mature. The mature oocytes were inseminated with TEFNA-retrieved sperm, and 18 (56.25%, 18/32) oocytes were fertilized normally following ICSI. Eleven embryos were transferred in five cycles and two pregnancies and two singleton births were achieved in two patients. CONCLUSIONS: This result demonstrates that the successful pregnancies and live births can be established from embryos produced from in vitro matured oocytes that fertilized with testicular sperm.

Fertilization of in vitro matured human oocytes by intracytoplasmic sperm injection (ICSI) using ejaculated and testicular spermatozoa.

AIM: To evaluate the fertilization competence of spermatozoa from ejaculates and testicle when the oocytes were matured in vitro following intracytoplasmic sperm injection (ICSI). METHODS: Fifty-six completed cycles in 46 women with polycystic ovarian syndrome were grouped according to the semen parameters of their male partners. Group 1 was 47 cycles that presented motile and normal morphology spermatozoa in ejaculates and Group 2 was the other nine cycles where male partners were diagnosed as obstructive azoospermia and spermatozoa could only be found in testicular tissue fragment. All female patients received minimal stimulation with gonadotropin. Immature oocytes were matured in vitro and inseminated by ICSI. The spermatozoa from testes were retrieved by testicular fine needle aspiration. RESULTS: A total of 449 and 78 immature oocytes were collected and cultured for 48 hours, 75.5 % (339/449) and 84.6 % (66/78) oocytes were matured in Groups 1 and 2, respectively. The percentage of oocytes achieving normal fertilization was significantly higher in Group 1 than that in Group 2 (72.9 % vs. 54.5 %, P 0.05). There were no significant differences in the rates of oocytes cleavage and clinical pregnancies in these two groups [87.4 % (216/247) vs. 88.9 % (32/36); 21.3 % (10/47) vs. 44.4 % (4/9)]. A total of 15 babies in the two groups were healthy delivered at term. CONCLUSION: It appears that IVM combined with ICSI using testicular spermatozoa can produce healthy infants, while the normal fertilization rate of in vitro matured oocytes after ICSI using testicular spermatozoa was significantly lower than using the ejaculated spermatozoa.

[In vitro maturation, fertilization and embryo transfer of human immature oocyte].

OBJECTIVE: To try a new method for treating infertile women by in vitro maturation technique. METHODS: To set up in vitro maturation (IVM) program, including stimulating regimen, oocyte retrieval technique, and culture system of the immature oocytes with the medium consisted of essential supplements and human mature follicular fluid. IVM was performed in 30 infertile patients (35 cycles), (1) with polycystic ovary syndrome (POCS) (14/30), (2) ovary hyperstimulation syndrome (OHSS) in the previous stimulating cycles (6/30), and (3) retardation of the oocyte growth in conventional IVF cycles (10/30). RESULTS: Totally 203 oocytes were obtained, with a mean of 5.8 oocytes each cycle. First polar bodies were observed in 156 oocytes, maturation rate was 76.8%. There were 120 oocytes getting into pronucleus stage, normal fertilization rate was 76.9%. Embryo transfer was performed in 33 cycles. Eight pregnancies were established, pregnancy rate was 24%. Five patients have delivered 7 healthy babies. CONCLUSIONS: The result of our study indicated that IVM is one of the new assisted reproductive technologies for the infertile women with PCOS, poor response to stimulation and the history of OHSS. Human mature follicular fluid would be an ideal component in the maturation medium for oocyte maturation in vitro and provides a much simplified and practical method for clinical use.