Cumulus cells apoptosis as an indicator to predict the quality of oocytes and the outcome of IVF-ET

Purpose: Our purpose was to establish an evaluation system for oocyte quality based on the incidence of cumulus cells apoptosis and to examine the effect of coculture, using autologous cumulus cells, on the outcome of IVF–ET according to proliferative activities of helper cells and the incidence of cumulus cells apoptosis. Methods: Cumulus cell masses were collected from 91 mature oocytes among 330 oocytes retrieved from a total of 34 IVF–ET cycles with tubal infertility and unexplained infertility. The incidence of apoptosis in cumulus cells was assessed by apoptosis detection kit fluorescein. On ovum pick up, 2nd day embryos were cocultured with autologous cumulus cells. Prior to coculture, in vitro proliferative activity of cumulus cells was evaluated. Results: Cumulus cells from patient groups over 40 years old had a significantly increased apoptosis incidence, a lower fertilization rate, and the decreased number of oocytes retrieved compared to the other age groups (P < .05). The incidence of cumulus cells apoptosis was significantly lower when the number of oocytes retrieved was 5 or less (P < .05). Cumulus cells from fertilized oocytes (0.43 ± 0.07%) and those from patients who became pregnant (0.44 ± 0.11%) following IVF–ET showed a significantly lower incidence of apoptosis compared to those of unfertilized oocytes (1.80 ± 0.35%; P < .001) and the nonpregnant group (0.81 ± 0.10%; P < .05). Embryo quality also had a negative correlation with the incidence of cumulus cells apoptosis. Coculture of fertilized oocytes with cumulus cells with high proliferative activity resulted in improved rates of implantation and pregnancy compared to that with poor active cumulus cells. No significant difference was found between the in vitro proliferative activity of cumulus cells and the incidence of cumulus cells apoptosis (P < .063). Conclusions: The age of women might influence the incidence of apoptosis in cumulus cells, and the increased incidence of apoptosis is associated with the number of oocytes retrieved, the fertilization rate, and the pregnancy outcome following IVF–ET. These results suggest that the incidence of cumulus cells apoptosis can be used in predicting oocyte quality, outcome of IVF–ET, and age-related decline in fertility.     

Relationship of the Human Cumulus-Free Oocyte Maturational Profile with In Vitro Outcome Parameters After Intracytoplasmic Sperm Injection

Purpose: We investigated whether the human oocyte maturational profile at the removal of cumulus/corona cells affects the fertilization rate and subsequent embryo quality after intracytoplasmic sperm injection. Methods: A total of 1011 oocytes from 150 cycles was included in this retrospective analysis. Cumulus-free oocytes that were in prophase or metaphase I of meiosis at the removal of cumulus/corona cells were incubated in vitro until they reached metaphase II (in vitro-matured oocytes) and were then immediately injected with a single spermatozoa. Oocytes that were in metaphase II at the removal of cumulus/corona cells (MII oocytes) received sperm injection after 3–4 hr of preinjection incubation. Results: The fertilization rate of the MII oocytes was significantly higher than that of in vitro-matured oocytes (81 vs 62%; P < 0.001). The cleavage rates were similar in the two groups (MII oocytes, 94%; in vitro-matured oocytes, 91%). However, MII oocytes had significantly higher percentages of good-quality embryos (grade 1–3 embryos, 87 vs 58%, P < 0.001) and embryos with high cumulative embryo scores (score 10–32 embryos, 62 vs 33%, P < 0.001). The mean cumulative embryo score of MII oocytes after fertilization was also higher than that of in vitro-matured oocytes (12.1 ± 3.8 vs 8.8 ± 3.4; P = 0.014). Conclusions: MII oocytes that extruded the first polar body at the removal of cumulus/corona cells had better fertilization rates and embryo morphology than in vitro-matured oocytes that extruded the first polar body following the removal of cumulus/corona cells and in vitro culture.     

Coculture of mouse embryos with cryopreserved human oviduct epithelial cells

Purpose The effect of coculturing mouse embryos with cryopreserved human oviduct epithelial cells was investigated. The cryopreserved cells in Cellbanker were thawed and cultured in Richard D. Goldsby culture medium to establish monolayers. Two-cell-stage mouse embryos were cultured alone (control group) or cocultured with monolayers established from cryopreserved cells (cryopreserved coculture group) or from fresh cells (fresh coculture group). The rates of embryo development and the qualities of the blastocysts in the three groups were compared.Results The two coculture groups had significantly higher blastocyst development rates (cryopreserved coculture group, 81.6%; fresh coculture group, 82.2%) than the control group (63.1%). The two coculture groups had significantly more blastomeres (cryopreserved coculture group, 108.3 ± 25.9; fresh coculture group, 108.4 ± 25.1) than the control group (87.7 ± 31.9).Conclusion The method of cryopreservation of human oviduct epithelial cells using Cellbanker is simpler than conventional cryopreservation methods. These cryopreserved human oviduct epithelial cells may provide a constant supply of cells for coculture for in vitro fertilization and embryo transfer.     

Autologous endometrial coculture in patients with a previous history of poor quality embryos

Purpose : To evaluate the effect of autologous endometrial coculture in patients (less than 36 years old) with a history of a single IVF failed cycle associated with poor quality embryos. Methods : Design: Controlled clinical study. Setting: University-based in vitro fertilization center. Patients: Twenty-six patients with a history of a single prior failed IVF-ET with poor preembryo quality. Intervention(s): Autologous endometrial coculture. Main outcome measures: Preembryo blastomere numbers and cytoplasmic fragmentation rates were compared between the treatment and previous cycle. Clinical pregnancy rates were analyzed. Results : Twenty-six women with an average age of 32.8 ± 2.9 years underwent treatment. On Day 3 the overall mean number of blastomeres per preembryo on coculture compared to conventional medium in a previous cycle was 6.1 ± 1.8 vs. 5.1 ± 1.3 (P = 0.01; Wilcoxon test). The average percentage of cytoplasmic fragments on coculture compared to the conventional medium in a previous cycle was 14% ± 10 vs. 22% ± 13 (P = 0.003; Wilcoxon test). At transfer the mean number of blastomeres per preembryo on coculture was 7.4 ± 1.8 compared to 6.7 ± 1.5 on conventional medium in a previous cycle (P = 0.02; Wilcoxon test). The clinical pregnancy rate (positive fetal cardiac activity) per patient was 88.5%. The delivery rate was 73.1% (19/26). Conclusions : There was an improvement in the preembryo quality for preembryos on autologous endometrial coculture compared to noncocultured preembryos from the same patient in a previous cycle. An excellent delivery rate was subsequently found.     

Histologic characteristics of the endometrium predicts success when utilizing autologous endometrial coculture in patients with IVF failure

Purpose : To analyze the success of autologous endometrial coculture (AECC) in improving embryo quality and pregnancy outcome based on the histologic characteristic of the biopsy. Methods : Prospective study of 86 consecutive patients undergoing IVF utilizing AECC. Results : The patients were on average 37.4±4.0 years with a history of 2.6±1.8 failed previous attempts. An overall clinical pregnancy rate of 45.3% per ET was found. The embryos grown in AECC were of an improved quality in comparison to those grown in conventional media. 33.7% (29/86) of the biopsies were out of phase (>3 days). In-phase (IP) and OOP (out of phase) specimens both demonstrated an improvement in embryo quality. However, OOP endometrial biopsies that displayed significant retarded endometrial development (< cycle day 19) did not demonstrate an improvement in embryos grown on AECC as compared to IP endometrial biopsies or OOP endometrial biopsies that demonstrated at least an endometrial development of cycle day 19. Conclusions : We have demonstrated a significant improvement in embryo quality with AECC. We have also demonstrated that histologic dating of the endometrium is predictive of IVF outcome when utilizing AECC.     

A prospective, randomized comparison of two commercial media for ICSI and embryo culture

Purpose:The aim of this prospective, randomized study was to compare the results obtained in ICSI with two culture media, P-1 (Irvine Scientific) and IVF-50 (Scandinavian IVF Science). Methods: A total of 182 patients undergoing ICSI treatment were randomly included in this study and divided in two groups: Group I: P-1 medium (n = 91) or Group II: IVF-50 medium (n = 91). All the embryos were transferred on the second day. Results: Patient age did not differ (p = .29) between Group I (34.8 ± 4.8) and Group II (34.0 ± 4.5). The number of oocytes retrieved from Group I (10.6 ± 6.7) was also similar (p = .49) to that retrieved from Group II (11.1 ± 6.4). In addition, there was no difference (p = .25) in the number of oocytes retrieved at metaphase II between Group I (7.9 ± 4.6) and Group II (8.7 ± 4.6). Normal fertilization rates, abnormal fertilization rates, and cleavage rates were similar (p = .62, p = .48, and p = .9, respectively) between Group I (68.4 ± 23.3%, 6.7 ± 10.3%, and 98.7 ±4.6%) and Group II (65.3 ± 26.2%, 9.0 ± 13.8%, and 98.9 ± 3.9%, respectively). The embryo score was also similar (p = .62) for both groups (Group I: 31.9 ± 14.0 and Group II: 33.4 ± 15.8). There was no difference in the number of embryos transferred (p = .69) between Group I (2.8 ± 1.0) and Group II (2.8 ± 1.1). In addition, pregnancy rates/puncture, pregnancy rates/transfer, implantation rates, and abortion rates were also similar for Group I (36.2%, 37.0%, 17.4%, and 12.1%, respectively) and Group II (31.8%, 33.7%, 15.8%, and 10.3%, respectively) (p = .64, p = .75, p = .72, and p = 1.0, respectively). Conclusions: There were no differences in the results obtained with culture media P-1 (Irvine Scientific) and IVF-50 (Scandinavian IVF Science) for ICSI and embryo culture.     

The effects of coculture with autologous cryopreserved endometrial cells on human in vitro fertilization and early embryo morphology: A randomized study

Purpose: The aim of this study was to examine the influence of endometrial cells on the fertilization rate and early embryonic morphology following routine in vitro fertilization (IVF). Cryopreservation with subsequent thawing allowed the use of autologous somatic cells, thus minimizing the risk of transmission of infective agents. Interpatient variability was eliminated by randomizing oocytes from each cycle into the control or coculture group. Results: Two hundred ninety-four oocytes from 24 IVF cycles (21 patients) were included in the study (145 coculture and 149 control). The normal fertilization rate of control oocytes (56.4%) was not significantly different from that of oocytes cocultured with endometrial cells (61.4%). The mean number of blastomeres in cocultured embryos (3.65) was not significantly different from the number in control embryos (3.46) 2 days after insemination, but the proportion of embryos with minimal or no fragmentation was significantly higher in the coculture group [34/84 (40.5%) vs.17/80 (21.3%);P<0.01]. Conclusions: The inclusion of cryopreserved autologous endometrial cells in routine clinical IVF procedures does not influence fertilization or the early cleavage rate but may reduce the extent of embryo fragmentation during the early cleavage divisions.     

Coculture with homologous oviductal cells improved the implantation of human embryos—A prospective randomized control trial

Purpose: The efficacy of homologous oviductal cell coculture on the success of a humanin vitro fertilization program was investigated in a prospective randomized control clinical trial. Methods and Results: One hundred eighty-one couples were randomized into the control and the coculture groups. Pronu-clear-stage zygotes were either cultured in Earles' balanced salt solution supplemented with 15% preovulatory serum (control) or cultured with human oviductal cells (coculture) for 24 hr before embryo transfer. There was no difference in the age of the patients, indication for treatment, number of oocyte retrieved or fertilized, or number of embryo replaced between the two groups. The pregnancy rates per transfer for the control and the coculture group were 12.8 and 19.3%, respectively. The number of viable fetus was significantly higher (P<0.01, chi-square test) in the coculture group (25/264) than in the control group (8/262). The coculture group also showed a higher multiple pregnancy rate, lower abortion rate, and more spare embryos suitable for cryopreservation.     

Blastocyst culture: Toward single embryo transfers

The routine culture and transfer of viable human blastocysts has been made possible by the development of sequential culture media, formulated to account for the changes in nutrient requirements of the embryo as it develops and differentiates. Resultant implantation rates of blastocysts transferred on day 5 are significantly higher than those obtained by the transfer of cleavage stage embryos transferred on day 2 or day 3 within the same programme. As a direct result of this increase in implantation rate, fewer blastocysts than cleavage stage embryos need to be transferred to obtain acceptable pregnancy rates, thereby reducing the incidence of multiple gestations. Blastocysts developed in sequential culture media are readily cryopreserved. The efficiency of in vitro fertilization (IVF) in a general patient population can be calculated using a model that takes into account the number of embryos transferred and cryopreserved, together with their respective implantation rates. Blastocyst transfer is associated with about a 20% increase in the efficiency of IVF compared with the transfer of cleavage stage embryos on day 3. The development of a suitable scoring system has enabled identification of those blastocysts with the highest developmental potential (70% implantation rate). The culmination of this work should be the move to the transfer of a single blastocyst for a significant number of patients.     

The production of interleukin-1α immunoreactivity by human oviductal cells in a coculture system

Purpose: The aim of this study was to determine the concentration of interleukin-1 in human embryo culture medium with or without oviductal cell coculture and to correlate the interleukin-1 levels with pregnancy. Methods: Culture media from 32 in vitrofertilization and embryo transfer cycles were assayed for interleukin-1 by immunoassay technique. Human embryos were cultured in Earles' balanced salt solution supplemented with 15% preovulatory serum (sEBSS) in 16 of these cycles, while embryos in the rest of the cycles were cocultured with human oviductal cells in sEBSS. Results: Both sEBSS and spent sEBSS after embryo culture contained low or undetectable levels of interleukin-1 in the pregnant and nonpregnant cycles. On the other hand, oviductal cells significantly increased the amount of interleukin-1 immunoreactivity in the conventional culture medium or coculture medium (P<0.001, Mann-Whitney rank sum test). The concentrations of interleukin-1 in the spent sEBSS after oviductal cell culture and after coculture with human embryos were 1.5±1.0 and 1.3±0.9 pg/ml, respectively. There was no difference in the interleukin-1 concentration between the pregnant and the nonpregnant coculture cycles. Conclusions: These data showed that human oviductal cells produced interleukin-1 immunoreactivity in a coculture system. However, this production could not be used as a marker for successful embryo implantation.     

Implantation rates after in vitro fertilization and transfer of a maximum of two embryos that have undergone three to five days of culture

OBJECTIVE: To evaluate implantation and pregnancy rates in patients undergoing IVF after the transfer of a maximum of two embryos that had been cultured for 3-5 days. DESIGN: Prospective study. SETTING: An IVF laboratory at a tertiary referral university hospital. PATIENT(S): One thousand seven hundred eighty-seven couples who underwent their first IVF cycle between January 1995 and December 1997. INTERVENTION(S): In vitro fertilization and transfer of embryos after 3, 4, or 5 days of culture using a single medium without coculture. MAIN OUTCOME MEASURE(S): Implantation and pregnancy rates. RESULT(S): Overall implantation and pregnancy rates were not significantly different with different culture periods. Forty-one percent of all available embryos developed into blastocysts on day 5. The transfer of at least one good-quality blastocyst could be performed in 62% of patients. Blastocysts had an implantation rate of 26% per embryo, whereas the implantation rate of eight-cell embryos on day 3 was 18%. Implantation rates for retarded, normal, and advanced embryos were not significantly different with an extended culture period. CONCLUSION(S): Under the study conditions, the transfer of embryos after 5 days rather than 3 days of embryo culture did not change the overall implantation and pregnancy rates. The implantation potential of embryos available for transfer can be assessed better after an extended culture period. Five days of culture allows the transfer of a reduced number of embryos without decreasing overall pregnancy rates.     

Clinical Outcome of Day 2 Versus Day 3 Embryo Transfer Using Serum-Free Culture Media: A Prospective Randomized Study

Purpose: The objective was to evaluate whether extending the embryo culture period from 2 to 3 days would yield a more optimal selection of viable embryos, thereby increasing the implantation and live birth rates. Methods: Patients undergoing in vitro fertilization with at least one oocyte fertilized were prospectively randomized to 2 or 3 days of embryo culture in serum-free media. On the basis of their morphology and cleavage rate, a maximum of three embryos was selected for transfer. Results: Embryos transferred on day 2 or day 3 were similar morphologically, however, a higher proportion of retarded embryos was observed on day 3. The implantation rate was 15.8 and 14.3% for day 2 and day 3 transfers, respectively. The increase in live birth rate from 18.5 to 22.6%, possibly suggesting a better embryo selection on day 3, was not statistically significant. Conclusions: Extending the embryo culture period from 2 to 3 days had no effect on implantation and live birth rates.     

Effects of Reducing Insemination Time in Human In Vitro Fertilization and Embryo Development by Using Sibling Oocytes

Purpose: Recent studies showed a beneficial effect of reducing the time of sperm–oocyte interaction on fertilization, division, and implantation rates of the oocytes obtained from randomized patients. In the present study, the effects of reduced insemination time on fertilization and embryo development were evaluated by using sibling oocytes from the same patient. Methods: A total of 464 oocytes from 36 patients was randomly allocated to be inseminated for either 1 hr (reduced) or 18 hr (regular). Results: Fertilization rates were not significantly different between reduced (135/229; 59%) and regular (150/235; 64%) groups. Cleavage rates and embryo quality were similar in both groups. A total of 135 embryos (73 from the reduced and 62 from the regular group) was transferred to 36 patients. Thirty-four embryos implanted in 18 patients (25.2% implantation and 50.0% pregnancy rates). Conclusions: Fertilization, cleavage, and embryo development from 1-hr insemination is comparable, not superior, to those from an 18-hr insemination time, which is commonly used in in vitro fertilization programs. These data suggest that reduced insemination time can be used during in vitro fertilization to avoid unnecessarily longer exposure to spermatozoa.     

Human follicular fluid and mouse cumulus cells act synergistically to enhance preimplantation mouse Balb/cJ embryo development

Problem The development of preimplantation mammalian embryos in vitro is less than optimal. Follicular fluid and cumulus cells have both been used, independently, to improve preimplantation embryo quality in culture.Method To determine the ability of mouse cumulus cell coculture in the presence of human follicular fluid to support preimplantation mouse Balb/cJ embryo development in vitro.Results Culture of preimplantation mouse Balb/cJ embryos independently in human follicular fluid or on mouse cumulus cells had no significant affect on blastocyst development or total cell number per blastocyst. The coculture of mouse Balb/cJ preimplantation-stage embryos on mouse cumulus cells in the presence of human follicular fluid significantly (P<0.07) improved blastocyst development and the total number of cells per blastocyst.Conclusions Cumulus cells and follicular fluid have a positive synergistic affect on preimplantation mouse Balb/cJ embryo development and formation in vitro.     

Beneficial effects of coculture with cumulus cells on blastocyst formation in a prospective trial with supernumerary human embryos

Purpose: We reported previously on the use of coculture with cumulus cells in insemination medium for the development of human embryos in vitro. Here we describe a prospective trial to determine if this procedure has a significant beneficial effect. Methods: On the day after insemination, zygotes were randomized for culture in either a fresh drop of medium without (– cum) or were left in their insemination drop with (+ cum) cumulus cells. Embryos with the best morphological quality were replaced on the third day of development at the eight-cell stage. The remaining embryos were cultured for a further 3 days and cryopreserved if they reached the fully expanded blastocyst (FEB) stage. Three different culture media were used over the period of this study. Results: In 11 patients, supernumerary embryos were available only for continued culture in + cum and three patients had embryos cultured in only – cum. Thirty-nine other patients had embryos assigned to both + cum and – cum treatments. In the + cum group, 98 blastocysts developed from 216 embryos cultured for 6 days (45%), and this was significantly greater (P<0.01) than the 48 blastocysts from 156 embryos (31%) developing in the absence of cumulus cells. In basal HTF medium (HTF medium with EDTA and glutamine) and basal XI HTF medium (similar to basal HTF but devoid of glucose and phosphate), culture of embryos with cumulus cells produced significantly more FEBs than in the absence of cumulus cells. There was no significant difference between the two culture treatments when regular HTF medium was used. Preliminary results indicate that pronectin-coated dishes provide a good substratum for cumulus cell attachment and embryo development. Conclusions: The culture of human embryos with their cumulus cells in insemination drops of medium produces a significantly greater proportion of FEBs than when the zygotes are transferred to fresh culture drops devoid of cumulus cells. This is the first report of a significantly higher blastocyst rate with coculture in which a real comparison has been made between two culture treatments which differ only in the presence or absence of homologous cumulus cells in insemination drops.Presented in part at the IXth World Congress on In Vitro Fertilization and Alternate Assisted Reproduction. Vienna, Austria. April 3–7, 1995.     

A new model for embryo implantation: coculture of blastocysts and Ishikawa cells

Objective: To explore and develop a new in vitro implantation model that reflects the main process of embryo attachment and invasion. Study design: One of the limitations in human embryo implantation research is lack of an available in vitro model that faithfully replicates human embryo–uterine interactions. In the present study, we examined the attachment and invasiveness of blastocysts from mice in Ishikawa cell (IK), a human endometrial cell, to clarify whether this new model is suitable to study implantation of embryos. We used IK and placed it in contact with blastocysts to initiate coculture experiments using a specifically designed medium. The culture medium was composed of Ham F-12/Dulbecco’s modified Eagle medium (1:1), 30% fetal calf serum, 63.5 nmol/L progesterone, 7.14 nmol/L estradiol-17β, 100?mg/ml of insulin, and 20?ng/ml epidermal growth factor. The culture for 24?h clearly demonstrated that embryos were capable of attachment to the IK and displayed partial invasion. Results: Our results showed that embryos attached to the IK and displayed partial invasion after coculture of blastocysts with IK for 48?h. Conclusions: The model is capable of demonstrating the procedure of attachment and invasion of embryo into the endometrial cells and has promises to be used in studies related to early embryo implantation in human endometrium.     

Intracytoplasmic Sperm Injection Increases Embryo Fragmentation Without Affecting Clinical Outcome

Purpose: To examine the effect of intracytoplasmic sperminjection (ICSI) on embryo fragmentation and implantationrates in those embryos chosen for transfer compared toconventional in vitro fertilization (IVF). Methods: We compared 253 infertility patients (71 ICSI and182 IVF) with respect to age, semen analysis, number ofembryos transferred, embryo fragmentation, implantationrate, and pregnancy rate. Embryo fragmentation wasdetermined by one observer at the same laboratory over the entirestudy period. Results: A statistically significant difference was observedin mean embryo grade between IVF (2.2 ± 0.84) and ICSI(2.5 ± 0.77), P = 0.01. Additionally, the IVF patients hadsignificantly more nonfragmented (grade I) embryoscompared to the ICSI group, P < 0.01. Conclusions: These data suggest that ICSI, irrespective ofsemen parameters, may increase embryo fragmentation andproduce fewer nonfragmented grade I embryos while maintaining implantation and pregnancy rates similar toconventional IVF.     

The effects of sperm quality on embryo development after intracytoplasmic sperm injection

Purpose : To explore the possible relationship between sperm quality and embryo development, pregnancy and implantation rates, in patients undergoing intracytoplasmic sperm injection (ICSI). Methods : Fertilization and cleavage rates, quality of embryos, blastocyst development, pregnancy and implantation rates were analyzed in 1020 embryos from 219 couples undergoing first ICSI treatment cycle. The couples were allocated in five groups, according to semen parameters: Group 1: patients with normal semen parameters, Group 2: patients with mild oligo-astheno-teratozoospermia, Group 3: patients with severe oligo-astheno-teratozoospermia, Group 4: patients with obstructive azoospermia, Group 5: patients with non-obstructive azoospermia. Results : Fertilization and cleavage rates, quality of embryos as well as blastocyst development rates were significantly reduced, as semen quality decreased. However, no significant differences were observed in clinical pregnancy and implantation rates. Conclusion : Overall, a negative relationship was observed between semen quality and embryo development, even before activation of the embryonic genome, suggesting that sperm can affect embryogenesis from a very early stage.     

Randomized Autocontrolled Comparison of the Embryo Culture Performance of Nunc and Falcon Petri Dishes

Purpose: Our purpose was to compare the embryo culture performance of two types of petri dishes (Nunc and Falcon). Methods: Mouse zygotes were cultured up to the expanded blastocyst stage in both types of dishes. The oocytes from 50 in vitro fertilization cycles were randomly divided between the two types of dishes. Fertilization, cleavage, and embryo quality were compared. Oocytes from another 50 cycles were all cultured at random in either type of dish. Pregnancy and implantation rates were compared between the two types. Results: Of 91 mouse zygotes, 81 cleaved to two-cell-stage embryos, and 64 became expanded blastocysts in Falcon dishes; of 99 zygotes, 81 cleaved to two-cell-stage embryos and 66 became expanded blastocysts in Nunc dishes. Of 248 oocyte–cumulus complexes (OCC), 145 fertilized in Falcon dishes, and of 269 OCC, 175 fertilized in Nunc dishes. The high quality embryo ratio was 51 out of 118 in Falcon dishes, not different from that in Nunc dishes, 58 out of 139. In Falcon dishes 72 out of 118 embryos were at least at the four-cell stage after 45 hr, versus 70 out of 139 in Nunc dishes. Twenty-three clinical pregnancies were obtained in the first 50 cycles with sibling oocytes. In the second group, with randomization of the cycles between Nunc and Falcon, 8 pregnancies were obtained in the Nunc and 10 in the Falcon dishes. The implantation rate in this second group of 50 cycles was 9 out of 61 in Falcon and 11 out of 57 in Nunc dishes. Conclusions: No differences were observed.     

Deliveries from embryos fertilized with spermatozoa obtained from cryopreserved testicular tissue

Aim: The data of 167 TESE-ICSI-ET cycles performed with fresh or frozen, motile or immotile testicular spermatozoa were analyzed, retrospectively. Methods: The outcome measures studied were state/condition of spermatozoa, fertilization, embryo developmental, implantation and pregnancy/delivery and abortion rates. Results: No differences were found in fertilization, implantation and pregnancy rates of oocytes injected with fresh or frozen spermatozoa. However, difference was obtained in the fertilization rate of oocytes injected with motile vs. non-motile spermatozoa (72% vs. 62%; P < 0.04). Difference was also observed in embryo development between oocytes injected with fresh vs. frozen spermatozoa (83% vs. 75%; P < 0.03). But, no difference was obtained in embryo development between oocytes injected with motile vs. immotile spermatozoa. No difference was also found in the implantation rate of embryos developed from oocytes injected with motile vs. non-motile spermatozoa. No difference was found in abortion rates either. Conclusions: State/condition of injected testicular spermatozoa has impact to fertilization and embryo development. Pregnancy/delivery can be achieved with frozen/immotile spermatozoa.State and condition of testicular spermatozoa injected has influence the developmental capacity of embryo derived from ICSI.