The effect of sperm DNA fragmentation on ICSI outcomes depending on oocyte quality.

BACKGROUND: Sperm deoxyribonucleic acid (DNA) fragmentation is commonly encountered in spermatozoa, and the oocyte assumes responsibility for repairing sperm DNA fragmentation during the oocyte-embryo transition. OBJECTIVES: This study aimed to investigate whether the effect of sperm DNA fragmentation on intracytoplasmic sperm injection outcomes depends on the incidence of oocyte dimorphisms. MATERIALS AND METHODS: For the present cohort, 2942 fertilized oocytes from 525 patients submitted to intracytoplasmic sperm injection cycles were assessed. The present study was conducted in a private in vitro fertilization center affiliated to a university from June 2016 to July 2019. Semen samples were divided into the following two groups depending on the sperm DNA fragmentation index: a low fragmentation index group (<30% sperm DNA fragmentation, n = 1468) and a high fragmentation index group (≥30% sperm DNA fragmentation, n = 486). In addition, mature oocytes were examined before sperm injection, and intracytoplasmic and extracytoplasmic defects were recorded. The effect of the sperm DNA fragmentation index on laboratory and clinical intracytoplasmic sperm injection outcomes (depending on the presence of oocyte defects) was evaluated. RESULTS: Significant increases in the rates of fertilization, high-quality embryo, implantation, and pregnancy were noted for cycles with <30% sperm DNA fragmentation than cycles with ≥30% sperm DNA fragmentation (regardless of the presence of oocyte dimorphisms). The presence of dimorphisms significantly impacted laboratory and clinical outcomes. The lowest fertilization and high-quality embryo rates were observed when a high sperm DNA fragmentation index was associated with the presence of dark cytoplasm, vacuoles, resistant membrane, and non-resistant membrane. The lowest implantation and pregnancy rates were observed when a high sperm DNA fragmentation index was associated with the presence of vacuoles, defective perivitelline space, and fragmented polar body. The effect of sperm DNA fragmentation on miscarriage rates was significantly influenced by the presence of centrally located cytoplasmic granulation, a defective perivitelline space and non-resistant membrane. CONCLUSION: A high sperm DNA fragmentation index increases the likelihood of miscarriage in intracytoplasmic sperm injection cycles, an effect that may potentially be magnified by the presence of oocyte dysmorphisms.

Increasing paternal age and ejaculatory abstinence length negatively influence the intracytoplasmic sperm injection outcomes from egg-sharing donation cycles.

BACKGROUND: The paternal role in embryogenesis is much more than providing a haploid genome. The spermatozoa have crucial roles related to embryogenesis' epigenetic regulation, fusion of gametes, and cleavage, which may influence intracytoplasmic sperm injection outcomes. OBJECTIVES: To study whether paternal age, length of ejaculatory abstinence and semen parameters influence the outcomes of intracytoplasmic sperm injection in the oocyte recipients from egg-sharing donation cycles. MATERIALS AND METHODS: This historical cohort study involved 427 oocyte recipient intracytoplasmic sperm injection cycles, from 321 patients undergoing egg-sharing donation cycles from January 2015 to May 2017, in a private in vitro fertilization center. Electronic medical records of donors and recipients intracytoplasmic sperm injection cycles were reviewed. The impact of male age, length of ejaculatory abstinence, and semen parameters on recipients' intracytoplasmic sperm injection outcomes were investigated using general mixed models. Implantation, pregnancy, and live birth rates were the primary outcome measures. RESULTS: Paternal age negatively affected fertilization, day 3 top-quality embryos, day 3 normal embryo cleavage speed, blastocyst formation, high-quality blastocysts, and implantation, and was correlated with diminished pregnancy and live birth rates. Length of ejaculatory abstinence negatively influenced the rates of day 3 top-quality embryos, normal embryo development on day 3, blastocyst development, and implantation. Sperm count positively affected the rates of fertilization, blastocyst development, and implantation. The percentage of progressive sperm motility positively affected day 3 normal embryo development rate and implantation rate. Total motile sperm count positively affected blastocyst development rate and implantation rate. DISCUSSION AND CONCLUSION: Additional tracking of the influence of male characteristics on intracytoplasmic sperm injection outcomes should be stimulated. Paternal age is not manageable, but reducing length of ejaculatory abstinence could be an alternative approach to improve the outcomes of intracytoplasmic sperm injection.

Concomitant use of FSH and low-dose recombinant hCG during the late follicular phase versus conventional controlled ovarian stimulation for intracytoplasmic sperm injection cycles.

The objective of this study was to investigate the effects of low-dose hCG supplementation on ICSI outcomes and controlled ovarian stimulation (COS) cost. Three hundred and thirty patients undergoing ICSI were split into groups according to the COS protocol: (i) control group (n = 178), including patients undergoing conventional COS treatment; and (ii) low-dose hCG group (n = 152), including patients undergoing COS with low-dose hCG supplementation. Lower mean total doses of FSH administered and higher mean oestradiol level and mature oocyte rates were observed in the low-dose hCG group. A significantly higher fertilization rate, high-quality embryo rate and blastocyst formation rate were observed in the low-dose hCG group as compared to the control group. The miscarriage rate was significantly higher in the control group compared to the low-dose hCG group. A significantly lower incidence of OHSS was observed in the low-dose hCG group. There was also a significantly lower gonadotropin cost in the low-dose hCG group as compared to the control group ($1235.0 ± 239.0×$1763.0 ± 405.3, p < 0.001). The concomitant use of low-dose hCG and FSH results in a lower abortion rate and increased number of mature oocytes retrieved, as well as improved oocyte quality, embryo quality and blastocyst formation and reduced FSH requirements.

Protein expression in human cumulus cells as an indicator of blastocyst formation and pregnancy success.

PURPOSE: The goal for the present study was to implement a technique for protein extraction and identification in human cumulus cells (CCs). METHODS: Forty samples of CCs were collected after ovum pick-up from patients undergoing intracytoplasmic sperm injection (ICSI). Samples were split into the blastocyst group (n = 10), including patients in which all embryos converted into blastocysts, and the non-blastocyst group (n = 10), including patients in which none of the embryos reached the blastocyst stage or the positive-pregnancy (n = 10) and negative-pregnancy group (n = 10). Proteins were extracted and injected into a liquid chromatography system coupled to a mass spectrometer. The spectra were processed and used to search a database. RESULTS: There were 87 different proteins in samples from the blastocyst and non-blastocyst groups, in which 30 were exclusively expressed in the blastocyst group and 17 in the non-blastocyst group. Among the 72 proteins detected in the pregnancy groups, 19 were exclusively expressed in the positive, and 16 were exclusively expressed in the negative-pregnancy group. CONCLUSIONS: CC proteomics may be useful for predicting pregnancy success and the identification of patients that should be included in extended embryo culture programs.

Freeze-all, oocyte vitrification, or fresh embryo transfer? Lessons from an egg-sharing donation program.

OBJECTIVE: To compare the outcomes of ETs using cryopreserved embryos, cryopreserved oocytes, or fresh embryos. DESIGN: Observational, cohort study. SETTING: Private university-affiliated fertility center. PATIENT(S): This study included 8,210 mature oocytes obtained from 425 oocyte donors. Of those, 5,440 were used for the donors' own cycles (Fresh Oocyte Cycles Group), and 2,770 were cryobanked for 425 recipients (Banked Donor Egg Group). All of the oocytes were sperm injected, resulting in 4,585 embryos from the donors' own cycles and 2,128 embryos from the recipients' cycles. For the donor cycles, embryos were either cryopreserved and transferred during a subsequent cycle (Thaw Cycles Group, 3,209 embryos), or they were transferred during a fresh cycle (Fresh Cycles Group, 1,307 embryos). For the recipient cycles, embryos derived from vitrified oocytes were transferred (Vitrified Oocytes Group, n = 425 cycles, 2,128 embryos). INTERVENTION(S): Oocyte/embryo vitrification and intracytoplasmic sperm injection. MAIN OUTCOME MEASURE(S): Embryo quality, pregnancy, and implantation rates. RESULT(S): Decreased embryo quality and lower rates of blastocyst formation were observed among embryos derived from vitrified oocytes. The highest pregnancy and implantation rates were noted for the Thaw Cycles Group, followed by the Banked Donor Egg Group; the Fresh Cycles Group had the lowest rates. CONCLUSION(S): Oocyte vitrification followed by intracytoplasmic sperm injection leads to lower embryo developmental competence compared with when fresh insemination methods are used. However, pregnancy and implantation rates are higher when embryos are transferred into a "more receptive" endometrium, free of the adverse effects of gonadotropin. Moreover, the freeze-all method leads to exceptional clinical outcomes.

Strategies for the management of OHSS: Results from freezing-all cycles.

OBJECTIVE: To compare the use of GnRH agonist (GnRHa) or hCG trigger in potential OHSS patients undergoing freeze-all programs. We also compared the clinical outcomes when fresh versus freeze-thawed embryo transfers were performed in cycles with a high number of retrieved oocytes. METHODS: The study included potential OHSS patients who received GnRHa (n=74) or hCG (n=49) trigger. The protocols were compared with respect to the clinical outcomes. We also compared the clinical outcomes of cycles in which hCG trigger was used and more than 20 MII oocytes were retrieved when: fresh embryo transfer protocol (n=153) or freeze-all protocol (n=123) were performed. RESULTS: A decreased serum estradiol level, a decreased number of retrieved oocytes, an increased MII retrieved rate, and decreased fertilization rate was observed in the hCG when compared with the GnRHa group. No significant differences were noted concerning clinical outcomes. When fresh cycles were compared with frozen-thawed cycles, the estradiol serum level and the number of cryopreserved embryos were higher in the frozen-thawed cycles. The clinical pregnancy rate was higher among freeze-all cycles, as well as the implantation and cumulative pregnancy rates, when compared with fresh embryo transfer cycles. CONCLUSION: The use of GnRHa trigger may be a good alternative to prevent the OHSS in patients presenting an extreme ovarian response to COS, leading to similar clinical outcomes, when compared with the traditional hCG trigger. Moreover, our findings demonstrated that the strategy of freezing-all embryos not only decreases the risk of OHSS but also leads to a better pregnancy rate.

Racial and ethnic differences in assisted reproduction treatment outcomes: the benefit of racial admixture.

The goal of the present study was to determine whether racial and ethnic differences affect the outcomes of assisted reproductive technology in the Brazilian population. 1497 patients undergoing intracytoplasmic sperm injection (ICSI) cycles were split into groups according to the patient's ethnicity: Caucasian (n = 2131), Mestizo (n = 358), Asian (n = 174), Black (n = 115) and Indian (n = 260). ICSI outcomes were compared among the groups. Body mass index was highest in the Black group, followed by the Mestizo, Indian, Caucasian and Asian groups (p > 0.001). The FSH dose (p > 0.001) was highest among Indians, followed by Asians and Caucasians, and the dose was lowest among Blacks and Mestizos. In contrast, the oocyte yield was highest among Mestizos, followed by Indians, Blacks and Caucasians, and lowest among Asians (p = 0.005). The fertilisation rate was highest among Mestizos, followed by Blacks, Indians and Caucasians, whereas Asians had the lowest fertilisation rate (p = 0.004). Pregnancy and implantation rates were also highest among Mestizos, followed by Blacks, Indians and Caucasians, whereas the Asian patients had the lowest rates (p = 0.008 and p > 0.001, respectively). In conclusion, our evidence suggests a possible beneficial effect of racial admixture on ICSI outcomes.

Blastocyst Morphology Holds Clues Concerning The Chromosomal Status of The Embryo.

BACKGROUND: Embryo morphology has been proposed as an alternative marker of chro- mosomal status. The objective of this retrospective cohort study was to investigate the association between the chromosomal status on day 3 of embryo development and blas- tocyst morphology. MATERIALS AND METHODS: A total of 596 embryos obtained from 106 cycles of intra- cytoplasmic sperm injection (ICSI) followed by preimplantation genetic aneuploidy screening (PGS) were included in this retrospective study. We evaluated the relation- ship between blastocyst morphological features and embryonic chromosomal altera- tion. RESULTS: Of the 564 embryos with fluorescent in situ hybridization (FISH) results, 200 reached the blastocyst stage on day 5 of development. There was a significantly high- er proportion of euploid embryos in those that achieved the blastocyst stage (59.0%) compared to embryos that did not develop to blastocysts (41.2%) on day 5 (P<0.001). Regarding blastocyst morphology, we observed that all embryos that had an abnormal inner cell mass (ICM) were aneuploid. Embryos with morphologically normal ICM had a significantly higher euploidy rate (62.1%, P<0.001). As regards to the trophectoderm (TE) morphology, an increased rate of euploidy was observed in embryos that had nor- mal TE (65.8%) compared to embryos with abnormal TE (37.5%, P<0.001). Finally, we observed a two-fold increase in the euploidy rate in high-quality blastocysts with both high-quality ICM and TE (70.4%) compared to that found in low-quality blastocysts (31.0%, P<0.001). CONCLUSION: Chromosomal abnormalities do not impair embryo development as ane- uploidy is frequently observed in embryos that reach the blastocyst stage. A high-quality blastocyst does not represent euploidy of chromosomes 13, 14, 15, 16, 18, 21, 22, X and Y. However, aneuploidy is associated with abnormalities in the ICM morphology. Further studies are necessary to confirm whether or not the transfer of blastocysts with low-quality ICM should be avoided.

The impact of food intake and social habits on embryo quality and the likelihood of blastocyst formation.

The aim of this study was to evaluate the influence of patients' lifestyle factors and eating habits on embryo development. A total of 2659 embryos recovered from 269 patients undergoing intracytoplasmic sperm injection cycles were included. The frequency of intake of food items and social habits were registered and its influences on embryo development evaluated. The consumption of cereals, vegetables and fruits positively influenced the embryo quality at the cleavage stage. The quality of the embryo at the cleavage stage was also negatively correlated with the consumption of alcoholic drinks and smoking habits. The consumption of fruits influenced the likelihood of blastocyst formation, which was also positively affected by the consumption of fish. Being on a weight-loss diet and consumption of red meat had a negative influence on the likelihood of blastocyst formation. The likelihood of blastocyst formation was also negatively influenced by the consumption of alcoholic drinks and by smoking habits. The consumption of red meat and body mass index had a negative effect on the implantation rate and the likelihood of pregnancy. In addition, being on a weight-loss diet had a negative influence on implantation rate. Our evidence suggests a possible relationship between environmental factors and ovary biology.

Poor-responder patients do not benefit from intracytoplasmic morphologically selected sperm injection.

PURPOSE: To compare the outcomes of ICSI and IMSI in women presenting with poor ovarian response. METHODS: Data of IMSI cycles performed from January 2011 to December 2013 were included in this retrospective cohort study. Patients were divided into two groups: normoresponder patients (NR group; patients with > 4 oocytes retrieved) and poor-responder patients (PR group; patients with ≤ 4 oocytes retrieved). Patients who underwent IMSI were matched with patients who underwent ICSI in the same period. The ICSI and IMSI outcomes were compared in the NR and PR groups. RESULTS: A total of 414 matched cycles were included in this study. The NR group comprised 324 cycles (164 ICSI and 160 IMSI cycles), and the PR group comprised 90 cycles (43 ICSI and 47 IMSI cycles). In the NR group, no significant differences were observed between the ICSI- and IMSI-treated couples regarding cycle outcomes. In the PR group, fertilisation rate was significantly lower in IMSI-treated couples (53.9% ± 36.7% vs. 79.8% ± 29.3%). The proportion of cycles with embryo transfer (57.4 vs. 79.1%) and the number of transferred embryos (1.5 ± 0.8 vs. 1.9 ± 0.7) were significantly lower in IMSI compared with ICSI. Implantation, pregnancy and miscarriage rates were similar when ICSI or IMSI were performed. CONCLUSIONS: Our results suggest that unselected couples undergoing ICSI that present with poor ovarian response to controlled ovarian stimulation do not benefit from sperm selection under high magnification prior to ICSI.

The combination of pronuclear and blastocyst morphology: a strong prognostic tool for implantation potential.

PURPOSE: To (i) investigate a possible association between different features of pronuclear (PN) morphology and different features of blastocyst morphology, (ii) evaluate the combination of PN and blastocyst morphologies as a predictive factor for ICSI outcomes and (iii) identify possible contributing factors to poor PN morphology. METHODS: This study included 908 normally fertilised zygotes reaching full blastocyst stage, obtained from 350 patients undergoing ICSI cycles, in which the implantations rates were 0 % or 100 %. The influence of PN morphology on blastocyst morphology and on the rates of pregnancy and miscarriage was investigated. Embryos were graded and split into three groups, taking into consideration both the PN and the blastocyst status. The pregnancy rate was compared among these groups. RESULTS: Inner cell mass (ICM) alterations were correlated with the number of nucleolar precursor bodies (NPB), while trophectoderm alterations were correlated with the size of the pronuclei and the distribution of the NPB. The distribution of the NPB had an impact on the chances of pregnancy. A significant difference was observed among the groups regarding the pregnancy rate. The maternal age, number of aspirated follicles and number of retrieved oocytes influenced the incidence of PN defects. CONCLUSIONS: These findings suggest that a lower oocyte yield may lead to higher-quality PN zygotes. In addition, different PN features may influence further embryo development, especially the quality of the blastocyst. Moreover, the association between PN and blastocyst morphology may be used as a prognostic tool for implantation.

Influence of oocyte dysmorphisms on blastocyst formation and quality.

OBJECTIVE: To identify the association between oocyte dysmorphisms and blastocyst developmental competence. DESIGN: Case-control study. SETTING: Private assisted reproduction center. PATIENT(S): This study included 5,516 embryos, which were obtained from 605 patients undergoing intracytoplasmic sperm injection cycles in which ET was performed on day 5 of embryo development. INTERVENTION(S): Intracytoplasmic sperm injection and ET. MAIN OUTCOME MEASURE(S): The morphologic characteristics of the blastocysts, namely the degree of blastocoel expansion and hatching status, the size and compactness of the inner cell mass (ICM), and the cohesiveness and number of trophectoderm cells, were correlated with oocyte dysmorphisms. RESULT(S): Of the 5,516 embryos evaluated, 2,920 (52.9%) reached the blastocyst stage. The blastocysts' degree of expansion and hatching status were decreased by the presence of aggregates of smooth endoplasmic reticulum clusters, large perivitelline space, and shape abnormalities. The presence of a large perivitelline space also decreased the ICM and trophectoderm cell quality. Aggregates of smooth endoplasmic reticulum clusters also decreased the ICM quality, pregnancy rate, and the miscarriage rate. CONCLUSION(S): The results presented here suggest that the individual identification of oocyte dymorphisms may be a prognostic tool for blastocyst development and quality. However, the results do not diminish the importance of cleavage-stage embryo morphology assessment to predict blastocyst development competence.

The predictive value of high-magnification sperm morphology examination on ICSI outcomes in the presence of oocyte dysmorphisms.

PURPOSE: To evaluate the relationship between oocyte dysmorphisms and IMSI outcomes METHODS: Data of IMSI cycles performed in 332 patients were included in this study. Patients were included only if presented more than four and less than 30 oocytes upon oocyte retrieval. Patients who underwent IMSI were matched, concerning female age and sperm parameters, with patients who underwent ICSI in the same period (n = 332). The two groups (ICSI and IMSI) were compared with regard to treatment outcomes. The influence of IMSI on fertilization and embryo quality on D3 and D5, when oocyte dysmorphisms were present was analyzed. RESULTS: A total of 6444 oocytes were morphologically evaluated and injected. Regardless of the oocyte quality, IMSI performance was a determinant of the increased odds of development to high quality embryo on D3 (OR: 1.98; CI: 1.54-2.56) and D5 (OR: 3.27; CI: 1.61-6.66). CONCLUSIONS: The selection of a morphologically normal spermatozoon under high-magnification is associated with increased embryo quality, regardless of the oocyte morphology. The injection of a spermatozoon, selected under high-magnification, into a morphologically normal oocyte leads to the highest probability of developing high quality embryos.

Patient selection criteria for blastocyst transfers in extended embryo culture programs.

PURPOSE: To identify the correlation between different cycles, patient factors and blastocyst characteristics. METHODS: The study included 420 patients undergoing ICSI cycles and 2781 graded blastocysts, which took into account the blastocyst quality. The correlations between the blastocyst parameters and the patient and cycle characteristics were assessed. RESULTS: The blastocyst development was negatively correlated with the maternal age, BMI and dose of FSH. The ICM was negatively correlated with the FSH dose, whereas the TE quality was influenced by the FSH dose, the maternal age and the number of retrieved oocytes. The embryo morphology on days two and three may predict the blastocyst developmental competence. CONCLUSIONS: Older patients and patients with high BMI should not be included in extended embryo culture programmes. The extended culture may not favour embryos with poor morphology on days two and three of development. Additionally, a lower ovarian stimulation and decreased oocyte yields may lead to the development of high-quality blastocysts.

Oocyte yield and dysmorphisms as indicators of biological efficiency in intracytoplasmic sperm injection cycles.

The aim of the study was to examine whether oocyte yield could be an indicator of morphological oocyte quality and biological competency in patients younger than 36 years undergoing controlled ovarian stimulation (COS). Three hundred and thirty-five intracytoplasmic sperm injection (ICSI) procedures were arbitrarily subdivided into five groups according to the number of retrieved oocytes. Patients' demographic characteristics and treatment success were compared among the groups. The influence of the morphological oocyte abnormalities on outcomes was also investigated. The proportion of oocytes that gave rise to viable embryos and high-quality embryos decreased significantly according to oocyte yield. Similarly, the number of foetal heartbeat per retrieved oocyte in fresh embryo transfer cycles was higher in patients with fewer oocytes collected. Finally, a negative correlation was observed between the occurrence of intracytoplasmic oocyte dysmorphisms and the number of foetal heartbeat per oocyte. High oocyte yield may be considered an indicator of low oocyte biological efficiency and intracytoplasmic dysmorphisms may contribute to this biological wastage suggesting that protocols of minimal or mild stimulation should be used.

Effect of acupuncture on assisted reproduction treatment outcomes.

BACKGROUND: Acupuncture has recently been used as a complementary technique in the management of infertility. It has physiological and psychological effects and may be considered an alternative for stress reduction in women undergoing infertility treatments. OBJECTIVE: To examine the hypothesis that acupuncture treatment may increase the pregnancy rate in patients undergoing intracytoplasmic sperm injection cycles. METHODS: Patients enrolled in the study were stratified according to age and randomised to either a control group (n=208) or acupuncture group, (n=208). The pregnancy, implantation and abortion rates of the two groups were compared. RESULTS: No influence of acupuncture treatment on clinical outcomes was seen; however, when cycles in which the causes of infertility were exclusively tubal-uterine or idiopathic were evaluated separately, a positive influence of acupuncture on pregnancy (OR=5.15, 95% CI 1.03 to 34.5; p=0.048) was noted. Moreover, trends toward an increase in implantation were seen when acupuncture was performed (regression coefficient: 0.645; p=0.092). CONCLUSION: The results suggest that acupuncture treatment had no influence when performed immediately before and immediately after embryo transfer, on clinical outcomes overall. In a subgroup analysis, when the embryo was not affected by an ovarian or seminal influence, a benefit was noted.

Poor ovarian response in patients younger than 35 years: is it also a qualitative decline in ovarian function?

OBJECTIVE: To investigate whether poor response to controlled ovarian stimulation (COS) is due to a qualitative decline in ovarian function. METHODS: This retrospective cohort study included 436 patients younger than 35-years old, undergoing COS for intracytoplasmic sperm injection (ICSI). Patients with four or fewer MII oocytes after COS (poor-responder group, PR, n = 52) were age-matched with normoresponder patients (NR, n = 364). RESULTS: Although similar duration of stimulation (10.5 +/- 0.4 and 9.3 +/- 0.8 days; p = 0.1358), increased doses of gonadotrophins (2510 +/- 865 and 2253 +/- 572 IU; p = 0.0061) were used in the PR. The results show a increased chance of cycle ending of PR (PR: 26.9% and NR: 3.1%; p < 0.0001). Although the lower total number of oocytes retrieved (2.4 +/- 1.4 and 16.2 +/- 9.3; p < 0.0001), equal rate of fertilization (70.2% and 72.0%, p = 0.1190) and high quality embryos were obtained (50.0% and 45.2%; p = 0.4895), resulting in similar implantation (14.5% and 19.7%; p = 0.2246) and abortion (10.0% and 15.4%; p = 1.00) rates, respectively. A trend towards increased pregnancy rate per embryo transfer in NR group was noted (PR: 26.3% and NR: 42.2%; p = 0.0818). CONCLUSIONS: Low ovarian response could be associated mainly with a quantitative rather than a qualitative decline in ovarian function. Therefore, even if the ovarian response to stimulation is low, patients aged < or =35 years should process to oocyte retrieval.