The role of mode of conception in the outcome of twin pregnancies.

Artificial reproductive techniques (ART) have become a routine and successful treatment of infertility. Twin pregnancies are more frequent after ART compared to spontaneous conception. Varying findings have been reported for the obstetric and perinatal outcome of ART twin pregnancies as compared to spontaneously conceived twin pregnancies. To evaluate the specific risks of twin pregnancies achieved through ART, literature has been reviewed for obstetric and perinatal risks and differences between artificially and spontaneously conceived twin pregnancies. Eleven studies have been involved in this review. Three of the studies were matched and eight were non-matched studies. Results of analysed studies differed widely. Twin pregnancies resulting from ART showed an increased rate of Caesarean section and tend toward higher risk for preterm birth and low birthweight compared to spontaneously conceived twin pregnancies. Various conditions can influence twin pregnancies outcome after ART. In in-vitro fertilization programs twin pregnancies should be avoided and physicians have to inform patients of the specific risks when transferring more than one embryo.

Morphological analysis at compacting stage is a valuable prognostic tool for ICSI patients.

In terms of treatment outcome, little prognostic power is attributed to day-4 morphology. A day-4 score was applied to 56 patients separating non-compacting embryos from compacting (some areas of compaction) and fully compacted embryos. The latter were further subdivided according to the morphology of compaction. Grade C1 embryos represented optimal quality, while grades C2 (exclusion of fragments) and C3 (exclusion of blastomeres) were characterized by a loss of cytoplasm. Grade 4 embryos (C4) showed incomplete compaction with several blastomeres not yet incorporated into cell mass. Pooled embryos without compaction showed a reduced (P < 0.001) blastulation (28.8%) as compared with compacting embryos with the same cell number (68.8%), which, in turn, revealed lower (P < 0.05) rates of blastulation as compared with concepti that completed compaction process (84.6%). Among fully compacted embryos grade C1 had a better (P < 0.01) blastocyst formation rate (94.4%) as compared with grade C3 (68.2%). Grade C1 embryos showed significantly higher rates of top-quality blastocysts as compared with grade C2 (P < 0.05) and C3 (P < 0.01). Blastocysts deriving from grade C1/C4 embryos led to a higher pregnancy rate as compared with the C2/C3 counterparts (P < 0.05). This modified score allows for adequate prediction of both blastocyst formation/quality and pregnancy.

Blood clots in the cumulus-oocyte complex predict poor oocyte quality and post-fertilization development.

Assessment of oocyte maturity and quality (morphological appearance) at the time of retrieval is difficult as the egg is obscured by a large cumulus mass that hinders adequate scoring. Since no data are available on the possible relationship between the cumulus-oocyte complex (COC) and oocyte morphology, this prospective intracytoplasmic sperm injection study was set up in 87 consecutive patients. COC were grouped according to expansion of both corona radiata and cumulus matrix. Special emphasis was placed on recording morphological anomalies of COC (inclusion of blood clots and amorphous clumps). For all mature ovae, quality was assessed and preimplantation development followed up to blastocyst stage if fertilized. The risk of not harvesting an oocyte was higher in COC with blood clots compared with normal cumulus matrices (P = 0.004). COC expansion did not allow for prediction of either nuclear status or quality of the egg. The presence of blood clots within the cumulus matrix was associated with reduced oocyte quality (dense central granulation), fertilization rate and blastocyst formation, compared with unaffected COC (P < 0.05). It may be postulated that COC showing blood inclusions derive from poor quality follicles, which has a detrimental effect on oocyte quality and further cleavage to blastocyst stage. Consequently, mechanical removal of blood clots cannot rescue the corresponding embryo.

Developmental fate of ovoid oocytes.

BACKGROUND: Irregularities in composition, thickness and/or color of the zona pellucida may impair optimal function and result in reduced outcome. Anomalies of oocyte shape have not been investigated in detail in this respect. METHODS: Therefore, all patients attending our clinic within a period of 1 year were screened for the presence of ovoid gametes and the corresponding developmental potential was evaluated. For all elongated gametes, a roundness index (RI; length divided by width) was calculated in order to quantify shape. RESULTS: RI did not affect fertilizability (P > 0.05). The degree of dysmorphism was found to be related to cleavage pattern. The more ovoid a gamete was, the higher was the risk of the corresponding zygote not cleaving like a tetrahedron (P < 0.01). Abnormal cleavage (a rather flat array of blastomeres) was associated with delayed compaction (P < 0.01) and blastocyst formation (P < 0.001). The quality of blastocysts was not affected at any stage in ovoid concepti (P > 0.05). CONCLUSIONS: Ovoid oocytes with abnormal cleavage pattern show delayed preimplantation development, probably due to a reduced number of cell-to-cell contacts.

[Multiple pregnancies after ART: problems and possible solutions]. / Mehrlingsschwangerschaften nach ART: Probleme und Losungsansatze.

Multiple pregnancies following an assisted reproduction technique (ART) should be seen as a complication, and for that reason they should be avoided. In contrast to singleton pregnancies following ART, the multiple pregnancies are associated with a higher prenatal, neonatal and maternal risk; furthermore this results in a financial burden for the health care system. This paper gives an overview of the latest literature and different attempts of European countries, trying to reduce the multiple pregnancy rate. An efficient reduction is only possible by single-embryo transfers. There should be strict and cross-national regulation for the choice of women who should have a transfer of more than one embryo.

Incomplete denudation of oocytes prior to ICSI enhances embryo quality and blastocyst development.

BACKGROUND: Granulosa cells are essential mediators of oocyte maturation and fertilization. Because of the denudation of oocytes in preparation for ICSI, any potential positive effect of surplus cumulus cells (CCs) on further development would be unable to exert further effect. In order to evaluate the actual influence of adhering cumulus cells on further preimplantation development, this prospective study was carried out. METHODS: Sibling cumulus-oocyte complexes for 57 ICSI patients were split into a study group (incomplete denudation, n = 314) and a control group (complete denudation, n = 336). According to the cumulus cell pattern after partial denudation, mature gametes from the study group were further subdivided into type A oocytes, which showed several prominent CC clusters (n = 202), and type B (n = 75), which showed a more homogeneous pattern with CC covering the whole surface of the gamete. RESULTS: In immature oocytes, presence of adhered CCs led to a significant increase in resumption of meiosis (P < 0.01). Fertilization rate (P < 0.05) and ability to cleave (P < 0.01) was impaired in the study group, because of difficulties in ICSI of type B oocytes. By contrast, embryo morphology on days 2 (P < 0.01) and 3 (P < 0.05), as well as blastocyst formation, was better (P < 0.05) in the study group (55 blastocysts out of 88 zygotes) as compared to that in the control group (49/105). CONCLUSION: These data indicate that co-culture of oocytes with attached CCs may enhance preimplantation development.

Basal level of anti-Müllerian hormone is associated with oocyte quality in stimulated cycles.

BACKGROUND: Serum anti-Müllerian hormone (AMH) levels provide a powerful means for predicting ovarian response, which is reflected not only by the size of the primordial follicle pool but also by the quality of the oocytes. Considering a mutual interdependence between AMH-expressing somatic cells and gametes, this prospective morphological study was set up to evaluate whether extreme AMH levels represent diminished oocyte quality and developmental incompetence. METHODS: A total of 141 consecutive ICSI patients were subdivided into three groups using the 25th and 75th percentiles of the serum AMH levels (cycle day 3). In these three groups, morphology of all oocytes and fertilization rate, embryo quality and blastocyst formation were evaluated, and FSH, LH and estradiol (E(2)) levels were also measured. RESULTS: Cycle cancellation rate was correlated with AMH levels (P < 0.05). AMH groups 1 (<1.66 ng/ml) and 3 (>4.52 ng/ml) showed oocytes of lower quality [dark central granulation, aggregation of smooth endoplasmic reticulum (sER)] compared with the median group 2 (1.66-4.52 ng/ml). Basal serum FSH did not allow for adequate prognosis in terms of gamete appearance. Fertilization and further cleavage up to blastocyst stage was not affected by AMH levels. CONCLUSIONS: AMH seems to be superior to FSH in predicting both oocyte number and quality.

Complete oocyte activation failure after ICSI can be overcome by a modified injection technique.

BACKGROUND: Complete fertilization failure after ICSI is a rare event, and it may happen repeatedly even in cases of normal sperm parameters and good ovarian response. In these cycles, alternative ICSI techniques may prove useful. METHODS: Our modified ICSI (mICSI) is characterized by aspiration close to the opposite membrane (the region of the mitochondria with a high inner mitochondrial membrane potential) which is followed by central deposition of the sperm. The method was applied prospectively to ICSI cycles of patients with a history of complete fertilization failure in previous ICSI cycles. In parallel, mICSI was compared with conventional ICSI in terms of further preimplantation development and treatment outcome. RESULTS: In patients with previous ICSI failures using conventional ICSI (no 2Pn zygotes out of 70 oocytes that had been injected) application of mICSI led to adequate fertilization (53.6%) and pregnancy rates (33.3%) (P < 0.001; P < 0.01). In patients without previous failed fertilization, no improvement in the rates of fertilization, blastocyst formation, implantation or clinical pregnancy could be seen. CONCLUSIONS: Our data indicate that the present version of ICSI is a reliable alternative to conventional ICSI. However, although it overcomes oocyte-dependent activation failure, routine application does not improve the overall results.

Laser-assisted zona pellucida thinning prior to routine ICSI.

BACKGROUND: In MII oocytes showing difficult oolemma breakage, ICSI can cause an increase in the degeneration rate. This may be overcome by laser-assisted ICSI using a 5-10 micro m opening in the zona pellucida for injection. However, such a small opening might impair the hatching process, especially if assisted hatching is applied in addition. In order to prevent this, the present study used routine injection through an area of zona pellucida in which laser zona thinning had been applied, providing for both a reduced mechanical stress to the oocyte and assisted hatching. METHODS: This prospective study involved 100 cycles with 1016 MII oocytes. Conventional ICSI (control group) was compared with a modified laser-assisted ICSI (study group) in sibling oocytes. In the latter group oocytes were injected through an extended area of zona thinning. RESULTS: Degeneration rate was significantly lower in the study group (P < 0.004). There were no differences in fertilization, or formation and quality of blastocysts. In the study group embryo quality on day 2 was significantly better (P = 0.004) and herniation of day 5 blastocysts was increased (P = 0.005). Rates of implantation and pregnancy were not affected. However, on day 3 laser-assisted ICSI proved beneficial (P = 0.038) in terms of clinical pregnancy rate. CONCLUSIONS: The new method combines a less invasive ICSI technique with assisted hatching. Our preliminary data indicate that in addition to an improved oocyte survival, this new approach increases the hatching rate in vitro, which may explain the increase in pregnancy rate, at least in day 3 transfers.

Presence, but not type or degree of extension, of a cytoplasmic halo has a significant influence on preimplantation development and implantation behaviour.

BACKGROUND: Since there is considerable disagreement in grading cytoplasmic haloes, this prospective study was set up to evaluate if certain subtypes of haloes are related to further development. METHODS: Out of a total of 152 patients, 713 zygotes could be checked for the formation of a halo. Where present, haloes were subdivided into concentric (symmetric) and polar (asymmetric) types. In addition, each halo was measured accurately to see if the extension of the halo might influence further development. In parallel, pronuclear patterns were checked. RESULTS: Halo-positive zygotes did not differ from halo-negative ones in terms of embryo quality and blastocyst formation rate. However, quality of blastocysts (assessed by their inner cell mass consistency) was significantly increased (P < 0.001) if a halo appeared at zygote stage. This phenomenon was not related to type of halo or degree of halo. In terms of pronuclear pattern, pattern 0 (0A, 0B) led to significantly more blastocysts (P < 0.001) of better quality (P = 0.002) compared with patterns 1-5. A stepwise logistic regression showed no relationship between different halo types and pronuclear pattern 0. CONCLUSIONS: The present study indicates that any halo has a positive prognostic value on blastocyst quality, irrespective of the fact that it is light or extreme, polar or concentric. In addition, the developmental advantage of pattern 0 is confirmed.

Selection based on morphological assessment of oocytes and embryos at different stages of preimplantation development: a review.

In contrast to IVF, in ICSI the surrounding cumulus and corona cells must be removed completely, as only denuded oocytes can be successfully manipulated by the holding pipette. This ancillary effect of ICSI allows us to focus on the morphology of preimplantation development from the earliest stages. Early prognosis regarding the developmental fate of oocytes would help to limit a negative impact of culture conditions. However, little evidence is available that non-invasive selection at the oocyte stage (first polar body, granular cytoplasm) may be of prognostic value. Recently, certain patterns of pronuclei (number and the distribution of nucleoli) at the zygote stage were found to correlate with treatment outcome in IVF and ICSI cycles, offering an additional prognostic tool prior to cleavage. As there is evidence that embryo selection on day 2 or 3 based on morphological criteria (fragmentation, number of blastomeres, multinucleation, uneven cleavage) may be imprecise, patients might benefit from extended embryo culture to day 5. However, not all major chromosomal aberrations are incompatible with blastocyst formation, and prolonged culture in vitro does not exclusively select embryos with a normal chromosomal complement. Consequently, special care should be taken to minimize the presence of aneuploid concepti in culture. In addition, multiple selection at different stages of development will be required to filter out the correct 'candidate' embryo which will result in a healthy newborn.

Developmental competence of oocytes showing increased cytoplasmic viscosity.

BACKGROUND: The objective of the study was to investigate the developmental fate of oocytes with increased cytoplasmic viscosity as assessed by the persistence of the injection funnel after withdrawal of the ICSI pipette. METHODS: For this purpose, 1008 oocytes showing a characteristic injection funnel during ICSI were subdivided into two groups according to the oocyte's ability to restore its spherical shape within 2-3 min after ICSI. Fertilization and further development was evaluated in both groups. In addition, implantation and pregnancy rates were analysed. RESULTS: In the funnel positive cohort (group 1) significantly fewer oocytes degenerated after injection (P < 0.01) compared with oocytes without persistent funnel (group 2). However, at zygote stage, presence of a halo (P < 0.05) and a optimal pronuclear pattern 0 (P < 0.01) was increased in group 2. In addition, significantly fewer poor quality embryos were found in this group (P < 0.01). The number of good quality blastocysts but not blastocyst formation was increased in group 2 (P < 0.05). This resulted in an increased clinical pregnancy rate if embryos which derived exclusively from funnel negative oocytes were transferred (P < 0.05). CONCLUSIONS: Our data suggest that cytoplasm of higher viscosity delays development up to cleavage stage and impairs optimal development. Injection funnel persistence was found to be a negative prognostic marker of preimplantation development.

[Sonographic measurement of endometrium thickness as a predictive value for pregnancy through IVF]. / Die sonographische Endometriumdicke als Prediktionsfaktor für eine Schwangerschaft in einem IVF-Programm.

AIM: This prospective study was performed to evaluate the predictive value of endometrium thickness in patients undergoing IVF-ET. METHOD: Measurements of endometrium thickness as well as pattern assessments were performed in 936 cycles (722 patients) on the day of administering human chronic gonadotropin (HCG). RESULTS: The overall pregnancy rate was 31.1 % (291/936). The age of non pregnant patient was significantly higher (p < 0.029). There was a significant influence of oestradiol (p < 0.029), number of transferred embryos (p < 0.004), and embryo quality (p < 0.002) on the pregnancy rate. Overall, the mean (+/- SD) endometrium thickness was 11.16 mm (+/- 2.13). The mean (+/- SD) endometrium thickness of pregnant patients was 11.25 mm (+/- 2.19). The mean (+/- SD) endometrium thickness of non-pregnant patients was 11.12 mm (+/- 2.10). A stepwise logistic regression analysis showed no statistically significant correlation between endometrium thickness and pregnancy rate (p < 0.23). CONCLUSION: Sonographic measurement of endometrium thickness on the day of human chorionic treatment with gonadotropin is not useful in predicting IVF outcome.

First polar body morphology and blastocyst formation rate in ICSI patients.

BACKGROUND: It may be beneficial to identify, at a very early stage of development, concepti that will result in viable blastocysts by using a non-invasive technique. METHODS: Homogeneous groups in terms of first polar body (PB) morphology were analysed with regard to fertilization, embryo quality and blastocyst formation. The strategy was to transfer a maximum of two blastocysts with an adequate inner cell mass deriving from oocytes with identical first PBs in order to obtain information about the actual implantation potential. RESULTS: A significant relationship between first PB morphology and embryo quality was found. Fragmentation after 2 days was increased in embryos derived from oocytes with fragmented first PBs (P < 0.05) in comparison with those derived from oocytes with intact PBs. No similar correlation could be demonstrated for fertilization rate. Embryos in the intact first PB group showed an increased rate of blastocyst formation as compared with the fragmented first PB group (P < 0.05). In addition, a significant difference in implantation rate (48.6 versus 22.0%; P < 0.025) and ongoing pregnancy rate (68.4 versus 34.8%; P < 0.05) was observed for the intact versus fragmented groups respectively. CONCLUSION: In conclusion, the current study provides further evidence that preselection at a very early stage may be helpful in identifying a subgroup of preimplantation embryos with a good prognosis to form blastocysts and, consequently, to implant.

Prospective hatching of embryos developed from oocytes exhibiting difficult oolemma penetration during ICSI.

BACKGROUND: The hormonal milieu during ovarian stimulation is known to affect oolemma behaviour as well as zona pellucida thickness and structure. This led us to investigate whether a special subgroup of patients with oocytes where penetration of the oolemma is difficult during ICSI may benefit from assisted hatching. METHODS: A total of 77 couples (mean age: 32.9 +/- 4.6 years; range: 22-38) had oocytes that could hardly be penetrated by the ICSI pipette. Nineteen patients underwent two ICSI cycles, giving a total number of 96 cycles, which were randomly split into either the study group (n = 52) or the non-hatching group (n = 44). Hatching was done using a non-contact 1.48 mm wavelength diode laser. Implantation and pregnancy rates were recorded. RESULTS: The pregnancy rate was 36.6% (19/52) in the study group and 13.6% (6/44) in the non-hatching group (P < 0.05). In addition, a higher number (P < 0.05) of embryos implanted in the study group (23/106; 21.7%) than in the non-hatching group (9/92; 9.8%). CONCLUSIONS: Once oolema penetration during ICSI has proven difficult, prospective hatching of embryos considered for transfer may increase their implantation behaviour.

Laser assisted immobilization of spermatozoa prior to intracytoplasmic sperm injection in humans.

BACKGROUND: The conventional method of immobilization of spermatozoa prior to intracytoplasmic sperm injection (ICSI) is mechanical breakage of the tail by pressing it against the bottom of the injection dish. METHODS: This prospective self-controlled study was set up to evaluate the potential of a non-contact 1.48 microm wavelength diode laser in terms of immobilization. In addition, the fertilization rate and further development potential of such zygotes were investigated. The patients included in our study (n = 60) had oestradiol concentrations >2000 pg/ml, and thus a relatively high number of MII oocytes could be expected. Approximately half the oocytes were injected with laser treated spermatozoa (n = 262, study group) and the other half with mechanically immobilized spermatozoa (n = 252, control group). RESULTS: No significant differences between the two groups in terms of fertilization rate, early cleavage or blastocyst formation were observed. However, time required for identification, aspiration and injection of a potential spermatozoa was significantly shorter in the laser immobilized sperm group (P < 0.001). CONCLUSIONS: The application of a non-contact diode laser for sperm immobilization prior to ICSI is a potentially useful alternative to the conventional mechanical approach.

Acute puerperal inversion of the uterus--treatment by a new abdominal uterus preserving approach.

OBJECTIVE: Inversion of the uterus is still a rare (1, 2) but serious and life-threatening obstetric complication. It is said to be complete when the fundus uteri protrudes through the cervix and into the vagina. Within minutes a state of shock is reached due to pulling forces on the peritoneum as well as blood loss. METHODS: The vagina was entered by a longitudinal incision (3 m) below the contraction ring. Through this opening it was possible to advance two fingers (second and third finger of the left hand) into the vagina above the invaginated corpus uteri. The invaginated cavum uteri was loaded on these two fingers and, exerting counterpressure with the right hand, the inside was turned out. CONCLUSION: The present operative method guarantees easy reposition of the uterus in cases of failed vaginal manual repositioning. Furthermore, the cavum uteri remains intact and hysterectomy can be avoided.

Embryo fragmentation in vitro and its impact on treatment and pregnancy outcome.

OBJECTIVE: To determine the impact of embryo fragmentation on pregnancy, obstetric, and perinatal outcome. DESIGN: Retrospective analysis of embryo transfers that were homogeneous in regard to the degree of fragmentation. SETTING: Fertility center. PATIENT(S): A cohort of 460 fresh embryo transfers. INTERVENTION(S): A total of 164 pregnancies were analyzed for the incidence of antepartum complications during gestation, obstetric (multiple pregnancy, preterm delivery, cesarean section), and perinatal outcome (sex, birth weight, admission to neonatal intensive care unit, malformations). MAIN OUTCOME MEASURE(S): Implantation and clinical pregnancy rate, obstetric and perinatal outcome. RESULT(S): Embryo fragmentation and number of embryos per transfer showed a significant influence on clinical pregnancy and implantation rate. No such relation was found concerning complications, multiple pregnancy rate, incidence of cesarean section, gestation week, birth weight, and average time at the neonatology. On the other hand, pregnancies derived from bad-quality embryos had a significantly higher rate of malformations. CONCLUSION(S): The higher percentage of malformations found in bad-quality embryos may be due to a higher percentage of apoptotic features and chromosomal disorders. For ethical reasons, the transfer of embryos with >50% fragmentation should be considered only after consultation with the patient.

A prospective study on oocyte survival rate after ICSI: influence of injection technique and morphological features.

PURPOSE: To determine the influence of technical pitfalls and oocyte morphology on survival rate and cleavage behavior after ICSI. METHODS: A total of 2210 injection procedures was examined for morphological and technical deviations. Survival rate and cleavage behavior were evaluated. RESULTS: In 77.8% of all cases ICSI was unsuspicous. Out of 491 deviations from optimal injection deep penetration of the oocyte and abundant presence of cumulus cells showed significant correlation with degeneration rate (p < 0.001). Morphological anomalies associated with the periphery of the oocyte were rather related to degeneration than cytoplasmic anomalies (p < 0.001). Early embryonic development was not impaired by technical or morphological parameters. CONCLUSIONS: To conclude, these prospective data may be of prognostic value in regard of the number of embryos available for transfer and may help to improve treatment outcome.