Embryo quality in natural versus stimulated IVF cycles

BACKGROUND: The impact of controlled ovarian stimulation (COS) on oocyte and subsequent embryo quality remains controversial. In the present study we have compared embryo quality in natural and stimulated cycles in the same group of patients. METHODS: This retrospective study was comprised of patients with a regular menstrual cycle who had IVF after COS using rFSH in a long GnRH agonist protocol. In all stimulated cycles the patients had fresh embryos transferred and surplus good quality embryos cryopreserved. Subsequently the same patients were treated with a modified FER cycle (mFER) where thawing of the frozen embryos was combined with aspiration of the dominant follicle in the natural cycle. The embryo cleavage stage and quality score were compared between the stimulated and the natural cycle for the patients having an embryo in the natural cycle. RESULTS: In 177 cases patients returned for mFER in a natural cycle. Spontaneous ovulation had occurred in 35 cycles. In 17 cycles no oocyte was retrieved at aspiration and in 125 cycles 128 oocytes were aspirated. In the stimulated cycles from these patients we had obtained 950 embryos (cleavage rate 70.4%) versus 85 embryos (cleavage rate 66.4%) (P = 0.34) in the natural cycles. Comparing the embryos in the natural and stimulated cycles in all patients having an embryo in the natural cycle, we found no difference in the distribution between the different cleavage stages. Of the cleaved embryos, 53% in the stimulated cycles had >or=4 cells versus 59% in the natural cycles after 2 days culture (P = 0.31). In the stimulated cycles 61% of the embryos had <10% fragmentation at the time of transfer on day 2, compared to 69% in the natural cycles (P = 0.15). CONCLUSION: The administration of exogenous gonadotrophins was not reflected in cleavage capacity or quality assessment of the resulting embryos.     

Plasmatische Gerinnungsfaktoren und Wundheilung

Ohne Zusammenfassung a. G. Mit 3 Abbildungen Vortragender:D. Borm.     

Embryo quality and developmental potential is compromised by age

AIMS: The aim of this retrospective study was to assess whether and how the age of the woman affects the quality and developmental potential of the oocytes and embryos in an ART program. METHOD AND MATERIAL: A total of 878 IVF cycles was included as a consecutive series of single transfers (n=292), dual transfers (n=366) and triple transfers (n=220), where all the transferred embryos in each cycle were of identical quality score and identical cleavage stage. RESULTS: We found a highly significant decrease in oocyte recovery with increasing age with about one oocyte per 2.3 years (95% CI 1.8 years to 3.1 years, p<0.0001). Further, we found that the number of oocytes that cleaved declined significantly with increasing age with one per 3.7 years (95% CI 2.7 years to 5.5 years, p<0.0001). This decline was mainly due to the decline in number of oocytes retrieved as the ratio of aspirated oocytes that cleaved with increasing age (approx. -0.04/10 year 95% CI: -0.10; +0.009) was not significantly different (p=0.10). The percentage of transfers using fragmented embryos did not increase significantly with increasing age (p=0.08). The odds of fragmentation increased by 3% per year. The average number of embryos transferred decreased significantly (p=0.03) with age from approximately 2.1 at the age of 25 to approximately 1.8 at the age of 40. In a selected subgroup of embryos all consisting of good quality embryos, a significant decrease was found in implantation rate with increasing age (approx. -0.08/10 years, 95% CI: -1.6; +0.00, p=0.05). Of the 357 pregnancies achieved in this study we found a significantly decreased ongoing pregnancy rate and a significantly increased abortion rate with increasing age (p=0.03). The decrease in the rate of ongoing pregnancies was almost linear, decreasing by approximately 1.5% per year. CONCLUSIONS: We conclude that age has an impact throughout a woman's reproductive life and that it is important to realize that the age-related decline in fertility may start already in the late twenties and not in the mid-thirties as is generally assumed.     

Nuclei size in relation to nuclear status and aneuploidy rate for 13 chromosomes in donated four cells embryos

Purpose The aim was to elucidate if the nuclear size and number are indicative of aberrant chromosome content in human blastomeres and embryos. Methods The number of nuclei and the nucleus and blastomere size were measured by a computer controlled system for multilevel analysis. Then the nuclei were enumerated for 13 chromosomes by a combination of PNA and DNA probes. Results In the mononucleated embryos there was no difference in the mean size of chromosomally normal and abnormal nuclei but a significant difference in the mean nuclei size of nuclei that had gained chromosomes compared to nuclei that had lost chromosomes. The nuclei from multinucleated blastomeres had a significant smaller mean size and the frequency of chromosomally aberrant blastomeres was significantly higher. Conclusion The mean nuclear size is not a marker for the chromosome content in mononucleated embryos. However, it seems that the nuclei size can be related to multinucleation and maybe to the chromosome content.     

Computer-controlled, multilevel, morphometric analysis of blastomere size as biomarker of fragmentation and multinuclearity in human embryos

BACKGROUND: Little is known about blastomere size at different cleavage stages and its correlation with embryo quality in human embryos. Using a computer system for multilevel embryo morphology analysis we have analysed blastomeres of human embryos and correlated mean blastomere size with embryonic fragmentation and multinuclearity. METHODS: A consecutive cohort of 232 human 2-, 3- and 4-cell embryos from patients referred for ICSI treatment were included. Sequences of digital images were taken by focusing at 5- micro m intervals through the embryo. Blastomere sizes and number of nuclear structures were evaluated based on these sequences. The degree of embryonic fragmentation was evaluated by normal morphological assessment prior to transfer and correlated to the blastomere sizes. RESULTS: As a result of normal cell cleavage, mean blastomere size decreased significantly from a volume of 0.28 x 10(6) microm(3) at the 2-cell stage to 0.15 x 10(6) microm(3) at the 4-cell stage (P < 0.001). Mean blastomere size decreased significantly (P < 0.001) with increasing degree of embryonic fragmentation, where highly fragmented embryos showed a 43-67% reduction in blastomere volume compared with embryos with no fragmentation. Multinucleated blastomeres were significantly larger than non-multinucleated blastomeres (P < 0.001). On average, multinucleated blastomeres were 51.5, 67.8 and 73.1% larger than their non-multinucleated sibling blastomeres at the 2-, 3- and 4-cell stage, respectively. Furthermore, the average volume of non-multinucleated blastomeres originating from multinucleated embryos was significantly smaller than the average volume of the blastomeres from mononucleated embryos (P < 0.001). CONCLUSIONS: The results of this study show that the average blastomere size is significantly affected by degree of fragmentation and multinuclearity, and that computer-assisted, multilevel analysis of blastomere size may function as a biomarker for embryo quality.     

Changes induced in serum protein profiles by ovarian stimulation during in-vitro fertilization--embryo transfer treatment: a comparison between conception and non-conception cycles.

The profiles of plasma protein concentrations during the follicular phase in unstimulated women and in women undergoing ovarian stimulation for in-vitro fertilization--embryo transfer (IVF-ET) treatment are described. Plasma protein concentrations are correlated with those of total oestradiol (protein-bound and free) and total progesterone. In addition, 10 conception cycles and 18 non-conception cycles are compared in an attempt to identify predictors of successful treatment. Ovarian stimulation caused a significant increase in follicular phase in serum concentrations of sex hormone binding globulin (SHBG), cortisol binding protein (CBP) and insulin-like growth factor binding protein 1 (IGFBP1). In contrast no increase was observed in unstimulated cycles. Serum levels of endometrial protein PP14 decreased significantly during the follicular phase in both stimulated and unstimulated cycles. Levels of pregnancy zone protein (PZP) were more than doubled at the time of oocyte aspiration compared to the unstimulated cycles. Albumin concentrations were unchanged by the stimulation. Throughout the follicular phase, levels of SHBG were significantly higher, and total oestradiol significantly lower in women who became pregnant, than in those who did not. Therefore, a low concentration of free, biologically active oestradiol seemed to favour pregnancy, as the concentration of albumin is similar in the two groups. The endometrial protein PP14 was significantly lower during the follicular phase in conception than in non-conceptional cycles. On day 2 of the treatment cycle, the PP14 concentration showed a 75% correct prediction of conception and non-conception cycles. These results suggest that levels of PP14 may predict successful IVF cycles even before hormonal treatment is commenced.     

Resumption of mitosis during post-thaw culture: a key parameter in selecting the right embryos for transfer

This retrospective study of 701 thaw cycles analysed the clinical importance of whether or not embryos resumed mitosis during 24 h of post-thaw culture. A total of 3360 frozen embryos were thawed; 1922 embryos survived the freeze-thaw procedure with at least one intact blastomere and were then cultured for 24 h before transfer. All transfers were registered into either the 'cleaved embryo group' (n = 459), which was defined as transfers where at least one of the transferred embryos cleaved during the post-thaw culture period, or the 'non-cleaved embryo group' (n = 153), where none of the transferred embryos cleaved during the post-thaw culture period. A total of 1408 thawed embryos were transferred in 612 cycles; 459 embryo transfers were in the cleaved embryo group, resulting in an implantation rate of 10%, significantly higher than the 4% in the non-cleaved embryo group (P = 0.0003). A total of 130 pregnancies (28% per transfer) were obtained in the cleaved embryo group which was significantly higher than the 17 pregnancies (11% per transfer) obtained in the non-cleaved embryo group (P = 0.0001). However, the average number of transferred embryos was significantly higher in the cleaved embryo group (2.46 +/- 0.03) compared to the non-cleaved embryo group (1.82 +/- 0.07). No difference was found in the age of the women between the two groups. When analysing transfers where all transferred embryos had cleaved during the post-thaw culture period the clinical pregnancy rate increased significantly from 13% transferring two embryos to 36% transferring three embryos (P = 0.0136). In this latter subgroup an implantation rate as high as 17% was obtained. The overall multiple pregnancy rate was 16%. The multiple pregnancy rate was 19% in the cleaved embryo group. In conclusion, 24 h post-thaw culture may allow a better selection of the embryos and thereby we may be able to increase the implantation and pregnancy rates. This may enable us further to reduce the number of embryos transferred.      

Time to pregnancy in relation to semen quality assessed by CASA before and after sperm separation.

BACKGROUND: In order to provide a reference for infertile men, we defined normal values of semen quality in a population of fertile men, using computer-assisted semen analysis (CASA) before and after sperm separation. Additionally, we investigated the relationship between semen quality and time to pregnancy (TTP). METHODS AND RESULTS: Semen samples were obtained from 315 proven fertile men. The median sperm concentration in fresh samples was 107 x 10(6)/ml (5-95 percentiles: 16-322 x 10(6)/ml), the median percentage of motile sperm cells was 65% (14-87%) and the median percentage of progressively motile cells was 37% (5-64%). After density gradient sperm separation, the median total sperm count was 46 x 10(6) (4-350 x 10(6)), the median percentage of motile sperm cells was 77% (16-95%) and the median percentage of progressively motile cells was 63% (11-84%). No significant associations were found between TTP and sperm counts or sperm motility, either before or after sperm separation. This may be due in part to the fact that the study comprised couples with proven fertility. CONCLUSION: We have defined semen parameters, including the results of density gradient separation, in a population of normal fertile men which may be of interest in the evaluation of semen samples from infertile men.