Three different protocols were utilized for patients undergoing IVF during the study interval, including long gonadotropin releasing hormone GnRH agonist, micro-dose flare and GnRH antagonist protocols. The particular protocol chosen was based on anticipated patient response, with the majority of patients undergoing the long agonist protocol.
A blood sample was taken at the last ultrasound, prior to hCG administration and analyzed for estradiol. Luteal phase supplementation was by daily IM injection of progesterone 50 mg. Clinical pregnancy CP was documented by observing a fetal sac s and heart rate by vaginal ultrasound at 6. Inter-assay and intra-assay coefficients of variation were 9. Fertilization was checked between 17 and 18 hours and normally fertilized oocytes having 2 pronuclei were placed in groups of and cultured as described previously.
On day 2, embryos were graded and embryos allocated to new culture drops based on quality good, fair or poor. A decision was made to perform embryo transfer on day 3 or day 5, based on the number of good quality embryos. If this criterion was not met, there were a low number of embryos or poor quality embryos; transfer on day 3 was elected. New culture plates were made daily and embryo development and video documentation was also conducted daily at approximately 24 hour intervals.
Cleavage stage embryos and blastocysts were graded as being good 1 , fair 2 or poor quality 3 and numerical conversion of embryo grades were performed in order to statistically analyze the data. The embryos or blastocysts that were transferred for each patient were averaged to yield an average embryo quality score AEQS.
The numeric scoring is similar to that used by the Society for Assisted Reproductive Technology [ 23 ]. For cleavage stage embryos, the quality was determined by stage of development, blastomere symmetry and fragmentation of the embryos. A good quality embryo score of 1. A fair quality embryo score of 2. The minimum stage of development of good quality blastocysts was 3 expanded or in the case of an early observation, 2 cavitated. A day 6 good quality blastocyst was similar to day 5 except with more advanced development.
A fair quality blastocyst score of 2 for day 5; the minimum quality was 2BB- and for day 6 was 4BB-. A poor quality blastocyst score of 3 for day 5 or day 6, included any blastocyst having a B-B, or lower quality regardless of stage of development. The poor quality group had an average embryo quality score range of 2. To focus on LB rates, we examined if there was an association between maternal estradiol levels and LB after controlling for potential confounders using multivariate logistic regression.
We examined if estradiol levels and other response variables were different by live birth status. We compared LB rate, number of embryos transferred and embryo transfer quality scores in patients having embryo transfer on day 3 or day 5.
The same approach was taken when examining NLB and LB differences in the high estradiol populations by day of embryo transfer day 3 or day 5. Patient demographics for the entire population are shown in Table 1. For the entire population Table 2 , the higher estradiol groups were associated with greater biochemical and clinical pregnancy rates.
In the multivariate analysis Table 5 however, estradiol level was not related to live birth. Only the main effects of AEQS, patient age, and the transfer of two embryos as opposed to 1 or 3, were found to significantly affect LB in the final model.
IVF response variables in the total population and in the subgroup containing only good and fair quality embryos on the day of embryo transfer were evaluated by live birth status. In the entire population the LB rate was greater for day 5 than for day 3 transfers. However, when the subgroup containing only good and fair embryos was evaluated, although the LB rate was slightly higher When embryo transfer was conducted on day 3 or day 5 with high quality embryos, patients had similar estradiol levels, with the number of oocytes retrieved, mature oocytes and number of 2PN embryos all being higher in the day 5 transfer group.
In high responders there is usually an increased number of eggs retrieved resulting in more embryos and therefore, better embryo selection, leading to more patients undergoing transfer on day 5. In the case of poor responders the opposite is true as was demonstrated in the low estradiol group in Table 2. Here, there were fewer oocytes in spite of a higher dosage of FSH, which also resulted in fewer 2PN embryos and ultimately poorer embryo selection at the time of transfer.
In our experience, patients having embryo transfer on day 3 can attain high pregnancy rates, but only if their embryo quality is not compromised. Therefore, we evaluated a population of patients undergoing their initial IVF cycle to determine the relationship between serum estradiol level and IVF outcomes.
We next evaluated a subgroup that included patients having only good or fair quality embryos transferred to study the influence of embryo quality on pregnancy rate and subsequent live birth outcome. Although our initial categorical analysis showed that in an unselected population of first time IVF patients, estradiol level was significantly related to better pregnancy rates Table 2 , in patients having only good or fair quality embryos transferred, to control for embryo quality influence , estradiol level was no longer related to pregnancy outcome Table 3.
They were divided into three groups according to their age. Percentile E 2 curves according to E 2 levels were plotted. High responders were those patients with E 2 levels above the 90th percentile, normal responders had E 2 between the 10th and 90th percentiles, and poor responders had E 2 below the 10th percentile. Pregnancy rates were higher for high responders, but the difference did not reach statistical significance.
Controlled ovarian stimulation COS for IVF cycles is usually monitored by serum estradiol E 2 levels and pelvic ultrasonography with two purposes: i to obtain an adequate number of mature oocytes, and ii to prevent the risk of severe ovarian hyperstimulation syndrome OHSS. In order to decide whether to proceed with an ongoing cycle and to define the so-called poor and high responders, Phelps et al.
However, the influence of high E 2 levels on the outcome of IVF cycles is still controversial. Simon et al. The use of E 2 levels to differentiate poor, normal and high responders, and to predict IVF outcome, has been the subject of debate. In most studies, cut-off levels were chosen arbitrarily and depend on various factors.
Percentile curves offer a more objective definition of these three categories of patients. To our knowledge, only one study has used percentile curves to adjust the doses of gonadotrophins to the ovarian response and to evaluate IVF outcome in stimulation cycles Forman et al. Zorn, unpublished data. Gonadotrophin dose and ovulation induction by hCG are adjusted according to the pattern of E 2 levels on the percentile curve of the corresponding protocol.
The objectives of the present study were: i to determine the ovarian response in COS with recombinant r FSH in a short GnRH agonist protocol by using E 2 percentile curves; ii to define low, normal and high responders; and iii to predict the IVF outcome by using percentile E 2 serum levels at day 5 and d-hCG.
Due to the retrospective study design based on routine practice, consultation with the hospital ethics committee was not required. Triptorelin Decapeptyl; Ipsen, Paris, France was administered in a dose of 0. A total of cycles were stimulated with Gonal-F and with Puregon. Patients received norethisterone NE Primolut Nor; Schering, Paris, France 10 mg daily for 8—16 days starting from day 3 of a natural cycle.
Triptorelin, 0. Generally, the initial dose for patients in group A was 3 ampoules, for group B, 4 ampoules and for group C, 6 ampoules. From day 5 of the stimulation cycle, doses of rFSH were adjusted according to the number of follicles found on ultrasound and to the E 2 levels.
Oocyte retrieval was performed transvaginally under ultrasound guidance. The oocytes were checked 16—20 h after insemination or microinjection for evidence of fertilization. The oocytes were considered to be normally fertilized when two pronuclei were visible. The embryos were evaluated according to the number of blastomas and the degree of fragmentation.
Up to four embryos were transferred, and any remaining high grade embryos were cryopreserved. IVF outcomes were expressed as the number of oocytes, number of embryos obtained, number of high grade embryos, E 2 peak levels and pregnancy rates PRs at day 5 and d-hCG.
The P10 and P90 cut-offs were chosen based on previous reports for other fields of gynaecology—obstetrics Lubchenco et al. E 2 determination was performed using an immunoenzyme assay by sequential competitive technique on an Immuno-1 multiparameter analyser Bayer reference ES2. The capture antibody used was a rabbit polyclonal antibody against E 2 conjugated to fluorescein.
The competitive marker was constituted by E 2 conjugated to alkaline phosphatase. Separation was performed using a dispersed solid phase that is achieved by magnetic particles linked to monoclonal antibody anti-fluorescein.
The signal was read on the spectrophotometer at and nm. E 2 standard USP in a saline buffer bovine albumin with sodium azoture was used for calibration. The molar conversion factor was 3. Samples with concentration values exceeding the maximal calibrator can be diluted in the zero calibrator or in an albumin buffer. Blood samples were collected in tubes without anticoagulant and separated by centrifugation.
Such overexposure may result in postmature oocytes and end in early abortion. The same group of investigators noted also that in good outcome cycles, E2 continued to rise until hCG was administered, but in nonpregnant cycles, E2 plateaued on the day before hCG administration, which suggests that luteinization or atresia of the more advanced follicles had commenced spontaneously.
Although adequate follicular development occurs with CC and hMG combination regimen, it is thought that one problem with that regimen is premature luteinization In general, it is believed that the rise in serum progesterone occurs 12 hours before or on the day of the onset of a spontaneous LH surge in a natural cycle, or in a controlled ovarian hyperstimulation for IVF-ET program However, there were reports that a significant rise in serum progesterone occurs in advance of the onset of the LH surge in regimens using a combination of CC and hMG It was reported that subtle progesterone rise occurred in A significantly higher serum E2 concentration and a greater number of developed and collected oocytes were observed in those cycles with subtle progesterone rise.
The rate of mature oocyte formation and fertilization were significantly lower, however, and the rate of embryos with cytoplasmic fragments was significantly higher in those cycles. A low pregnancy rate which did not progress to ongoing pregnancy was also observed in those cycles An excessive sensitivity of granulosa cells to LH might induce untimely progesterone production even for a low concentration of serum LH.
The low fertilization rate may be due to the direct determining effect of the subtle progesterone rise on oocyte quality and maturity. A low pregnancy rate after ET may be related to the poor quality of embryos as well as to the direct effect of progesterone on the receptivity of the endometrium due to asynchrony between it and the embryos.
It could be that those patients have diminished ovarian reserves and consequently a poor prognosis for future IVF based on the findings of Navot et al. A reverse effect of CC on ovarian steroid synthesis however cannot be excluded 18, These patients may benefit from a combination of GnRHa and gonadotropin therapy 1, It has been demonstrated that basal LH levels decline, and LH surges are often absent in gonadotropin-treated cycles of humans 17 and animals Suppression of LH secretion during stimulated cycles in which serum E2 is often elevated beyond normal mid cycle levels is considered to be due to inhibition of the pituitary by ovarian factors, or a direct effect of exogenous gonadotropins In some cases however, increase in LH is often observed during gonadotropin therapy Mizunuma et al.
They could recognize three types of premature LH release. The sustained type, defined as an LH release that occurred during the treatment and lasted until hCG administration. The transient type, so-called when LH release occurred during treatment, but returned to normal levels before LH administration. If LH was released only on the day of hCG administration, it was called the onset type.
Those cycles with premature LH release were accompanied by increased FSH levels, and a high incidence of ovarian hyperstimulation. They concluded that ovarian hyperstimulation can be reduced by modulating the dose of FSH and the intervals of administration in cycles showing premature LH release when it occurs early in the cycle and is discovered early.
The doses of hCG administered are in the range , IU. The regimen used by many clinics 37 calls for the administration of hCG on the sixth day of a sustained increase in serum estradiol levels. Patients who fail to achieve adequate follicular development after days of ovarian stimulation do not receive hCG, and the treatment cycle is cancelled. Patients with poor follicular development or with only one developing follicle are not given hCG.
It is inadvisable to give hCG to patients in whom the serum estradiol level is seen to increase rapidly i. Just prior to hCG injection, a serum LH can be drawn and compared to values earlier in the cycle. This helps to identify women who have initiated a premature LH surge LH value 2. However, without frequent sampling of LH every 3 hours , the onset of the surge cannot be identified with precision LH sampling is not required in patients who are treated with GnRHa. If a spontaneous LH surge occurs in a stimulated cycle, some centres cancel the treatment cycle, whereas others give hCG if there is a satisfactory estradiol response and adequate follicular growth has taken place In these cases, it is necessary to adjust the timing of oocyte recovery.
As a general rule, hyperstimulation is associated with the presence of many follicles. It is advisable that hCG not be administered if there are more than follicles of 14 mm or more in diameter Mild hyperstimulation has been associated with an increased number of intermediate size follicles and severe hyperstimulation with an increase in small follicles 2.
A large number 11 or more of small follicles should also preclude hCG administration. Check and colleagues 4 used hCG to trigger ovulation in their patients in whom ovulation was induced by hMG. Some programmes measure the estrogen level on the day following hCG, and if there is a marked drop in the value at that time, retrieval is cancelled because that pattern is associated with a poor chance of pregnancy Oocyte retrieval is performed approximately 35 hours after the hCG injection.
Nowadays, ultrasonically-guided retrieval has replaced laparoscopic oocyte retrieval to a large degree. After completion of the procedure, the ovarian pedicles should be thoroughly observed for torsion, especially in the presence of enlarged ovaries, as the repeated manipulations in the course of the procedure to reach the follicles located at different parts of the ovary may directly lead to iatrogenic torsion especially when the ovaries are enlarged.
The final decision regarding how many embryos to transfer and what to do with any remaining embryos is made on the day of transfer. The decision is made collaboratively among you, your partner, the doctor, and the embryologist. If the embryo s survives in the IVF lab for five days after the egg retrieval, it is likely that it will have reached the blastocyst stage. At this stage we are able to transfer just one or two embryos to achieve the same pregnancy rate as transferring three embryos on day 3.
This decreases the instances of high order multiple pregnancies. For many patients, an elective single embryo transfer eSET is recommended by the fertility team to achieve the primary goal of a singleton pregnancy which is ideal for both mother and baby.
The drawback to having a transfer on day 5 is that not all embryos survive that long. The embryos that do not survive to day 5 in the lab probably would not have created a pregnancy had they been transferred on day 3, but there is no way to know this for certain. Fourteen days after egg retrieval, a blood test is performed to determine if a pregnancy resulted from this process. Spotting sometimes even heavy bleeding can occur even if you are pregnant. Please remember that the progesterone medication you are taking is vital to the pregnancy and should not be stopped or interrupted unless you are specifically told to do so by a doctor or a nurse.
Click here or call Download Brochures. Below is a more detailed description of the IVF process. It is important to remember that IVF is a step-by-step process and one step cannot begin until the previous step has been completed.
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