Document Type : Research Article
Authors
1 Yazd Research and Clinical Center for Infertility, Shahid Sadoughi University of Medical Sciences, Yazd, Iran
2 Department of Obstetrics and Gynecology, Faculty of Medicine, Bushehr University of Medical Sciences, Bushehr, Iran
Abstract
Keywords
Cryopreserved-thawed embryo transfer began
in 1983 and became a popular, vital component of
assisted reproduction technology (
Various protocols (gonadotropin/GnRH agonists,
clomiphene citrate, or exogenous estrogen
and progesterone) have been discussed in literature
reviews with regards to the endometrium
preparation for frozen-thawed embryo transfer (
Because the natural cycle protocol does not require
exogenous hormones‚ it is favored by many
patients (
There is an idea that the endogenous production
of progesterone is enough to support
implantation in a natural cycle. However, an
inadequate progesterone level at the time of
implantation or during early pregnancy may
happen naturally due to luteal phase deficiency
(LPD), which can result in infertility or abortion
(
The reported frequency of LPD ranges from
3.7% to 20% among infertile patients (
The study was designed as a prospective randomized
clinical trial. A total of 102 women
each underwent an embryo transfer in a natural
cycle in Yazd Research and Clinical Center for
Infertility affiliated by Shahid Sadoughi University
of Medical Sciences, from March 2011
to March 2012. This study was approved by the
Ethics Committee of Yazd Research and Clinical
Center for Infertility. Prior to starting the
study‚ an informed consent was signed by each
couple. The inclusion criteria were: cryop reserved
embryos after conventional
Patients were randomized to either group in a ratio of 1:1 by means of computer-generated random numbers on the day of participation. Group selection and randomization were performed by a nurse not involved in the study, by using opaque sealed envelopes. Both the patients and the clinicians were aware of the allocated arm.
Of the initial 109 women invited to participate,
102 were included in the study. All women
had previously undergone IVF or ICSI with
embryo cryopreservation. They were randomly
allocated to either the progesterone (n=51) or
the no-progesterone (n=51) groups. In the progesterone
group, we excluded four women. One
patient had an endometrial polyp and three patients
had thin endometria. Similarly, three patients
were excluded from the no-progesterone
group because of endometrial polyps (
The progesterone group received 100mg/day of progesterone (Aburaihan Pharmaceutical Co., Tehran, Iran) IM, that began 36 hours after the hCG administration and continued until ten weeks of gestation if pregnancy occurred. Control patients received no progesterone. In both groups, cryopreserved embryo transfer was performed with a Cook catheter (Cook Ireland Ltd.) five days after hCG administration. Serum β-hCG level was measured 14 days after the transfer.
Morphology of fresh cleavage-stage embryos
was evaluated according to the number of blastomeres
and degree of fragmentation. Embryo
selection for transfer or freezing was performed
in the morning of the transfer day. Embryos
were considered suitable for freezing if they
had <30% fragmentation. Cryopreservation of
all embryos was undertaken with vitrification
by the cryotop method on day 3 of pre implantation
development in both groups. After two-step
loading with equilibration solution that contained
ethylene glycol and dimethyl sulfoxide
and a vitrification solution that contained ethylene
glycol, dimethyl sulfoxide and sucrose,
a narrow glass capillary was used to load the
embryos onto the cryotop. After loading, the
majority of the solution was removed to leave
only a thin layer that covered the embryos, after
which the sample was quickly immersed into
liquid nitrogen. Subsequently, the plastic cap
was pulled over the film part of the cryotop and
the sample stored in liquid nitrogen. At warming,
the protective cap was removed from the
cryotop while it was still submerged in liquid
nitrogen and the cryotop was immersed directly
into a 37˚C medium that contained sucrose.
Next, the embryos were sequentially incubated
in diluent solution before further
The main outcome measures concerned clinical pregnancy and implantation rates. Chemical pregnancy was defined as serum β-hCG>50 IU/L at 14 days after the embryo transfer. Clinical pregnancy was defined as the presence of a gestational sac with heart beat identified by ultrasound 4-5 weeks after the embryo transfer. Implantation rate was defined as the ratio of gestational sacs to the number of embryos transferred. Clinical abortion rate was determined as clinically recognized pregnancy losses before 20 weeks of gestation.
The SPSS 19 package program was used to perform all statistical analyses. The normality of distribution of variables was tested by the Kolmogorov- Smirnov test. Independent sample t test was used for continuous variables which were normally distributed and Mann-Whitney U test for data not normally distributed. Chi-square or Fisher exact tests were used for qualitative variables as appropriate. A p value <0.05 was considered statistically significant. The data are presented as the mean ± standard deviation unless otherwise indicated.
There were no significant differences noted in the fertilization rate between study and control groups (55.4% vs. 64.3%; p=0.16). Of the 102 patients included in this study, 51 received progesterone and the other 51 did not. Table 1 describes the basic characteristics of the patients in the two groups. The demographic parameters were similar in both groups in terms of age, basal FSH levels‚ body mass index (BMI)‚ the number of previous cycles‚ etiology of infertility, and infertility duration. Table 2 compares the previous fresh cycle characteristics in the two groups.
The mean number of oocytes retrieved‚ mean number of mature oocytes and the number of embryos obtained and vitrified did not differ between the groups. There were no significant differences noted in the fertilization rate (55.4% vs. 64.3%; p=0.16). In addition, the previous stimulation protocols and fertilization procedures were similar in the two groups. Only ejaculated sperms had used for conventional IVF or intracytoplasmic sperm injection and percent of sperms with progressive motility and sperms with normal morphology‚ also sperm count were not different in those groups. There was no significant difference observed between the groups regarding the reasons for embryo freezing. Table 3 compares the cycle characteristics of the two groups. Endometrial thickness and estradiol‚ progesterone and LH levels on the day of hCG administration were similar between groups.
The cycle length until the day of hCG administration‚ number of embryos transferred, and the number of good-quality embryos did not differ in the two groups. Table 4 presents a comparison of the pregnancy outcomes of the study groups. Again, no statistically significant differences were observed in the clinical pregnancy rate between the groups (33.3% vs.27.5%, p=0.66). Although there was a trend toward an increased clinical pregnancy rate with luteal supplementation‚ the difference was not significant. There were no differences between the implantation rates (16.6% vs. 15.3%‚ p=0.93) or clinical abortion rates (11.8% vs.14.3%‚ p=0.83). The flowchart of the study is shown in figure 1.
Characteristics of patients
Outcome variable | Progesterone N=51 | No progesterone N=51 | P value |
---|---|---|---|
29.0 ± 3.8 | 28.7 ± 4.6 | 0.71 | |
23.8 ± 2.8 | 24.3 ± 2.4 | 0.35 | |
6.0 ± 3.8 | 6.7 ± 4.5 | 0.71 | |
5.8 ± 1.9 | 6.0 ± 2.0 | 0.90 | |
14 (27.5) | 17 (33.3) | 0.51 | |
0.62 | |||
35 (68.6) | 32 (62.7) | ||
7 (13.8) | 6 (11.8) | ||
9 (17.6) | 13 (25.5) | ||
Patients’ previous fresh cycle characteristics
Outcome variable | Progesterone N=51 | No progesterone N=51 | P value |
---|---|---|---|
0.84 | |||
29 (56.9) | 31(60.8) | ||
22 (43.1) | 20 (39.2) | ||
11 (21.6) | 19 (37.3) | ||
40 (78.4) | 32 (62.7) | ||
10.0 ± 4.3 | 9.6 ± 3.4 | 0.16 | |
8.3 ± 3.4 | 7.6 ± 2.8 | 0.22 | |
6.2 ± 1.6 | 5.7 ± 2.2 | 0.19 | |
4.3 ± 1.0 | 4.0 ± 0.6 | 0.07 | |
55.4 | 64.3 | 0.16 | |
12.6 ± 7.7 | 11.9 ± 7.0 | 0.62 | |
15.0 ± 5.8 | 14.5 ± 6.9 | 0.72 | |
15.3 ± 9.8 | 14.2 ± 7.3 | 0.51 | |
0.59 | |||
30 (58.8) | 26 (45.1) | ||
19 (37.3) | 21(41.2) | ||
2 (3.9) | 4 (7.8) | ||
Frozen-thawed embryo replacement cycle characteristics
Outcome variable | Progesterone N=51 | No progesterone N=51 | P value |
---|---|---|---|
8.7 ± 1.3 | 8.9 ± 1.4 | 0.64 | |
208.4 ± 60.2 median: 200 | 196.9 ± 85.3 median: 170 | 0.11 | |
0.77 ± 0.09 | 0.80 ± 0.07 | 0.08 | |
4.9 ± 1.9 | 4.6 ± 1.7 | 0.39 | |
14.3 ± 1.8 | 13.7 ± 1.5 | 0.07 | |
1.7 ± 0.5 median: 2 | 1.9 ± 0.5 median: 2 | 0.07 | |
54.9 | 60.8 | 0.54 | |
Pregnancy outcomes
Outcome variable | Progesterone N=51 | No progesterone N=51 | P value |
---|---|---|---|
18 (35.3) | 16 (31.4) | 0.83 | |
17 (33.3) | 14 (27.5) | 0.66 | |
16.6 | 15.3 | 0.93 | |
2 (11.8) | 2 (14.3) | 0.83 | |
Flowchart of study patients
The granulosa cells of the developing follicle generate
estradiol in response to gonadotropin stimulation
in natural cycles. The endometrium acquires
receptivity to embryo implantation by responding
to progesterone action on an appropriately primed
endometrium. Estrogenic stimulation would result in
endometrial proliferation and the induction of progesterone
receptors. The endometrium undertakes
profound conformational and biochemical changes,
from proliferative to secretory, with a concomitant
induction of endometrial receptivity and opening of
the window of implantation in response to progesterone
(
The transfer of frozen-thawed embryos has important
implications for the management of women undergoing
ovarian hyperstimulation for IVF (
Our hypothesis was that progesterone support has a
beneficial effect on pregnancy rate after frozen embryo
transfer in natural cycles, but the results did not support
our hypothesis. In our study, hCG was used for final oocyte
maturation. It was suggested that hCG administered
for the final oocyte maturation in stimulated IVF cycles
would cause a luteal phase defect by suppressing LH
production through a short-loop feedback mechanism
(
Bourgain et al. have reported that progesterone induces
a secretory transformation of the endometrium in
the luteal phase (
In contrast to our study, Bjuresten et al. have reported
that progesterone supplementation improved the live
birth rate after embryo transfer in natural cycles (
Our study was in agreement with a study by Kyrou et
al. that reported luteal phase support did not affect ongoing
pregnancy rates in natural hCG-induced frozenthawed
embryo transfer cycles (
There emerged no significant differences between the two groups in our study with regards to the implantation or clinical pregnancy rates‚ but there was a trend toward an increased clinical pregnancy rate with luteal supplementation. Thus, further studies are needed to confirm our findings.