Document Type : Original Article
Authors
1 4Department of Reproductive Imaging, Reproductive Biomedicine Research Center, Royan Institute for Reproductive Biomedicine, ACECR, Tehran, Iran
2 Department of Endocrinology and Female Infertility, Reproductive Biomedicine Research Center, Royan Institute for Reproductive Biomedicine, ACECR, Tehran, Iran
3 5Department of Midwifery and Reproductive Health, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran
4 6Department of Biostatistics and Epidemiology, School of Medicine, Babol University of Medical Sciences, Babol, Iran
5 Department of Endocrinology and Female Infertility, Reproductive Biomedicine Research Center, Royan Institute for Reproductive Biomedicine, ACECR, Tehran, Iran;Department of Obstetrics and Gynaecology, Tehran University of Medical
Abstract
Keywords
Successful implantation of an embryo requires a receptive endometrium, a good quality embryo, and embryoendometrial synchronization in all species (
Endometrial receptivity during the implantation window is often assessed by ultrasonography markers such as
endometrial thickness, echogenic pattern, blood flow, and
biochemical markers (
Despite significant advances in ovarian stimulation protocols, treatment of repeated, unresponsive thin endometrium is still a challenge in assisted reproductive cycles,
which usually results in cycle cancellation or repeated implantation failures (
There is a growing body of evidence that the endometrial growth has a relationship with the state of uterine
blood flow (
Sildenafil citrate (Viagra®, Pfizer, NY, USA) is a 5-phosphodiestrase inhibitor that increases smooth muscle relaxation and vasodilation by preventing cGMP breakdown
(
It has been reported that vaginal sildenafil significantly
reduced peripheral natural killer cell (NK-cell) activity
and improved successful pregnancy rates in women with
histories of recurrent miscarriages. Although the mechanism of influence by sildenafil on natural killer cell activity is unclear, it seems that enhancement of uterine artery
flow has an effective influence on the local endometrial
NK-cell population (
This comparative pilot study evaluated the effect of vaginal sildenafil suppositories on endometrial proliferation and IVF outcome in infertile patients with a history of repeated IVF failure.
This phase II randomized, double-blind, placebo-controlled trial was performed at Royan Institute, Reproductive Biomedicine Research Center, Tehran, Iran, between February 14, 2014 and November 14, 2016 (NCT03192709). The assessors and patients were not aware of the treatment allocated. The Institutional Review Board and Ethics Committee of Royan Institute, Tehran, Iran reviewed and approved this study in compliance with the Declaration of Helsinki (EC/88/1045). Informed consent was obtained from all patients prior to their participation in the study.
The study population consisted of 66 infertile women aged ≤38 years. The inclusion criteria was met when the women had normal ovarian reserve [blood anti-mullerian hormone (AMH) levels >1.5 ng/mL] with at least two prior cycles with follicle stimulating hormone (FSH) <10 mIU/ml; a history of two prior consecutive failed IVF/ ICSI attempts with at least a transfer of two good quality fresh or frozen-thawed embryos; hCG day endometrial thickness <7 mm in all prior IVF/ICSI attempts; and normal endometrial appearance according to either hysterosonography, hysterosalpingography, or hysteroscopy. Women were excluded if they had a history of myomectomy or Asherman’s syndrome.
A preliminary colour Doppler transvaginal sonography
with a 4-8 MHz probe (ProSound Alpha 10; Aloka, Japan) was performed by an expert radiologist on day 14
of the patient’s prior menstrual cycle to investigate the
uterine and adenexes for any abnormal findings. The endometrial parameters of endometrial thickness, endometrial pattern, pulsatility index (PI), and resistance index
(RI) were measured. The uterine artery PI and RI were
obtained through flow velocity waveforms from the ascending branch of the uterine artery at the point near to
the internal cervical orifice and calculated as previously
described (
In the IVF treatment cycle, the patients were randomly assigned to three groups according to a random allocation sequence generated by a randomized block design. The size of each block was 3. In group A, the sildenafil (vaginal suppositories, 100 mg/day, Parnian Daroo, Co., Tehran, Iran) were administered from the first day of the FSH injection until the day of oocyte retrieval. In group B, placebo (vaginal suppositories, Parnian Daroo, Co., Tehran, Iran) was initiated from the first day of the HMG injection until 2 days before the hCG injection, after which sildenafil vaginal suppositories were initiated and continued until the day of oocyte retrieval. In group C, the placebo was given from the first day of HMG injection until the day of oocyte retrieval. Participants received 10 000 IU of hCG (Choriomon, IBSA, Switzerland) when at least two dominant follicles were 18 mm in diameter. Endometrial thickness, pattern, PI and RI were measured on the day of hCG administration and compared with the data obtained in the previous cycle without sildenafil citrate treatment. Oocytes were retrieved 36 hours later via transvaginal ultrasound-guided needle aspiration.
Embryo transfer was performed after 48 hours of oocyte retrieval. Progesterone in oil (100 mg, IM daily) or intravaginal progesterone (400 mg, twice daily) was used for luteal support and maintained until the pregnancy test was conducted. Serum hCG levels were measured on the 14th day following oocyte retrieval. Vaginal ultrasound confirmation of pregnancy was performed at 4-6 weeks after embryo transfer.
A chemical pregnancy was determined by a positive
ß-hCG test result. A clinical pregnancy was confirmed
by ultrasonography visualization of one or more gestational sacs or definitive clinical signs of pregnancy.
The spontaneous abortion was defined as a pregnancy loss of an intrauterine pregnancy before 22 weeks’ gestation (
Suppositories containing 100 mg of sildenafil were
prepared from the oral tablets by a local pharmacy (Parnian Daroo, Iran). We defined the endometrial thickness
threshold cut-off of <7 mm as a thin endometrium based
on other studies (
Data are expressed as mean ± standard error (SE) and proportion. Continuous and categorical outcome variables were compared between three intervention groups by one-way analysis of variance (ANOVA) and the chi-square test. All statistical analyses were performed using SPSS version 22 for Windows 7 (IBM Analytics, Armonk, NY). The significance level was set at 0.05.
During the recruitment process, we enrolled 66 patients and allocated 22 patients to each study group. A
total of 10 patients were lost to follow up for measuring clinical pregnancy for the following reasons: all
of the embryos of their treatment cycle were cryopreserved (n=3), they had no oocytes at retrieval (n=2),
or no embryos to transfer (n=5). The flow diagram explicitly shows the number of participants at the beginning, intervention allocation, follow-up, and analysis
(
Table 1 shows the baseline characteristics of the three
study groups. There were no significant differences
among the study groups. The majority of patients had
normal endometrial patterns, which were similar between the three study groups (
Of note, the embryo transfer days were similar between the three groups. All of the embryos were transferred either two or three days after ovum pickup.
The endometrial thickness and patterns after the interventions were not statistically different between the
study groups. Additionally, the three intervention groups
were not different in left and right uterine artery PI and
RI. Implantation rate was not statistically significant
over the three groups (P=0.290). Clinical pregnancy
rates were 33.3 (sildenafil), 33.3 (sildenafil+placebo),
and 17.6 (placebo) as seen in Table 2. Although the
clinical pregnancy rates varied among the intervention
groups, they were not statistically different (
Flowchart of participants in this randomized controlled trials. IVF/ICSI;
Comparison of baseline characteristics and cycle related factors between the sildenafil, sildenafil + placebo, and placebo groups prior to intervention
Variable | Sildenafil n=22 | Sildenafil+placebo n=22 | Placebo n=22 | P value | |
---|---|---|---|---|---|
Age (Y) | 33.2 ± 4.6 | 31.7 ± 4.8 | 32.8 ± 4.6 | 0.568 | |
BMI (kg/m2) | 24.7 ± 3.7 | 26.2 ± 3.6 | 25.2 ± 2.9 | 0.379 | |
Infertility duration | 8.8 ± 4.9 | 10.5 ± 5.1 | 8 ± 4.1 | 0.220 | |
Infertility type | |||||
Primary | 19 (86.4) | 20 (90.9) | 17 (77.3) | ||
Secondary | 3 (13.6) | 2 (9.1) | 5 (22.7) | 0.438 | |
Infertility reason | |||||
Tubal factor | 1 (4.5) | 1 (4.5) | 1 (4.5) | ||
Male factor | 14 (63.6) | 11 (50) | 11 (50) | ||
Endometriosis | 0 (0) | 0 (0) | 1 (4.5) | 0.910 | |
Unexplained | 1 (4.5) | 2 (9.1) | 1 (4.5) | ||
Two or more | 6 (27.3) | 8 (36.4) | 8 (36.4) | ||
Endometrial pattern | |||||
Normal | 18 (81.8) | 18 (81.8) | 19 (86.4) | ||
Heterogenic | 2 (9.1) | 3 (13.6) | 3 (13.6) | 0.683 | |
Ecogene | 2 (9.1) | 1 (4.5) | 0 | ||
Uterine artery PI | |||||
Right | 2.4 ± 0.7 | 2.5 ± 0.7 | 2.9 ± 0.9 | 0.134 | |
Left | 2.5 ± 0.8 | 2.7± 0.9 | 2.9 ± 1 | 0.440 | |
Uterine artery RI | |||||
Right | 80.1 ± 19.3 | 69.7 ± 32.7 | 74.2 ± 31.2 | 0.474 | |
Left | 76.5 ± 25.2 | 66.4 ± 35.7 | 74 ± 31 | 0.527 | |
Endometrial thickness | 8.0 ± 2.4 | 8.9 ± 2.0 | 7.6 ± 2.1 | 0.146 | |
Type of gonadotropins | |||||
FSH (75 IU/mL) | 5 (22.7) | 8 (36.4) | 3 (13.6) | 0.209 | |
FSH+LH (75 IU/mL) | 17 (77.3) | 14 (63.6) | 19 (86.4) | ||
Ovulation duration | 9.9 ± 2.1 | 10.3 ± 2.2 | 9.1 ± 1.3 | 0.100 | |
Ampoules (n) | 9.1 ± 12.2 | 9.6 ± 14.6 | 7.8 ± 8.5 | 0.871 | |
Oocytes (n) | 11.5 ± 5.6 | 11.6 ± 6.7 | 8.1 ± 5.5 | 0.098 | |
MII (n) | 9.3 ± 5.1 | 9.4 ± 5.8 | 6.3 ± 4.1 | 0.079 | |
BMI; Body mass index, PI; Pulsatility index, FSH; Follicle stimulating hormone, LH; Luteinizing hormone, RI; Resistance index, and MII; Mature metaphase II.
Comparison of treatment cycle outcomes between the sildenafil, sildenafil+placebo, and placebo groups after intervention
Variable | Sildenafil n=22 | Sildenafil+placebo n=22 | Placebo n=22 | P value | |
---|---|---|---|---|---|
Endometrial pattern | |||||
Normal | 17 (77.3) | 21 (95.5) | 20 (90.9) | ||
Heterogenic | 3 (13.6) | 0 | 2 (9.1) | 0.263 | |
Ecogene | 2 (9.1) | 1 (4.5) | 0 | ||
Uterine artery PI | |||||
Right | 2.1 ± 0.6 | 2.2 ± 0.7 | 2.0 ± 0.5 | 0.515 | |
Left | 2.2 ± 0.4 | 2.3 ± 0.7 | 2.1 ± 0.3 | 0.357 | |
Uterine artery RI | |||||
Right | 79.8 ± 7.0 | 82.3 ± 5.9 | 80.0 ± 5.0 | 0.318 | |
Left | 78.2 ± 18.1 | 79.0 ± 18.2 | 81.5 ± 4.0 | 0.747 | |
Endometrial thickness | 10.1 ± 2.4 | 10.30 ± 2.5 | 9.60 ± 2.5 | 0.656 | |
Embryo (n) | 4.8 ± 3.3 | 4.80 ± 2.7 | 3.60 ± 3.2 | 0.322 | |
ET (n) | 2.5 ± 1.2 | 2.20 ± 1.4 | 2.10 ± 1.6 | 0.603 | |
ET grade | |||||
A | 2.1 ± 2.0 | 2.13 ± 2.3 | 2.14 ± 2.2 | >0.999 | |
B | 1.0 ± 1.0 | 1.0 ± 1.1 | 1.0 ± 1.1 | 0.957 | |
C | 0.4 ± 0.8 | 0.10 ± 0.3 | 0.40 ± 1.0 | 0.555 | |
PI; Pulsatility index, RI; Resistance index, and ET; Embryo transfer.
Comparison of reproductive outcomes between the sildenafil, sildenafil+placebo, and placebo groups after intervention
Variable | Sildenafil n=21 | Sildenafil+placebo n=18 | Placebo n=17 | P value | |
---|---|---|---|---|---|
Chemical pregnancy | 7/21 (33.3) | 6/18 (33.3) | 3/17 (17.6) | 0.490 | |
Clinical pregnancy | 4/21 (19.0) | 6/18 (33.3) | 3/17 (17.6) | 0.464 | |
Implantation rate | 5/56 (8.9) | 8/51 (15.7) | 3/47 (6.4) | 0.290 | |
Miscarriage rate | 0 | 1 (16.7) | 0 | 0.562 | |
Data are presented as n (%).
The importance of the endometrial pattern as a predictor
of treatment cycle outcome in IVF-treated patients is
well-documented (
Several treatment modalities have been offered to
patients with ‘‘thin’’ endometrium, including hormonal
manipulation by oestrogen and gonadotropin therapy,
low-dose hCG, tamoxifen, L-arginine or sildenafil,
vitamin E, pentoxifylline, low-dose aspirin, hysteroscopic
adhesiolysis, intrauterine infusion of growth factor such
as G-CSF, and the recent application of regenerative
medicine. Despite the large variety of treatment,
most options lead to only minor modifications in the
endometrium thickness and subsequent pregnancy,
and when this modality fails, patients are eventually
candidates for surrogacy. Treatment of thin endometrium
remains a challenge and future investigations are required
to further clarify and ideally manage patients with thin
endometrium (
In the present study, the clinical and chemical
pregnancy rates in the two groups of women who were
taking sildenafil alone and the women who took placebo and sildenafil showed a twofold increase compared to
the placebo-only-treated women, which according to the
sample size, this increase is not significant. The results
reported by Dehghani Firouzabadi et al. (
In this study, the clinical pregnancy rate was higher in
the sildenafil and sildenafil+placebo groups than in the
placebo group. Although this increase was not statistically
significant, it was clinically shown to be twofold. These
results were consistent with the findings of AbdelKader
Fahmy et al. (
The endometrial receptivity is an important stage
in the ART cycles, the results of our study showed
a higher rate of implantation in the women who took
sildenafil+placebo than in the other two groups, which
was consistent with the results of Dehghani Firouzabadi
et al. (
In the present study, 2 out of 7 cases of pregnancy
ended in abortion in the sildenafil group, 1 out of 6 cases
of pregnancy in the sildenafil+placebo group, and 0 of
3 cases of pregnancy in the placebo group. The rate of
abortion was negligible in the sildenafil group. Follow-up
evaluations showed a molar pregnancy in the sildenafil
group. Dzieciol et al. (
In our study, higher numbers of MII oocytes were
retrieved in the sildenafil groups compared to the
placebo group, which was clinically important; however,
one-way ANOVA results showed that this finding was
not statistically significant. To our knowledge, there
is only one study (abstract available) by Vidal et al.
(
Vaginal sildenafil might conceivably improve chemical and clinical pregnancy rates in repeated IVF failure patients. Since this was a pilot study, we recommend that clinical trials should be conducted with vaginal or oral sildenafil on larger numbers of these patients.