Comparative study of the influence of dehydroepiandrosterone versus growth hormone on the improvement of ovarian reserve markers in women with diminished ovarian reserve

INTRODUCTION

The ovarian reserve reflects the woman’s pregnancy chance; it differs between individuals and, at least partially, is under genetic control operated by Fragile X mental retardation 1 gene.¹ Ovarian reserve defines both total ovarian reserve, a pool of growing follicles at different stages of maturation, and functional ovarian reserve, which is related to the size of the follicle pool and the follicle recruitment rate.² The European Society of Human Reproduction and Embryology published the Bologna criteria in 2011 in order to standardize the definition of poor ovarian response (POR).³

Dehydroepiandrosterone (DHEA), an endogenous androgen, is produced in the zona reticularis of the adrenal gland and by ovarian theca cells, and converted to testosterone and estradiol (E2) in peripheral tissues. ^{4, 5} The oral administration of DHEA before gonadotrophin stimulation to increase ovarian response in poor responder patients was proposed by. ⁶

Growth hormone (GH) regulates the effect of follicle-stimulating hormone (FSH) on granulose cells, by increasing the synthesis of Insulin-like growth factor1, augments the effect of gonadotropin on granulose and theca cells, and plays an essential role in ovarian function, including follicular development, estrogen synthesis and oocyte maturation. ^{7, 8}

This study aimed to evaluate the effects of DHEA and GH supplementation on the improvement of the ovarian reserve markers in women with history of POR in the previous In vitro fertilization (IVF) cycles. The primary outcomes of the study were the number of oocytes retrieved and the clinical pregnancy rate.

PATIENTS AND METHODS

After obtaining approval from the Institutional Ethics Committee and written informed consent from all the patients, 90 adult female with POR were included in this prospective randomized controlled trial (RCT). They were divided into 3 groups; each group included 30 patients. This study was conducted in Private Based IVF Units. All the data of patients were confidential with secret codes and private file for each patient, all given data were used for the current medical research only. Couples were counseled about the treatment protocols.

Inclusion criteria: Adult females with POR defined according to the Bolonga            criteria⁹ that included presence of at least two of the three following features; maternal age (≥ 40 years), a previous POR (≤ 3 oocytes with a conventional stimulation protocol) and an abnormal ovarian reserve tests including antral follicle count (AFC) < 5 and Anti-Müllerian hormone (AMH) < 1.1 ng/mL and / or elevated FSH on days 2 or 3 of the menstrual cycle more than 10 IU/L. All the included female had normal uterine cavity.

The exclusion criteria included body mass index (BMI) ≥ 30 mg/m², FSH >20 IU/L, endocrine or metabolic disorders, such as diabetes mellitus, thyroid disorders, and polycystic ovarian syndrome, severe endometriosis, abonormal gynecological bleeding of undetermined origin, allergy to the used medications, medical treatment with corticoids, azospermia or women with uterine malformation or abnormality.

The patients were randomized through a computer-generated randomization sequence into the three groups through using 90 sealed opaque envelopes. The patients were allocated in one of three groups (30 patients each):

Control group (HMG/GnRHant protocol) (30 patients): Women of this group received HMG/GnRH antagonist protocol.

GH group (GH/HMG/GnRHant protocol) (30 patients): Women of this group received GH/HMG/GnRH antagonist protocol. Patients received daily subcutaneous injection of 2.5mg of GH (equivalent to 7.5 IU) (Norditropin, Novo Nordisk) from day 21 of previous cycle, until the day of HCG injection.

DHEA group (DHEA/HMG/GnRHant protocol) (30 patients): Women of this group received DHEA/HMG/GnRH antagonist protocol. Patients received 25 mg of micronized DHEA, administered orally three times a day, for 12 weeks before starting the cycle of ovulation induction.

The participating females underwent full history taking, medical and gynecological examination. Transvaginal sonographic evaluation was done.

Treatment protocol:

Ovarian stimulation was started in all women in the three studied groups from the second day of menstrual cycle with 300 IU of HMG. Ovarian response monitoring was performed using serial vaginal ultrasonography and measuring serum E2 levels. When dominant follicles reached to 14 mm in mean diameter, 0.25 mg/day of GnRH antagonist was started and continued to prevent premature luteinization until the day of HCG injection in both groups. When at least two follicles with a mean diameter of 17 mm were observed, 10,000 IU HCG was injected. Endometrial thickness and serum E2 levels were measured in the day of HCG injection. Oocyte retrieval was done 34–36 h after HCG injection using a 17-gauge needle under vaginal ultrasonography guidance, and conventional IVF or intracytoplasmic sperm injection (ICSI) was performed appropriately. Embryos were transferred using a labotect catheter (labotect, Gottingen, Germany) 48–72 h after oocytes retrieval. At most three embryos were transferred in each In vitro fertilization-Embryo transfer (IVF-ET) cycle and excess embryos were cryoprecipitated. Luteal phase support was started with progesterone in oil 100 mg daily IM on the day of oocyte retrieval and continued until the documentation of fetal heart activity by ultrasound.

Quantitative β HCG was performed 14 days following embryo transfer and was considered positive if ≥ 50 IU/L. In cases with confirmed pregnancy, transvaginal sonographic was performed two weeks later to confirm the presence of an intrauterine sac with positive fetal pulsations. Pregnant cases were followed up till delivery.

Primary outcomes were the number of oocytes retrieved and the clinical pregnancy rate. Secondary outcomes were the number of fertilized oocytes, number of embryos transferred and live birth rate (LBR).

Fertilization of oocytes was defined with observation of at least one pronucleus or cleaved oocytes. Implantation rate defined by the number of gestational sacs per transferred embryos was calculated. Cycle cancellation rate, chemical pregnancy rate, ongoing pregnancy rate and early miscarriage rate were calculated. 

Cycle cancellation is identified when no embryo is transferred because of failed oocyte retrieval (no obtained oocyte on the day of ovarian puncture), or failed fertilization and/or cleavage (no obtained embryo after IVF/ICSI).Chemical pregnancy is defined as a serum beta HCG ≥ 50 IU/L, 14 days after embryos transfer. Clinical pregnancy is identified as observation of fetal heart activity by trans-vaginal ultrasonography performed 5 weeks after positive beta HCG. Ongoing pregnancy is defined as pregnancy proceeding beyond the 12th gestational week. Early miscarriage was defined as loss of pregnancy before 12 weeks of gestation. LBR was defined as the number of achieved live birth after 28 weeks of gestation.

Statistical analysis

Data were fed to the computer and analyzed using IBM SPSS software package version 20. (Armonk, NY: IBM Corp). The Kolmogorov-Smirnov test was used to verify the normality of distribution. Quantitative data were described using mean ± standard deviation and were analyzed using one way ANOVA with Post Hoc Tukey test for pair wise comparisons. Qualitative data were described using number and percent. Comparison between categorical data was performed using Chi square test. Significance of the obtained results was judged at P < 0.05.

RESULTS

A comparison between the three studied groups regarding demographic data including, age, BMI, duration of marriage and duration of infertility, and the basal levels of FSH, luteinizing hormone (LH), E2, AFC and AMH were insignificant different among the three groups (P > 0.05). Table 1

Data presented as mean ± SD. Data presented as mean ± SD.

Table 1: Basal patients' characteristics.

The mean values of the total dose of HMG (IU) and the duration of stimulation were statistically significant higher in the control group as compared to the DHEA and GH groups (P < 0.05). They were lower in the DHEA group than the GH group (P < 0.05). A Comparison between the mean values of the number of the oocytes retrieved, MII oocytes, the

fertilized oocytes and the transferred embryo were statistically significant higher in the DHEA group compared to the control and the GH groups (P < 0.05). While, there was no significant difference between the control group and the GH group (P > 0.05). Table 2

Table 2: cycle characteristics.

The cancelled cycle, the implantation rate and early miscarriage rate were statistically insignificant different among the three groups (P > 0.05). The chemical pregnancy rate, clinical pregnancy rate, ongoing pregnancy rate and LBR per cycle start or per embryo transfer were statistically significant

higher in the DHEA group compared to the control group and the GH group (P < 0.05), while they were statistically insignificant different between the control and the GH group (P > 0.05). Table 3, 4

Table 3: Reproductive outcomes.

Table 4: Reproductive outcomes/ embryo transfer.