Autologous oocyte cryopreservation in women aged 40 and older using minimal stimulation IVF

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Autologous oocyte cryopreservation in women aged 40 and older using minimal stimulation IVF Zhang, J et al Reprod Biol Endocrinol 2015

Dr. Daiter summarizes this research article

Target:  women aged over 40 years undergoing IVF with mature MII oocyte cryopreservation (for subsequent fertilization and transfer) for reasons including

(a) have no male partner,

(b) desire to delay childbearing and preserve future fertility,

(c) may lose their ovarian function due to aging, and

(d) have a family history of premature ovarian failure (early menopause) and fear that they may lose their ovarian function at a very early age.

The number of women over 40 seeking infertility treatment has been increasing over the past decade partially due to delaying childbearing until a career is established or the desire of some couples to have a 2nd family.

Goal:  Determine the reproductive potential of minimal-mild stimulation IVF for women 40 years of age or older following mature egg freezing, thaw, fertilization of thawed oocytes with ICSI, and embryo transfer.


(1) oral contraceptive pill pre-treatment at least until adequate suppression demonstrated (around 3 weeks),

(2) Clomiphene citrate (50 mg/day) or Letrozole (2.5 mg/day) started on CD3 and continued until the day prior to ovulation trigger,

(3) gonadotropin (Bravelle and/or Menopur, Follistim, or Gonal F) low dose (75 IU/day and increased if needed to 150 IU/day) starting on CD 4-7 depending on response (ultrasound and serum E2, LH, P4),

(4) ovulation trigger with nasal GnRHa (Synarel nasal spray, 2mg/mL, Pfizer) when lead follicle >18mm diameter,

(5) oocyte retrieval and cryopreservation by vitrification of mature oocytes (using Kitazato commercial kit: oocytes aspirated with a minimal volume of HEPES culture media; oocytes placed into a micro-droplet of “Basic Solution” and allowed to acclimate for 10-30 seconds; oocytes then transferred to drops of “Equilibrium Solution” containing 7.5% (v/v) ethylene glycol + 7.5% DMSO (at room temperature for 15 minutes) for 6 minutes; oocytes then transferred to “Vitrification Solution” drop containing 15% ethylene glycol + 15% DMSO + 0.5M sucrose for a total of 50 seconds; then oocytes loaded into a Cryotop (fine and thin strip) that is quickly plunged into liquid nitrogen),

(6) oocyte thawing and fertilization by ICSI (all sperm were motile to demonstrate viability for this study, beyond this the sperm with best strict morphology were selected),

(7) embryos cultured in a single global total medium (LifeGlobal Group LLC) that contains proteins, salts, amino acids, buffer (NaHCO3), glucose, pyruvate, lactate and antibiotics (gentamycin),

(8) all embryos placed into incubators with 5% CO2 resulting in a physiologic pH of about 7.30,

(9) a single embryo was transferred in a subsequent artificially prepared cycle of oral Estrace (4mg/day orally x 10 days starting on CD3, Estrace dosage increased to 6-8mg/day if the serum estradiol concentrations were <200 pg/mL and/or the ultrasound demonstrated an endometrial thickness of <7 mm) x 7-14 days until the endometrial thickness was >7mm,

(10) Prometrium vaginal insert (200mg three times a day) was added to the Estrace once the appropriate thickness was achieved and embryo transfer was on 5th day of Prometrium for cleavage stage embryos and on the 7th day of Prometrium for blastocysts,

(11) ultrasound guided ET assured placement at 1-2 cm from the fundus.

158 women were enrolled. The average age was 43.9 +/- 0.2 (range was 40-49 years old), mean BMI was 23.6 +/- 0.4 kg/m2 (range was 17-39 kg/m2), mean basal FSH was 12.6 +/- 0.6 IU/L (range was 2-48 IU/L), mean number of days of stimulation with gonadotropins was 6.1 +/- 0.3 days, and the mean peak serum Estradiol concentration was 661 +/- 37 pg/mL.  There was a postive correlation between age and basal FSH.  There was a negative correlation between age and peak E2, number of oocytes retrieved, and number of mature MII oocytes.  There was no correlation of BMI to number of oocytes retrieved or number of mature MII oocytes.  A total of 584 eggs were retrieved, 532 eggs were mature metaphase II oocytes, 4 patients had no eggs retrieved, 485 embryos resulted from the 532 MII eggs (1.9 +/- 0.1 embryo/patient), 57 relatively good embryos were transferred, 6 clinical pregnancies (10.5% clinical pregnancy/transfer), 3 live births (5.3% live births/transfer) and 3 spontaneous abortions.

Conclusion: One cannot place an absolute value on childbearing, so the upper age limit for considering oocyte cryopreservation may vary according to individual preferences, values and finances.  Women over 40 years of age can be counseled that if she forms embryos by minimal stimulation IVF each embryo transferred has a very low chance (only 5.3%) of yielding a live birth.

Dr. Daiter’s reviews this research article

​Dr. John J. Zhang is a pioneer in Reproductive Endocrinology and Infertility, including but not limited to the field of mini-IVF and natural cycle IVF.

This important article, published in Reproductive Biology and Endocrinology (2015), investigates the reproductive potential of mini IVF for women over 40 following cryopreservation of unfertilized mature eggs, thaw, assisted fertilization with ICSI, and embryo transfer.

The findings of this study include that for the 158 women enrolled with an average age of 43.9 years;  there was a negative correlation between age and number of eggs retrieved (older ~ less eggs) and number of mature MII eggs (older ~ less mature eggs retrieved); only 4 of the 158 women (<3%) had no eggs retrieved; 57 relatively good embryos were ultimately transferred and 6 women became pregnant (3.8% of women enrolled) for a clinical pregnancy rate of 10.5% per transfer; 3 pregnancies resulted in spontaneous abortion (miscarriage) so 5.3% of transfers resulted in a live birth.

The conclusion of this study is realistic: for women over 40 desiring to preserve their unfertilized eggs using cryopreservation by vitrification for the future, if embryos (fertilized eggs) are formed following the thaw then there is a low (5.3%) chance of a live birth per embryo transferred.  This information can be used to counsel this group of patients about this procedure.

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