In Vitro Fertilization (IVF) with or without assisted fertilization (ICSI) involves reproducing the first week of embryonic development in the laboratory, outside the female genital tract (in Vitro), from the fertilization stage to the blastocyst stage before hatching.
This technique is now over 40 years old, since the first birth of a child conceived by IVF was obtained in 1978 by Robert Edwards’ team in Great Britain.
1 OVARIAN STIMULATION
Unlike treatments for intrauterine insemination, where the aim is to obtain follicular development close to physiology, ovarian stimulation in IVF protocols aims to obtain simultaneous development of several follicles on each ovary.
To achieve this, higher doses of gonadotropin hormones (FSH ) are used (between 150 and 300 IU per day), from the start of the menstrual cycle, to “recruit” several follicles at the same time on each ovary. During this stimulation, spontaneous ovulation must be prevented by blocking the pituitary gland with either GnRH agonists or antagonists.
There are several protocols for ovulation stimulation for IVF, sometimes with small variations specific to each team.
For the sake of clarity, I have chosen to present the protocols I use most often in my current practice.
a) LONG AGONIST protocol
C’est le protocole le plus ancien, les agonistes de la GnRH ayant précédés de nombreuses années l’arrivée des antagonistes de la GnRH dans la panoplie des traitements d’AMP.
This is the oldest protocol, GnRH agonists having preceded by many years the arrival of GnRH antagonists in the panoply of MPA treatments.The principle is to desensitize the pituitary gland prior to ovulation stimulation, by injections of GnRH agonists. After about ten days of treatment, the anterior pituitary gland is put to rest, enabling ovulation to be stimulated without fear of premature ovulation.
In most cases, injections are started in the second half of the cycle before the actual stimulation begins, between days 20 and 23.
When the new menstrual period arrives, we check that the pituitary gland is properly blocked, using hormone assays, and that there are no ovarian cysts, using a pelvic ultrasound performed at the very beginning of the cycle.
Ovarian stimulation will then begin at the start of the new cycle, with daily injections of gonadotropins.
There are two phases in this protocol:
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- The pituitary desensitization phase, lasting around ten days, with daily injections of GnRH agonists.
- The ovarian stimulation phase, during which gonadotropin injections are added to the GnRH agonist injections, so there are two daily injections for a dozen days.
The total duration of this treatment is around 3 weeks, hence the name “long protocol”.
b) Short Agonist Protocol
Classically reserved for patients with low ovarian reserve, this protocol simultaneously uses low-dose GnRH agonists and high-dose gonadotropins.
The start of stimulation will be preceded by ovarian blockade with estrogen-progestin pills to “synchronize” the small follicles and obtain homogeneous growth of the small recruitable follicles.
The total duration of treatment is generally around twelve days.
c) ANTAGONIST protocol
GnRH antagonists began to be used in the late 1990s. Unlike GnRH antagonists, their mode of action is immediate, and pituitary blockade is achieved within hours of injection.
In antagonist protocols, ovulation stimulation begins at the start of the menstrual cycle, with the antagonist being introduced in the second part of the treatment around the 6th or 7th day of stimulation.
There are therefore two phases in this protocol:
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- The ovarian stimulation phase, lasting around twelve days, with daily injections of gonadotropins.
- The pituitary blocking phase, which begins around the 6th-7th day of ovarian stimulation. In addition to gonadotropin injections, there are daily injections of antagonists. There will therefore be two daily injections during the second part of the treatment for 5-6 days.
2 TREATMENT MONITORING
Regardless of the stimulation protocol used, monitoring is the same and relies on blood tests (estradiol, LH and progesterone) coupled with pelvic ultrasound performed vaginally with an empty bladder.
During each scan, the number and size of growing follicles are studied. On average, 3 ultrasounds are performed over a period of a dozen days.
After each scan, new treatment instructions are given.
3 OVULATION INDUCTION
Induction of ovulation enables the final stages of oocyte maturation to take place, rendering the oocyte fertile, and the scheduling of oocyte collection by ovarian puncture before in vivo ovulation occurs in the pelvic cavity.
In most cases, ovulation is triggered by subcutaneous injection of recombinant hCG, with oocyte collection taking place the day after injection.
When there is a risk of ovarian hyperstimulation (estradiol > 4000 pg/ml, number of follicles greater than 20, history of ovarian hyperstimulation), ovulation can be triggered by injection of GnRH agonists.
4 OOCYTE COLLECTION – OVARIAN PUNCTURE
Ovarian puncture enables oocytes to be collected before they are ovulated. The procedure is carried out in the operating theatre, under ultrasound control, with a brief general anaesthetic of around 10 minutes, without intubation. The patient is perfused and placed in the gynaecological position on the examination table. The bladder is emptied using a urinary catheter. The ultrasound probe, fitted with a needle guide, is positioned in the vagina. A needle connected to a continuous suction system is introduced through the needle guide. Under ultrasound control, each follicle is punctured via the Trans vaginal route. The follicular fluid collected is maintained in tubes at a constant temperature of 37 degrees. Both ovaries are punctured. Only small follicles are not punctured. The tubes containing the follicular fluid are immediately sent to the laboratory.
At the end of the procedure, the operator checks that there is no vaginal bleeding.
The patient is then monitored in the recovery room until she is fully awake.
5 SPERM COLLECTION
On the morning of the ovarian puncture, the partner will come to the AMP center to collect sperm. The abstinence period should be 2 or 3 days maximum. It is essential to bring identification with you. In some cases, a sperm sample frozen in the laboratory may be used.
6 FERTILIZATION – EMBRYO CULTURE
Once the ovarian puncture has been completed, the tubes containing the follicular fluid are transferred to the laboratory. The oocytes are isolated and placed in drops of culture medium. They are then placed in contact with a sperm preparation (IVF) or micro-injected (ICSI).
The oocytes are then placed in incubators for embryo development.
This short video illustrates the stages of embryonic development from day 3 to day 5, at the blastocyst stage.
