Intracytoplasmic sperm injection
|Intracytoplasmic sperm injection|
Oocyte is injected during ICSI
Intracytoplasmic sperm injection (ICSI, pronounced //, IK-see) is an in vitro fertilization (IVF) procedure in which a single sperm cell is injected directly into the cytoplasm of an egg. This technique is used in order to prepare the gametes for the obtention of embryos that may be transferred to a maternal uterus. With this method acrosome reaction is skipped.
There are several differences within classic IVF and ICSI. However, the steps to be followed before and after insemination are the same. In terms of insemination, ICSI needs one only sperm cell per oocyte, meanwhile IVF needs between 50 and 100 thousands. This is due to the fact that in IVF acrosome reaction has to take place and thousands of sperm cells have to be involved. Once fertilized, the egg is transformed into a proembryo and it has to be transferred to the uterus to continue its development.
Defective sperm function remains the single most important cause of human infertility. Although certain severe forms of male infertility have a genetic origin, others may be the result of environmental factors. During the past decade, ICSI has been applied increasingly around the world to alleviate problems of severe male infertility in human patients who either could not be assisted by conventional IVF procedures or could not be accepted for IVF because too few motile and morphologically normal sperm were present in the ejaculate of the male partner.. Low sperm count, poor quality of sperms and inability to fertilize the egg are some of the factors for which ICSI is suggested.
The first human pregnancy generated by ICS was carried out in 1991, by the hand of Gianpiero Palermo and his team.
This procedure is most commonly used to overcome male infertility problems, although it may also be used where eggs cannot easily be penetrated by sperm, and occasionally in addition to sperm donation.
It can be used in teratozoospermia, because once the egg is fertilized, abnormal sperm morphology does not appear to influence blastocyst development or blastocyst morphology. Even with severe teratozoospermia, microscopy can still detect the few sperm cells that have a "normal" morphology, allowing for optimal success rate.
The first child born from a gamete micromanipulation (technique in which special tools and inverted microscopes are used that help embryologists to choose and pick an individual sperm for ICSI IVF) was a child in Singapore-born in April 1989.
The procedure itself was first performed in 1987, though it only went to the pronuclear stage. The first activated embryo by ICSI was produced in 1990, but the first successful birth by ICSI took place on January 14, 1992 after an April 1991 conception.
There are some requirements that an IVF lab need to correctly perform ICSI technique. There are basically 5 main elements: laminar flow cabinet, inverted microscope, micromanipulators, microinjectors and anti-vibration table.
- Laminar flow cabinet. It has to contain a hot surface and an environment with 5% of CO2.
- Inverted microscope. It should have 20X and 40X lens, with Hoffman optics, and a 37 °C surface to have an optimal maintenance of the biological material.
- Micromanipulators. They allow us to move in three dimensions, so that there will be one on the left to control the holding of the micropipette and another to the right for the injection micropipette. In addition, there is an electric manipulator for large movements and an hydraulic one for the finest (adjustment of the pipettes is done with the hydraulic and quick movements to place them with the electric).
- Microinjectors. They are hermetic syringes filled with mineral oil controlled by the micromanipulators and connected to the microinjection pipettes (to aspirate and inject the spermatozoa) by a flexible tube.
- Anti-vibration table. It has to contain hydrogen to assure the optimal work conditions.
ICSI is generally performed following a transvaginal oocyte retrieval procedure to extract one to several oocytes from a woman.
In ICSI IVF, the male partner or a donor provides a sperm sample on the same day when the eggs are collected. The sample is checked in the lab, and if no sperm is present, doctors will extract sperm from the epididymis or testicle. The extraction of sperm from epididymis is also known as percutaneous epididymal sperm aspiration (PESA) and extraction of sperm from testicle is also known as testicular sperm aspiration (TESA).
The procedure is done under a microscope using multiple micromanipulation devices (micromanipulator, microinjectors and micropipettes). A holding pipette stabilizes the mature oocyte with gentle suction applied by a microinjector. From the opposite side a thin, hollow glass micropipette is used to collect a single sperm, having immobilised it by cutting its tail with the point of the micropipette. The oocyte is pierced through the oolemma and the sperm is directed into the inner part of the oocyte (cytoplasm). The sperm is then released into the oocyte. The pictured oocyte has an extruded polar body at about 12 o'clock indicating its maturity. The polar body is positioned at the 12 or 6 o'clock position, to ensure that the inserted micropipette does not disrupt the spindle inside the egg. After the procedure, the oocyte will be placed into cell culture and checked on the following day for signs of fertilization.
In contrast, in natural fertilization sperm compete and when the first sperm penetrates the oolemma, the oolemma hardens to block the entry of any other sperm. Concern has been raised that in ICSI this sperm selection process is bypassed and the sperm is selected by the embryologist without any specific testing. However, in mid-2006 the FDA cleared a device that allows embryologists to select mature sperm for ICSI based on sperm binding to hyaluronan, the main constituent of the gel layer (cumulus oophorus) surrounding the oocyte. The device provides microscopic droplets of hyaluronan hydrogel attached to the culture dish. The embryologist places the prepared sperm on the microdot, selects and captures sperm that bind to the dot. Basic research on the maturation of sperm shows that hyaluronan-binding sperm are more mature and show fewer DNA strand breaks and significantly lower levels of aneuploidy than the sperm population from which they were selected. A brand name for one such sperm selection device is PICSI. A recent clinical trial showed a sharp reduction in miscarriage with embryos derived from PICSI sperm selection.
'Washed' or 'unwashed' sperm may be used in the process.
Ultra-high magnification sperm injection (IMSI) has no evidence of increased live birth or miscarriage rates compared to standard ICSI.
Assisted Zona hatching (AH)
Patients with repeated failed implantation or who have a thick zona pellucida (covering around the embryo) are ideal candidates for Assisted Zona Hatching or AH. The procedure involves creating a hole in the zona, that improves the chances of normal implantation of the embryo in the uterus.
Pre-implantation genetic diagnosis (PGD)
PGD is a process in which one or two cells from an embryo on Day 3 or Day 5 are extracted and the cells genetically analyzed. Couples who are at a high risk of having abnormal number of chromosomes or who have an history of single gene defects or chromosome defects are ideal candidates for this procedure. It is used to diagnose a large number of genetic defects at present.
Success or failure factors
One of the areas in which sperm injection can be useful is vasectomy reversal. However, potential factors that may influence pregnancy rates (and live birth rates) in ICSI include level of DNA fragmentation as measured e.g. by comet assay, advanced maternal age and semen quality.
There is some suggestion that birth defects are increased with the use of IVF in general, and ICSI specifically, though different studies show contradictory results. In a summary position paper, the Practice Committee of the American Society of Reproductive Medicine has said it considers ICSI safe and effective therapy for male factor infertility, but may carry an increased risk for the transmission of selected genetic abnormalities to offspring, either through the procedure itself or through the increased inherent risk of such abnormalities in parents undergoing the procedure.
There is not enough evidence to say that ICSI procedures are safe in females with hepatitis B in regard to vertical transmission to the offspring, since the puncture of the oocyte can potentially avail for vertical transmission to the offspring.
Follow-up on fetus
In addition to regular prenatal care, prenatal aneuploidy screening based on maternal age, nuchal translucency scan and biomarkers is appropriate. However, biomarkers seem to be altered for pregnancies resulting from ICSI, causing a higher false-positive rate. Correction factors have been developed and should be used when screening for Down syndrome in singleton pregnancies after ICSI, but in twin pregnancies such correction factors have not been fully elucidated. In vanishing twin pregnancies with a second gestational sac with a dead fetus, first trimester screening should be based solely on the maternal age and the nuchal translucency scan as biomarkers are significantly altered in these cases.
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