Fertilization process of eggs in the IVF lab | Reasons for failed fertilization with ICSI
Sai Gundeti, Senior Embryologist
Most patients naively expect that if the doctor retrieves 10 eggs, all 10 will fertilize and form embryos in the IVF lab. However, this is not true. Human reproduction is not very efficient, and there is a lot of wastage in the process. Let's look at the process of fertilization in more detail to understand why.
Eggs are normally fertilized in a woman's body within her Fallopian tubes. This event occurs within the first 24 hours after the eggs have been released from the ovaries and can occur only if there are sperm present around the eggs at this critical time. The lucky few sperms after ejaculation which reach the egg in the Fallopian tube surround it, and begin competing for entrance. The head of each sperm, the acrosome, releases enzymes that begin to break down the outer shell of the egg, called the zona. Once a single sperm has penetrated the egg, its cell membrane changes its electrical characteristics. This electrical signal causes small sacs (called cortical granules) just beneath the egg's cell membrane to dump their contents into the space surrounding the egg. The contents swell, pushing the other sperm far away from the egg in a process called the cortical reaction. The cortical reaction ensures that only one sperm fertilizes the egg because it blocks further sperm from entering the egg.
Both the sperm and the egg are gametes and contain a single set of 23 chromosomes, 1 of every type. These are said to be haploid. As a result of fertilization, the single set of 23 chromosomes from the egg combines with the single set of 23 from the sperm to create a ZYGOTE. This is still a single cell (the fertilized egg ) BUT now it has 2 sets of chromosomes, or 46 chromosomes, which is why it is called diploid ). This zygote then divides serially ( by mitosis) to produce progressively more cells, resulting in an embryo, which continues to develop into a baby if all goes well.
In the IVF lab, we basically mimic the in vivo process. The sperm is processed; and 100,000 activated sperm are added to the egg(s) per ml of culture medium in a petri dish. The sperm then perform their usual biological function - the only difference being that instead of doing this in the fallopian tube in vivo, they are now doing it in vitro! Many motile sperm reach the egg and many sperm binds to the outer shell or zona pellucida. Of these, only a single sperm enters the eggs. At that point, the sperm and egg cell membranes chemically bind and the sperm is pulled into the interior, or cytoplasm, of the egg. When the sperm enters the interior of the egg, its tail breaks off. As a result, the sperm cell membrane is broken, and this allows water to enter the sperm head from the cytoplasm of the egg. Then, the genetic material of both the egg and the sperm form a nucleus within the egg. These two nuclei are called pronuclei until they merge to form the nucleus of the new embryo.
For semen samples with low sperm count /motility or with high abnormal sperms (teratozoospermia samples (abnormal sperms more than 30%), IVF is not recommended as chances of failed fertilization is very high. For these patients, ICSI is a better option.
InICSI, a single sperm is injected into each egg on an inverted microscope. This technique eliminates the need for the sperm to dissolve a path through the shell of the egg and to bind to the egg membrane. Some sperm do not have these capabilities ( which is why they will not be able to fertilize the egg on their own). ICSI allows us to give these sperm a "piggyback ride "into the egg, to ensure that fertilization will occur. Over 70% of mature eggs will fertilize after ICSI in a good lab.
INTRA CYTOPLASMIC SPERM INJECTION
The day of egg collection is called Day 0. Approximately 18 hours after IVF or ICSI, we do a fertilization check to see how many of the eggs have fertilized. This is called Day 1.
At the time of the fertilization check, we usually see that some of the eggs have not fertilized. These eggs fail to fertilize for a variety of reasons.
Sometimes, the reason may be a lab problem. This could occur if the incubator malfunctions; or if the culture medium gets contaminated.
The commonest cause of failed fertilization after IVF is abnormal sperm. This could be because of abnormally shaped sperm; or sperm dysfunction. In some cases, this may result in the total failure of fertilization. ICSI is a very efficient way of treating this problem.
However, even after ICSI, not all the eggs will fertilize. This is because some of the eggs are immature. Mature eggs are in metaphase II ( M2). As eggs mature, they go through stages of chromosomal division called meiosis, which halves their chromosome number. The extra chromosomes are contained in a small, round body located on the side of the egg called a polar body. If an egg does not have a polar body at the time of the fertilization check, this means it is still immature, and will not allow the sperm to fertilize it.
If the embryologist is not skilled, the reason for failed fertilization after ICSI maybe technical incompetence, because he did not break the oocyte membrane with the injection pipette. An unskilled embryologist may also damage the egg and kill it during the ICSI process ( for example, by using too much force).
It's very rare to have total failed fertilization of mature eggs after ICSI in a good lab. One of the reasons for failed fertilization after ICSI is a very rare condition called globozoospermia.
If you have experienced failed fertilization in your IVF cycle, and want to know what to do next, you can ask Dr. Malpani for a free second opinion !