Assisted Hatching: An Additional Step to Embryo Implantation

An embryo has a thick protein shell known as the zona pellucida (ZP). This structure has several functions during fertilization but it breaks apart after fertilization to be able to attach tightly to the uterus.

During in vitro fertilization, eggs are collected from the egg donor, fertilized with sperm under lab conditions and then implanted into the female partner’s uterus. Immediately following implantation to the uterus, the embryo in the blastocyst stage glides out of the zona pellucida through an occasional gap in the wall of the membrane so that it can successfully attach to the uterus. However, it doesn’t always occur naturally, and eventually lead to implantation failure.

What is Zona Pellucida?

The mature egg has a 13–15 μm thick outer membrane called the zona pellucida that protects it from the its surroundings. Between days 5 and 7 of development, the embryo breaks apart this layer in order to implant attach itself in the endometrium. This atatchment is called implantation. Thus breaking of zona pellucida in the right time plays important role on successfull pregnancy.

This membrane have multiple protective functions, including preventing the entry of multiple spermatozoa into the egg, controlling the first cell divisions of the embryo (the zona pellucida ensures that these cells remain together until day 4 and do not disintegrate), and preventing the embryo from implanting itself too soon.

What is Assisted Hatching?

Assisted Hatching (AH) is an additional step during in vitro fertilization treatment, where this zona pellucida is let to hatch by mechanical, chemical means or laser to increase the chances of successful embryo implantation. It is generally performed on the day 3 of embryo development.

Although it is not advised to every woman undergoing IVF treatment, several studies have shown that it may increase the chances of successful pregnancy.

According toa study published in Journal of Assisted Reproduction and Genetics, in spite of recent developments in IVF and ICSI procedures, the rate of implantation of the replacement embryos is still quite low, and it is believed that up to 85% of the replaced embryos do not implant. And assisted hatching may increase both the pregnancy and implantation rates.

Who is Advised for Assisted Hatching?

The following are the scenarios in which this method is utilized the most frequently:

• Women above maternal age because zona pellucida tends to get thickened by age

• Women with naturally thick zona pellucida

• Slow and disorganized embryonic development or cell division.

• Previous implantation failures

• When embryos have been cryopreserved, there is a higher risk of the zona pellucida becoming rigid. This is especially true in situations where the embryos have been frozen for an extended period of time.

Assisted Hatching is not always carried out on every patient. Only in certain circumstances is it recommended to do so. Therefore, in order to be able to recommend this procedure, the unique aspects of each patient should be evaluated and thorough diagnosis should be carried out in order to determine whether or not the embryo requires assistance for the successful implantation.

Methods for Assisted Hatching

Assisted hatching is typically performed on the third day after fertilization, when the embryo typically has about 8 cells and has not yet grown in size. This is because at this stage, the embryo is still relatively small.

There are several techniques to help an egg to hatch, including mechanical incision on the surface; chemical zona drilling with acid tyrode's medium; chemical zona thinning; laser-assistance and piezo vibrator manipulators.

1. Mechanical Hatching: The procedure uses partial zona dissection (PZD) to generate an artificial opening in the zona pellucida of early cleaved embryos, allowing them to hatch through the artificial opening. Although PZD is a fast process, the holes it creates can be of varying sizes, which isn't always desirable.

2. Chemical Hatching

   1. Zona Drilling: Applying Tyrode's acid in extremely dilute concentrations to the zona pellucida eventually causes the shell to crack.

   2. Zona Thinning: It entails using the Tyrode's acid biaxially to create a cruciate area of thinning around one-third of the circle of the zona pellucida without causing an opening.

3. Laser Assistance: Laser assisted microdissection of the ZP uses an infrared 1.48-μm diode laser to thin the zona pellucida without creating a hole. It has no deleterious effects on in vitro embryo development and can be conducted with great accuracy and consistency. Laser aided approach appears to have the lowest possible risk linked to it, and it is reasonably straightforward to implement with consistency between operators, despite the fact that the equipment may be expensive.

4. Piezo Technique: In this method, vibratory motion of a needle produced by a piezo-micromanipulator is utilized to carve a confined conical area in the zona pellucida.

Potential risk of Assisted Hatching

Embryo hatching is a delicate step for the embryo to attach to the endometrium safely. This is why assisted hatching is not advised to every woman who is applying to assisted reproductive technologies (ART) because this technique can also pose a threat to the embryo if not conducted carefully. Possible risks of AH include:

• During the hatching process, blastomeres, which are cells of an embryo at the cleavage stage, may become stuck in the tiny pores of the zona pellucida, and as a result, the embryo may not successfully finish the hatching process.

• On the contrary, blastomeres may be lost if holes are larger and junctions between blastomeres haven't been completed. This may cause monozygotic twins or even embryonic death.

How Assisted Hatching Leads to Multiple Pregnancies?

In vitro fertilization may increase the chances of having dizygotic pregnancies because generally more than one embryo is implanted to secure the successful pregnancy. This possibility can increase a little bit more if assisted hatching is applied. However, assisted hatching usually ends up with monozygotic twins, also called identical twins.

The higher incidence of monozygotic twins following assisted hatching may occur for two reasons:

1. The inner cell mass of an egg can expectedly divide because of theinsufficiently tight opening made by mechanical means on the zona pellucida.

2. A blastomere can break free before they create their tight junctions to form a single fetus. This can occur due to early hatching, or big holes on ZP. A released blastomere begins to divide on its own and forms another individual fetus, giving rise to monozygotic twinning.