In Vitro Fertilization (IVF) involves fertilizing an egg with sperm outside the body. A Frozen Embryo Transfer (FET) is a subsequent procedure where a cryopreserved embryo is thawed and placed into the uterus. When only one embryo is transferred, the expectation is a single birth. However, a frozen embryo can split after transfer, resulting in identical twins. This division of a single fertilized egg is known as monozygotic twinning and occurs after the embryo has been placed back into the uterine environment.
The Increased Rate of Identical Twinning in Frozen Embryo Transfer
The rate of identical twinning following a Single Embryo Transfer (SET) procedure is notably higher than in natural conception. In the general population, monozygotic twins occur in 0.4% to 0.5% of live births. Conversely, studies tracking outcomes after SET report an incidence of monozygotic twinning (MZT) ranging from 1.5% to over 2% of live births, a rate two to five times greater than spontaneous conception.
When a single embryo is transferred, any resulting twin pregnancy confirms the twins are genetically identical, as it must be the result of the embryo splitting. Analysis of single embryo transfer cycles indicated that using frozen-thawed embryos increased the risk of zygotic splitting by 34% compared to fresh embryo transfers. The splitting event most often occurs after the embryo has developed to the blastocyst stage, typically five to six days after fertilization.
Biological Mechanisms Behind Embryo Splitting
The elevated incidence of monozygotic twinning in frozen embryo cycles is believed to stem from the physical and biological stresses imposed by the assisted reproductive technology process itself. One of the leading theories centers on the stress induced by cryopreservation, the process of freezing and thawing the embryo. The rapid changes in temperature and the introduction of cryoprotectants can cause cellular stress that may compromise the integrity of the early embryonic cells, predisposing the embryo to divide after implantation.
Another factor involves the embryo’s protective outer shell, known as the zona pellucida. Cryopreservation techniques can sometimes cause this shell to harden, making it more difficult for the embryo to naturally escape, or “hatch,” before implantation. Micromanipulation procedures commonly performed in IVF, such as assisted hatching or embryo biopsy for preimplantation genetic testing, are also recognized as contributing factors. The increased pressure and cellular rearrangement required for an embryo to emerge from a compromised or manipulated zona pellucida may encourage the inner cell mass to separate into two distinct groups, initiating the twin process.
The practice of culturing embryos to the blastocyst stage (Day 5 or 6) before transfer is also associated with an increased risk of splitting. Extended culture periods, combined with the delayed timing of implantation common in FET cycles, allow more time for the cellular division that leads to twinning. Studies have shown that blastocyst culture increases the risk of zygotic splitting. The cumulative effect of cryopreservation, extended culture, and potential micromanipulation creates an environment that encourages the single embryo to split.
Medical Management Following Monozygotic Twinning
Once a twin pregnancy is confirmed following a single embryo transfer, the initial step in medical management is to determine the chorionicity and amnionicity via ultrasound, which dictates the level of subsequent monitoring. Chorionicity refers to the number of placentas, and amnionicity refers to the number of amniotic sacs. The majority of monozygotic twins resulting from a single blastocyst transfer are monochorionic diamniotic, meaning they share a single placenta but have separate amniotic sacs.
Monochorionic (MC) pregnancies carry greater risks than dichorionic (DC) pregnancies, which have two separate placentas. The shared placenta in MC twins can lead to unequal blood flow between the fetuses through vascular connections, resulting in complications such as Twin-to-Twin Transfusion Syndrome (TTTS), selective fetal growth restriction (sFGR), and the potential for a single-twin demise.
To mitigate these risks, MC twin pregnancies require intensive surveillance. Guidelines recommend serial ultrasonographic monitoring, often starting as early as 16 weeks of gestation, to facilitate the early detection of complications like TTTS. Patients carrying MC twins are often referred to specialized maternal-fetal medicine centers for expert care and timely interventions, such as fetal laser surgery for TTTS.