Cryopreservation of embryos is the process of preserving an embryo at sub-zero temperatures, generally at an embryogenesis stage corresponding to pre-implantation, that is, from fertilisation to the blastocyst stage.
Embryo cryopreservation is useful for leftover embryos after a cycle of in vitro fertilisation, as patients who fail to conceive may become pregnant using such embryos without having to go through a full IVF cycle. Or, if pregnancy occurred, they could return later for another pregnancy. Spare oocytes or embryos resulting from fertility treatments may be used for oocyte donation or embryo donation to another woman or couple, and embryos may be created, frozen and stored specifically for transfer and donation by using donor eggs and sperm.
Embryo cryopreservation is generally performed as a component of in vitro fertilization (which generally also includes ovarian hyperstimulation, egg retrieval and embryo transfer). The ovarian hyperstimulation is preferably done by using a GnRH agonist rather than human chorionic gonadotrophin (hCG) for final oocyte maturation, since it decreases the risk of ovarian hyperstimulation syndrome with no evidence of a difference in live birth rate (in contrast to fresh cycles where usage of GnRH agonist has a lower live birth rate).
The main techniques used for embryo cryopreservation are vitrification versus slow programmable freezing (SPF). Studies indicate that vitrification is superior or equal to SPF in terms of survival and implantation rates. Vitrification appears to result in decreased risk of DNA damage than slow freezing.
Direct Frozen Embryo Transfer: Embryos can be frozen by SPF in ethylene glycol freeze media and transfer directly to recipients immediately after water thawing without laboratory thawing process. The world's first crossbred bovine embryo transfer calf under tropical conditions was produced by such technique on 23 June 1996 by Dr. Binoy S Vettical of Kerala Livestock Development Board, Mattupatti
World usage data is hard to come by but it was reported in a study of 23 countries that almost 42,000 frozen human embryo transfers were performed during 2001 in Europe.
Pregnancy outcome and determinants
In current state of the art, early embryos having undergone cryopreservation implant at the same rate as equivalent fresh counterparts. The outcome from using cryopreserved embryos has uniformly been positive with no increase in birth defects or development abnormalities, also between fresh versus frozen eggs used for intracytoplasmic sperm injection (ICSI). In fact, pregnancy rates are increased following frozen embryo transfer, and perinatal outcomes are less affected, compared to embryo transfer in the same cycle as ovarian hyperstimulation was performed. The endometrium is believed to not be optimally prepared for implantation following ovarian hyperstimulation, and therefore frozen embryo transfer avails for a separate cycle to focus on optimizing the chances of successful implantation. Children born from vitrified blastocysts have significantly higher birthweight than those born from non-frozen blastocysts. For early cleavage embryos, frozen ones appear to have at least as good obstetric outcome, measured as preterm birth and low birthweight for children born after cryopreservation as compared with children born after fresh cycles.
Oocyte age, survival proportion, and number of transferred embryos are predictors of pregnancy outcome.
Pregnancies have been reported from embryos stored for 27 years. A study of more than 11,000 cryopreserved human embryos showed no significant effect of storage time on post-thaw survival for IVF or oocyte donation cycles, or for embryos frozen at the pronuclear or cleavage stages. In addition, the duration of storage had no significant effect on clinical pregnancy, miscarriage, implantation, or live birth rate, whether from IVF or oocyte donation cycles.
A study in France between 1999 and 2011 came to the result that embryo freezing before administration of gonadotoxic chemotherapy agents to females caused a delay of treatment in 34% of cases, and a live birth in 27% of surviving cases who wanted to become pregnant, with the follow-up time varying between 1 and 13 years.
The first baby born from a frozen human embryo was Zoe Leyland, on 28 March 1984 in Australia, reported by Alan Trounson. The first twins derived from frozen embryos were born in February 1985. Since then and up to 2008 it is estimated that between 350,000 and half a million IVF babies have been born from embryos frozen at a controlled rate and then stored in liquid nitrogen; additionally a few hundred births have been born from vitrified oocytes but firm figures are hard to come by.
It may be noted that Subash Mukhopadyay from Kolkata, India reported the successful cryopreservation of an eight cell embryo, storing it for 53 days, thawing and replacing it into the mother’s womb, resulting in a successful and live birth as early as 1978- a full five years before Trounson and Mohr had done so. A small publication of Mukherjee in 1978 clearly shows that Mukherjee was on the right line of thinking much before anyone else had demonstrated the successful outcome of a pregnancy following the transfer of a 8-cell frozen-thawed embryo into human subjects transferring 8-cell cryopreserved embryos.
- Youssef, Mohamed AFM; Van der Veen, Fulco; Al-Inany, Hesham G; Mochtar, Monique H; Griesinger, Georg; Nagi Mohesen, Mohamed; Aboulfoutouh, Ismail; van Wely, Madelon; Youssef, Mohamed AFM (2014). "Cochrane Database of Systematic Reviews: Gonadotropin-releasing hormone agonist versus HCG for oocyte triggering in antagonist-assisted reproductive technology". Reviews (10): CD008046. doi:10.1002/14651858.CD008046.pub4. PMID 25358904.
- Edgar, D. H.; Gook, D. A. (2012). "A critical appraisal of cryopreservation (slow cooling versus vitrification) of human oocytes and embryos". Human Reproduction Update. 18 (5): 536–554. doi:10.1093/humupd/dms016. PMID 22537859.
- Kopeika, J.; Thornhill, A.; Khalaf, Y. (2014). "The effect of cryopreservation on the genome of gametes and embryos: principles of cryobiology and critical appraisal of the evidence". Human Reproduction Update. 21 (2): 209–227. doi:10.1093/humupd/dmu063. ISSN 1355-4786. PMID 25519143.
- Cryopreservation of Embryos in Ethylene Glycol Freeze Media and Direct Transfer in Crossbred Cattle in the Tropics,9th International Congress on Biotechnology in Animal Reproduction, 2002,
- Deepika Malayalam daily Newspaper dated 23 June 1996.
- A. Nyboe Andersen, L. Gianaroli, R. Felberbaum, J. de Mouzon and K.G. Nygren. Results generated from European registers by ESHRE Assisted reproductive technology in Europe, 2001
- "Genetics & IVF Institute". Givf.com. Archived from the original on 6 December 2012. Retrieved 27 July 2009.
- Wennerholm, U. -B.; Soderstrom-Anttila, V.; Bergh, C.; Aittomaki, K.; Hazekamp, J.; Nygren, K. -G.; Selbing, A.; Loft, A. (2009). "Children born after cryopreservation of embryos or oocytes: A systematic review of outcome data". Human Reproduction. 24 (9): 2158–2172. doi:10.1093/humrep/dep125. PMID 19458318.
- Evans, J.; Hannan, N. J.; Edgell, T. A.; Vollenhoven, B. J.; Lutjen, P. J.; Osianlis, T.; Salamonsen, L. A.; Rombauts, L. J. F. (2014). "Fresh versus frozen embryo transfer: backing clinical decisions with scientific and clinical evidence". Human Reproduction Update. 20 (6): 808–821. doi:10.1093/humupd/dmu027. ISSN 1355-4786. PMID 24916455.
- Wikland M, Hardarson T, Hillensjö T, et al. (May 2010). "Obstetric outcomes after transfer of vitrified blastocysts". Hum Reprod. 25 (7): 1699–707. doi:10.1093/humrep/deq117. PMID 20472913.
- Riggs R; Mayer J; Dowling-Lacey D; Chi TF; Jones E; Oehninger S (November 2008). "Does storage time influence postthaw survival and pregnancy outcome? An analysis of 11,768 cryopreserved human embryos". Fertil. Steril. 93 (1): 109–15. doi:10.1016/j.fertnstert.2008.09.084. PMID 19027110.
- Courbiere, B.; Decanter, C.; Bringer-Deutsch, S.; Rives, N.; Mirallie, S.; Pech, J. C.; De Ziegler, D.; Carre-Pigeon, F.; May-Panloup, P.; Sifer, C.; Amice, V.; Schweitzer, T.; Porcu-Buisson, G.; Poirot, C. (2013). "Emergency IVF for embryo freezing to preserve female fertility: A French multicentre cohort study". Human Reproduction. 28 (9): 2381–8. doi:10.1093/humrep/det268. PMID 23832792.
- Extension given for embryo storage Google News. Retrieved on 11 September 2009
- AP News, February 1985
- Mukerji S., Mukherjee S., Bhattacharya, S.K. Indian J. Cryog., 1978, 3, 80.
- Kumar A. Architect of India's first test tube baby: Dr Subhas Mukerji (16 January 1931 to 19 July 1981). Current Science, 1997, V.72. N7. P. 526-531.
- Dhawale A.J. Dr Subhas Mukherjee and India's first test tube baby. PharmaTutor. 2014. V.2 N.12. P.99-106.