Boon of fertility

British physiologist Robert Edwards with Louise Brown (far right, carrying her son), the world's first test tube baby, and her mother, Lesley Brown, at the 30th anniversary celebrations of Bourn Hall, the fertility clinic he co-founded in Cambridge. A 2008 photograph.-REUTERS British physiologist Robert Edwards with Louise Brown (far right, carrying her son), the world's first test tube baby, and her mother, Lesley Brown, at the 30th anniversary celebrations of Bourn Hall, the fertility clinic he co-founded in Cambridge. A 2008 photograph.

The Nobel Prize in Medicine goes to Robert G. Edwards for developing the technique of in vitro fertilisation three decades ago.

IT is estimated that about four million individuals have so far been born through in vitro fertilisation (IVF), the medical technique that assists infertile couples to conceive through the fertilisation of the human egg into a viable embryo outside the uterus ( in vitro as against in vivo), popularly known as test tube baby. Many of these couples normal adults, and some have already become parents through normal sexual reproduction. The Nobel Prize in Physiology or Medicine 2010 has been awarded to the 85-year-old Robert G. Edwards, currently Professor Emeritus at the University of Cambridge, for developing this revolutionary IVF therapy over three decades ago.

Edwards' pioneering research into the biology of the human fertilisation process that began in the 1950s and finally succeeded on July 25, 1978, with the birth of Louise Joy Brown, has resulted in a medical advance that marks a paradigm shift in the treatment of many types of infertility affecting more than 10 per cent of all couples worldwide. According to the Indian Council of Medical Research (ICMR), around 15-20 million couples suffer from infertility in India alone. The first Indian IVF baby, though mired in controversy, came into the world just 67 days after the global first, and today it is estimated that IVF therapy is carried out over about a total of 30-40,000 (menstrual) cycles of patients a year in the country.

Research into IVF is not new. It, in fact, began in the mid-19th century in non-mammalian species. In 1935, Gregory Pincus showed, using the rabbit as the animal model, the experimental conditions necessary for mammalian oocytes (immature ova, or germ cells, or egg cells) to mature in vitro and reach the metaphase stage of meiosis (the appropriate developmental stage in cell division necessary for sexual reproduction). In 1959, Min Chueh Chang showed that in vitro-matured rabbit oocytes could be fertilised in vitro and also give rise to viable embryos. But the conditions used were not entirely in vitro because it was wrongly believed then that sperm required in vivo activation before being transferred to the petri dish for fertilisation. However, in 1963, Chang and Ryuzo Yanagimachi identified experimental conditions by which spermatozoa from hamsters could fertilise oocytes without prior in vivo activation and give rise to two-cell-stage embryos.

In the first half of the 20th century, researchers were also trying to define conditions suitable for the fertilisation of human oocytes in vitro. However, despite advances made in animal reproductive research, no significant progress could be achieved on the human front until the early 1960s. Basically, the following steps were involved in achieving IVF in humans:

The ability to control the oocyte maturation process;

The ability to retrieve oocytes at a developmental stage suitable for IVF;

The ability to activate sperm in vitro;

The ability to define conditions that would promote fertilisation as well as early embryo development in vitro; and

The ability to transfer early embryos back to the mother's uterus.

As early as the 1950s, Edwards, working at the National Institute for Medical Research in London, had realised the potential of IVF as a treatment for infertility. His research over many years in animal reproductive biology, in particular the mouse, was helpful in achieving the goal that he became committed to. Edwards made a number of fundamental discoveries. He clarified how human eggs mature, how different hormones regulated their maturation and at which point in time the eggs were susceptible to fertilisation. After several years of work, Edwards also succeeded, in 1965, in finding the right conditions that activated the dormant and immature egg cells in vitro and promoted their maturation. Contrary to the prevalent view from earlier research in mammalian IVF that maturation required only a few hours of cultivation in vitro, Edwards found that human oocytes required 24 hours of incubation before the maturation process began. He also found that this prolonged cultivation resulted in egg cells at a late developmental stage, which were suitable for IVF.

In his next challenge to find suitable conditions for the fertilisation of human oocytes in vitro, the possibility of which was already suggested by the Chang-Yanagimachi work on hamster eggs, Edwards was assisted by the work of his student Barry Bavister at Cambridge University, who in 1969 had identified the buffer conditions to support in vitro activation of hamster oocytes. Edwards used the same buffer conditions and showed that human spermatozoa thus activated could also promote the fertilisation of in vitro-matured oocytes. This discovery marked an important milestone in the development of treatment for infertility in humans.

This success, however, came with a serious limitation. The matured and fertilised eggs could not progress beyond the two-cell division stage. With his insight, Edwards correctly attributed this to the lengthy periods that the matured egg cells had to spend outside the body. So he decided to use oocytes that had completed their maturation process in vivo and hypothesised that if oocytes could be retrieved from the ovaries just before the onset of ovulation they would be more competent in early embryo development and thus useful for IVF treatment. Edwards also knew from his earlier work on mouse reproductive biology that the initiation of meiotic maturation of the egg cells could actually be controlled by externally provided gonadotropins, hormones that mimic the function of the internally acting leutenising hormone (LH). His in vitro studies on human oocytes also gave him an insight into the timing of the meiotic maturation process.

The new strategy of Edwards (who had now moved to Cambridge University), however, raised another important technical problem how to retrieve an adequate number of oocytes from the ovary at the correct stage of development. In the late 1960s, oocytes could be accessed only through the surgical removal of a small part of the ovary from infertile women. Edwards realised that this was not suitable for IVF. However, he got to know about the new technique of laparoscopy being pioneered by Patrick Steptoe of Oldham and District General Hospital that allowed the human female reproductive tract to be seen through a fibre-optic endoscope inserted through an incision near the navel. Steptoe had also succeeded in aspiring oocytes from the ovary by this method. Edwards immediately realised that this method could be employed to retrieve oocytes from the ovary at the appropriate developmental stage during a suitable period of the menstrual cycle. In 1970, Edwards and Steptoe demonstrated that mature pre-ovulatory oocytes could indeed be retrieved after priming ovaries with gonadotropins.

Using the above technique, Edwards and Steptoe demonstrated IVF of pre-ovulatory oocytes with in vitro-activated sperm that resulted in eight-cell-stage human embryos. This seminal finding was reported in Nature in 1970. This meant that in vitro-activated sperm were capable of embryo development beyond the two-cell stage, and for the first time, it was also shown that it was possible for human embryos to undergo cell division in vitro. A year later, Edwards and Steptoe further demonstrated that IVF human oocytes could undergo further cell division to form 16-cell-stage embryos and in vitro blastocysts (thin-walled hollow structures that form during a distinctive stage mammalian embryogenesis, which contain a cluster of cells with already differentiated cell types).

These findings set the stage for the next important phase in IVF development by Edwards and colleagues. Interestingly, despite this promising accomplishment, in 1971, the United Kingdom's Medical Research Council decided not to fund the IVF project of Edwards and Steptoe, chiefly on the basis of the criticisms by the medical fraternity on the safety of the embryos. Fortunately, the work was sustained through private donation.

When, in the early 1970s, Edwards and Steptoe transferred the early IVF embryos back into women, all of over hundred attempts by them resulted in short-lived pregnancies that ended in spontaneous abortions. Suspecting that the hormone treatment administered to induce oocyte maturation disturbed the implantation of embryos in the uterus, they altered the hormone treatment protocol. This resulted in the first successful pregnancy in 1976, but the embryo had implanted ectopically (outside the uterine cavity) in the fallopian tube and the pregnancy had to be terminated.

These events led to Edwards and Steptoe abandoning the hormone treatment altogether and instead relying on the natural menstrual cycle of the patients. This, however, meant that they could access only one egg a cycle. On the basis of the concentration of LH in the urine of the women, they could predict when the appropriate developmental stage of the oocyte would occur and then try to retrieve the egg before ovulation using laparoscopy. Their efforts were rewarded in 1978, and in a letter to the journal Lancet, they announced the historic achievement of the successful birth of a normal, healthy baby on July 25, 1978, after a full-term pregnancy through IVF of a human oocyte. The result of Edwards' nearly three decades of persistence, commitment and belief was the opening up of a new era in the treatment of infertility.

Following this success, Edwards and Steptoe founded an infertility clinic at Bourn Hall in Cambridge, where they developed the IVF technique further. The Bourn Hall Clinic quickly came to be known as a centre for IVF research. Rapid technical development, as a result of this emerging field in other parts of the world, too followed the achievements of Edwards' team. The laparoscopic recovery gave way to the ultrasound-guided oocyte recovery method and the cryopreservation of surplus human embryos (see also box). In 1994, successful IVF of in vitro-matured human oocytes was also reported. The development of the technique of intracytoplasmic sperm injection (ICSI) in which a single sperm is microinjected into the cytoplasm of a mature egg marked an important breakthrough that has enabled many categories of male infertility to be treated as well.

Edwards' work has been instrumental in later work leading to the derivation of human embryonic stem cells, which has been important for the understanding of cellular differentiation and is fast emerging as a medical therapy. The IVF method has also led to the development of pre-implantation genetic diagnosis, an in vitro procedure on fertilised early embryo cells that reduces the risk of genetic disorders or chromosomal abnormalities being transmitted to the offspring.

Ethical issues naturally arise when there is any interference with a natural biological process. Realising the importance of ethical considerations in IVF treatment, Edwards himself had initiated a debate on many of the issues that would arise in assisted reproduction through a key paper that he co-authored with a lawyer, David Sharpe. They argued for strict ethical guidelines for research and treatment using human germ cells and embryos. Edwards set an example by creating an ethics committee at Bourn Hall.

According to several follow-up studies, children born following IVF are, in general, as healthy as children born through normal conception. In general, 20-30 per cent of fertilised eggs lead to the birth of a child in IVF. But there is also some evidence of complications resulting from IVF, such as multiple births, an increased risk of major malformations and a twofold increased risk of a preterm birth, and a slightly increased frequency of two rare genomic imprinting disorders.

But most of these occur because of improper adherence to treatment protocols and ethical regulations, such as the practice of transferring two or more embryos into the uterus, and perhaps also because of the generally older age of women undergoing IVF. However, today IVF is an established therapy. The discovery of the technique by Edwards, followed by several improvements on it, marks a major medical advance that is a boon to infertile people all over the world.