July 2013 - Roger muses about Nature

Biological mysteries always drew me in, so it was natural to choose a career in research with the egg as its main subject. Remember the mythological sphinx with a human head and a lion’s body in Ancient Egypt? It guarded the temple secrets. Eggs may look plain and pure, but next time pause before you smash the shell of a biological sphinx on your breakfast table!

Eggs are the largest, the rarest, and the most paradoxical cells in the body. They possess contradictory characteristics: as ancestors of all other cells they are generalists, but also arch-specialists for no other cell engages in fertilization, except of course a sperm. You might think that these biological curiosities would keep young scientists employed in some quiet, arcane corner for their whole lives, but soon after wading into eggs they get into deep water.

I should have known better when I started because there were Egg and Sperm Wars as far back as the 18^th Century. Scientists and philosophers were speculating whether it is the egg or sperm that makes the baby. For a while the Spermists were winning because no one had ever seen eggs in any mammal, let alone a human being – there was nothing like a hen’s egg. But when the tiny mammalian egg was found in 1827 the Ovists counterattacked and soon afterwards it was known that eggs must be fertilized with a sperm, so both cells are needed. This settled a conflict that had been dangling since Aristotle’s days,* and put storks out of business.

But eggs can sometimes go it alone without help from a sperm. Some animals reproduce by virgin birth (parthenogenesis), but no mammal. That we are among the exceptions was an enigma until an old colleague showed that a subset of genes (fewer than 1%) are “imprinted” to make them behave differently in the sexes. Imprinted genes in sperm carry responsibility for the placenta whereas in eggs others are needed for the embryo. So that neatly explained why we always need fathers and mothers. Perhaps sperm are from Mars and eggs from Venus.

During fertilization male genetic material is injected into an egg, and if we think of the combined genes as hardware of the future embryo, the bulky cytoplasmic goo in the egg contains the clever software. So eggs and sperm make equal genetic contributions, but eggs are more equal than sperm!

Dolly the sheep came from Venus. Looking like any other in the flock outside Edinburgh, she was the only one to nuzzle in your pocket for a treat and the only celebrity there. My colleagues named her for Dolly Parton who once said of her second most important asset, “I wanted to be the first woman to burn her bra, but it would have taken the fire department four days to put it out.” Dolly the sheep was cloned from an adult udder cell.

Her arrival in 1996 shook modern biology. I remember receiving a call from a London newspaper asking for comments about the breakthrough, but I knew nothing because the secret was so closely kept. Dolly was created by removing the genetic material from an egg which was replaced by that from the udder cell. After delivering an electric jolt to the “Frankenegg”, a new creature was conceived and time’s arrow was reversed. She was a clone of a deceased animal.

Afterwards there was a furore whether humans should ever be cloned … or had been cloned! There were a few attention-seekers raising their megaphones to claim that several cloned babies already existed around the world. We found their stories preposterous, and it was just as unbelievable that the profession did not oust those doctors.

Cloning was momentous for science because by cracking the sphinx-like egg we can peep past the door into the temple of regenerative medicine. We can hope to create to order cells of a certain type and matching a particular patient who needs organ or tissue replacement. There would be cardiac cells to patch up dead tissue after a heart attack, pancreatic islet cells to treat diabetes, retinal cells for macula degeneration, and so on throughout the body.

By an odd coincidence another hugely important breakthrough hit the stage just two years after Dolly. Human embryo stem cells were created. No other stem cell is as versatile as those from the embryo’s inner core, because they make every cell type in the body, except those specific to the placenta. These cells are harvested at the blastocyst stage from embryos donated by patients in in vitro labs. Although only a minority among 100 or so cells in a blastocyst, when released into a culture fluid and surrounded by feeder cells they grow rapidly – and almost indefinitely (movie courtesy of Nikica Zaninovic).

George Bush blocked federal dollars for this research, but progress that was hampered in the U.S. continued nonetheless elsewhere. In 2004 there was news from South Korea that after injecting human eggs with genetic material cloned “embryos” were generated. Since regenerative medicine was the aim, the clones were used to harvest stem cells. It almost seemed at the time as if the holy grail had been discovered, creating the chance to help patients who were suffering or might die without a transplant. But then we heard a whistle blowing.

The Korean research director was accused by some of his associates of making fraudulent claims. His fortunes changed precipitately, absolutely, and irreversibly when this particular f-word was voiced, and to say that the innocent helpers around him had egg on their faces is an understatement. They suffered anyway because science is the most unforgiving profession when lied to. Moreover, the tide of hope was ebbing because no other lab had been successful, and public confidence was being lost. We even wondered if cloning was a biologically impossible feat in humans.

But soon after that low point, Shinya Yamanaka announced an amazing breakthrough that offered another path, and one that avoided bitter arguments about the use of human embryos. Like the rest of us, he had been intrigued when Dolly proved the molecular goo in eggs can reprogram the genetic software of body cells. For my part, then working at McGill, we were building an inventory of all the proteins in the egg, hoping to identify those with reprogramming potential. It was like looking for needles in a haystack, but Yamanaka succeeded because he took a clever short-cut to find four molecules needed to turn back the clock of adult cells, making them think they were embryos and avoiding the controversial use of human eggs and embryos. He called the cells treated with these molecules, induced pluripotent stem cells or iPS cells, and no one was surprised when last year he received flight tickets to Stockholm for a Nobel Prize.

But were we only sitting on a see-saw, not making as much headway as we thought? Are iPS cells too close to cancer for comfort? Could stem cells develop abnormalities after a sojourn in culture? Are they as good as embryo stem cells, if those cells can be made? They can. This year we heard that scientists in Oregon have succeeded in making stem cells from human clones after a long, labyrinthine struggle. So now there are two players for the prize, with supporters on both sides.

I wonder how I will regard this post in a year or two? Progress is accelerating with hopeful news of stem cell applications announced every month. I sometimes wish I were back in the lab squeezing pipettes again, but science is becoming more of a young man’s sport than for graybeards. At least we have a long view, remembering how biology turned out to be so much more plastic than we ever thought, but knowing there are always ups as well as downs before we know whether iPS cells or embryo stem cells open the door to revolutionary medical care.

Eggs are mothers of all stem cells but it is better to avoid controversy with human embryos and hope that iPS cells will succeed. But wait a minute! iPS cells are very similar to embryo stem cells, and improvements in technology bring them ever closer. On the other side of the same coin, they could be put to the same use as embryo stem cells – which can be made into babies, with genetic modification if desired, using a technique known as tetraploid complementation. For every bright side of a coin there is often a spooky opposite in science. The egg that started us down this road is not perhaps so much like the benign Egyptian sphinx but more the treacherous Greek variety.

*Read the full story in my article in Biology of Reproduction by email request and which will be publicly accessible online from 2014. To be published in fall 2013: The Oocyte: Biology, Pathology and Technology, Edited by Alan Trounson, Roger Gosden, and Ursula Eichenlaub-Ritter. Cambridge University Press. I doubt any of my readers has an appetite for this 450 page tome.

Next Post: Dear Jean

The first picture of a baby is usually in the mother’s arms soon after delivery, and becomes pasted into an album for posterity. My mother missed out on that snapshot since I was born in austere post-war London when a box Brownie camera was a luxury my parents couldn’t afford. But nowadays the picture is taken earlier and with a much more expensive camera. My granddaughter, Pippa, was snapped by an ultrasound scanner when she was barely past the fishy stage in her mother’s womb. The next picture taken a few weeks later showed her looking like a real child, and the obstetrician could confirm everything was fine, including beating heart and nuchal transparency. Pictures reassure us all is well, even if we can’t put absolute trust in them.

The first picture can be taken even earlier nowadays – in fact, as early as can be. For most of her thirty years as an embryology lab director, Lucinda gave prospective parents of IVF children a picture of their embryos which she had taken immediately before medical staff transferred them to their Mom’s body. These were embryos that had been fertilized in a Petri dish and grown to a ball of about 8 cells in three days, or had formed a blastocyst with about a hundred cells after a slightly longer incubation. Giving these photos to patients is now a common courtesy in fertility clinics, and more and more couples are having this extraordinary preview. Last year there were over 60,000 babies born after IVF procedures in the USA alone, or 1% of all births.

For such couples (and single mothers undergoing the same treatment) pictures of their embryos probably find a temporary place of honor on a kitchen bulletin-board until the next picture at the six week ultrasound scan – or taken down if the pregnancy test proves negative. But for embryologists and physicians, pictures of embryos are records of the best-looking in the bunch which they hope stand the best chance of getting Mom pregnant. Generally one or two are chosen (sometimes three) for transfer from the half-a-dozen eggs (more or less) that were fertilized, but these pictures say so much less about the embryos’ prospects than my first picture or Pippa’s, and even ours hardly say anything about things that most matter to us – future health, vigor, brains, and beauty.

Upwards of 50% of all human embryos conceived in the IVF lab (and probably the same in the body) have abnormal sets of chromosomes, and therefore no chance of making a healthy child. It is possible to genetically screen embryos so that only the healthy ones are placed in the uterus, but although this can be helpful it is far from foolproof. Some embryos are mixtures of normal and abnormal cells, meaning that a test produces misleading results if only one cell is removed, which is all that most embryologists dare take. And then there is the unanswered question of whether certain kinds of abnormalities can correct themselves spontaneously, implying that embryos could be discarded unnecessarily.

The upshot of this uncertainty is that deciding which one to transfer is still mainly based on how they look at the time when the photo is taken. It’s a beauty contest which Machiavelli would have derided because “men in general judge more from appearances than from reality.” Unfortunately, “reality” is much too deep to plumb, and so, despite the best efforts, the embryos with the best chances of success are not always chosen.

A single snapshot at the end of a growth process that had been going on for days mostly unseen inside the lab incubator is not much to go on. We wondered if a picture is worth a thousand words, perhaps a thousand pictures would better convey the whole story.

Back in the 1990s Lucinda had already started making movies of embryos in Petri dishes, but had to build her own equipment and record them on magnetic tapes. When I joined the project some years later, digital time-lapse recorders, more advanced microscopes, and miniature incubators were available for the job. We could take a snapshot of individual embryos every few minutes, which over five days added up to over two thousand frames. Imagine how eager we were to compile them for a motion picture of the first days of human embryo development! But we didn’t anticipate surprises, accustomed as we were only to static images, but the embryos educated us.

On the first day nothing seemed to be happening, although appearances are deceptive because there was a lot of activity at the sub-microscopic level. But the next day the fertilized eggs made an almighty heave and in a strange contortion never to be repeated divided into two equal halves within their shells. They tolerate only so much fragmentation of cells at this stage, which you can see in the movie. The next division to make four cells started later in the day, and more divisions created a ball of tiny cells without any net increase in overall size. Clinics are now using customized equipment to monitor the rate of embryo development to help to choose which to transfer.

On the fifth day, healthy embryos became blastocysts, hollow balls of cells with a tiny knot at one side that make up the embryo proper. The movies revealed that blastocysts undergo cycles of swelling and collapse rather like a beating heart, although far more slowly and not from muscles contracting because no such cells exist yet. Just as a heart sound or pulse is a sign of life, the blastocyst’s beat is a sign, and probably a better token of vitality than a static picture ever gave.

We had another surprise after pressing the PAUSE button at a random frame if the embryo looked sick, its cells more opaque and less plump. Sometimes it really was unhealthy and had reached the end of the road to development, but in other cases when we pressed RESUME the embryo would shrug off its frumpy appearance to continue growing, often even turning out to be a beauty. So Plato was right for embryos too – beauty really is in the eye of the beholder. A sad-looking embryo might have been discarded or, at the very least, frozen in reserve if its fate had hung on only one picture. Besides, we already knew that, like judges scrutinizing models on a catwalk, embryologists don’t always agree on their scores for embryos. They can tell you stories about patients who produced only a very few scrawny-looking embryos which were transferred to wombs “just in case”, only to find that at least one was successful and made a bonnie baby.

What about the rest, the ones that never survived long enough to be photographed? We seldom mourn things that pass beyond the focus of our eye or camera lens, nor give a moment’s thought to the far, far greater slaughter going on day-in and day-out in the wombs of the world where most embryos never thrive. Embryos may have started out looking hopeful after fertilization and even grow some, only to perish in the dish unnamed and unknown in a Greek tragedy of their own.

Sometimes I am moved to wonder about wasteful nature. I wonder why I can watch in deep sympathy a bee writhing after it has strayed into an insecticide spray or an earthworm wriggling its last as it dries out on the pavement, while we never spare a thought for the anonymous multitudes ending their lives prematurely by being trampled physically or chemically.

I suppose it is the connection with individuals, especially at tender ages, that makes the difference. No matter how alien to the subject we feel there is a biological link and by witnessing the life and death struggle of another creature we are jolted into thinking about our own. Patrick Steptoe (1913-88), who was Bob Edwards’ clinical partner in pioneering IVF, pondered the same thoughts as I do in idle moments. I remember him playing at a conference his musical elegy, For a Dying Embryo. Steptoe_PCS Before striking a key he told us he was sad for the couple who would never have that proud picture of their offspring, but he also had some sort of feeling for the embryo in his care, wondering what it might have become and achieved. Thank goodness Pippa and have pictures so we can muse in the future how we got here.

Next Post: Stem Cell Stuff

Month: July 2013

Sperm are from Mars, Eggs from Venus

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Baby’s First Picture

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