As the 1990s picked up speed, biologists continued to identify classes of stem cells that were hidden away in an animal and to make better sense of a hierarchy that had been discernible for years. Stem cells in the early embryo gave way to others in the developing fetus—among them, those in the developing testis and ovary that perpetuated the species—which in turn gave way to rare populations of stem cells that would renew certain organs and tissues of the adult. The essential finding was that although stem cells shared the ability to make more of themselves as well as generate specialized cells, stem cells were not all alike but differed in potential. Those in the embryo had immeasurable potential; those in organs, by comparison, had lost something in translation.

The cell that sat atop this hierarchy in humans—the stem cell inside the early embryo’s inner cell mass—still lay frustratingly beyond the reach of biologists. This was the pluripotent cell that could teach developmental biologists so much. It basically made the individual and every cell therein. One might have thought that because Martin Evans and Gail Martin had blazed the trail by pulling this cell from mouse embryos in the early ’80s, its counterpart in hu-

mans would have been easy game. Researchers had pried similar cells from the embryos of hamster, mink, rabbit, pig, and cow. Yet isolating stem cells in human embryos required more finesse. Success would count on a healthy blastocyst with plenty of inner stem cells; precise timing in respect to the blastocyst’s age; and a nutritive soup plus feeder cells that would keep the stem cells happily dividing in a dish while stifling their urge to differentiate.

One scientist who kept thinking about these coveted cells, and how they might serve as printing presses of sorts for a broad spectrum of mature cells, was Ariff Bongso, a fertility specialist at Singapore’s National University Hospital. If anyone did, he had the instinct, reverence, knowledge, and resources for obtaining them.

Born in Ceylon (now Sri Lanka), the eldest son of a Christian Dutch mother and a Muslim Ceylonese father, Bongso started his career as a veterinarian specializing in animal reproduction, launched by a doctorate from Ontario Veterinary College. For a while, he investigated ways of identifying the sex of an early animal embryo. This skill, coupled with the ability to transfer an early embryo from animal to dish, where its genes were fussed over, and then back to the womb, could supply farmers with genetically superior milk-producing cows on the one hand and genetically superior beef-producing steers on the other. He also worked at improving the culture conditions that permitted an embryo to survive outside the body in the first place. As he became familiar with a wider and wider circle of embryos, ranging from cows to sheep to water buffalo, he marveled over “the notion that the common beginnings of gametes, fertilization, embryo cleavage, and growth” gave way to such an awesome diversity of species, he shares today. “The more I sank into embryology, the more it strengthened my belief that one and all are God’s creations.”

Soon Bongso made the leap into human reproduction, drawn by the great potential of in vitro fertilization [IVF], the bold new fertility treatment that bypassed low sperm counts, fallopian tube blockage, and other obstacles to pregnancy by fusing egg and sperm

“in vitro”—literally, “in the glass” of a petri dish—in stark contrast to conception’s normal setting, the fallopian tube.

The public had first been introduced to this daring procedure in the summer of 1978, two years after Bongso had gotten his doctorate. It was then that IVF’s British pioneers, Robert Edwards and Patrick Steptoe, had announced the birth of Louise Brown, the world’s first “test-tube” baby—more accurately, petri-dish baby. Angry protests had followed. In vitro fertilization’s contrived merging of egg and sperm constituted an ungodly act, an unconscionable crime, some people charged. In the eyes of fertility expediters, however, the future had arrived. “I recall many accusations against Edwards and Steptoe, and how they were playing God,” relates Bongso. “But I knew there was nothing wrong with this approach, because all the technique was doing was mixing a husband and wife’s sperm and egg in a laboratory dish. I had faith these remarks would pass and the technique would become routine.”

In Asia, Bongso was part of a small contingent that enabled IVF to become routine. In ’83, he served on the fertility team responsible for Asia’s first test-tube baby, and two years later joined the staff at Singapore’s National University Hospital to help launch a state-of-the-art IVF program. “Singapore was the only Asian country to take the lead” in IVF, its university hospital having several visionary administrators, recalls Bongso. Before long, couples were singing the praises of fertilization in a dish, which gave them the children they otherwise might never have had, and interest in the hospital’s new program “exploded,” says Bongso. It wouldn’t be long before IVF clinics the world over would similarly thrive.

For better or for worse, in vitro fertilization would radically change the public’s perception toward conception and what an embryo could bear. Mouse embryos had been made to survive in culture since the ’40s. This added twist of starting a human one in a dish, however, and then transferring it back into a mother’s uterus seemed to some observers like one more step away from God and Nature.

IVF, however, caused no hideous consequences. Apparently the light of day did not deter a sperm from burrowing into an egg cell. Apparently a petri dish’s plastic didn’t do irreparable damage to a fledgling embryo. Louise Brown and other IVF babies weren’t deformed, and the fear that when they became adults they might end up as characterless as the sterile dishes their embryos had started in proved groundless. Tens of thousands of test-tube babies later, most physicians today believe that conception in a petri dish poses no major health risk for the great majority of embryos. Two 2002 studies reported that a small percent of IVF babies may be more prone to either birth defects or low birth weight than those conceived naturally. Yet further validation of these claims is needed, with some fertility experts suggesting that if the findings do bear out, they might relate to the infertile nature of women and men who seek IVF and not the treatment itself.

Interest in the new IVF wing at Singapore’s university hospital grew so rapidly that by the early ’90s Ariff Bongso and four other specialists found themselves treating 800 patients annually. Meanwhile, Bongso, the unit’s scientific director, was trying to improve the low survival rate of IVF embryos. When transferred back into the uterus for continued growth, only five to ten percent of embryos made it to birth. Fertility clinics everywhere were encountering the same low success rate, and to improve a woman’s ability to get pregnant, a practice had sprung up. Fertility technicians had taken to creating, in vitro, a dozen or more embryos for each woman treated, whereupon usually three or more of these embryos were transferred into the womb at the same time. A woman then stood a fighting chance of having at least one embryo go the whole nine yards. It also meant that she had a greater-than-average probability of bearing multiple infants. (According to one report, an IVF mother has a twenty percent or greater chance of having more than one child.)

As for the embryos that were left over, they were frozen, which provided a potentially useful stockpile. Should the first transfer of embryos fail and no pregnancy ensue, a second and even a third

attempt could be made without having the woman undergo the uncomfortable and expensive procedure of egg collection all over again. Or, if pregnancy did occur, the unused, frozen embryos might translate into more children for the couple at a later date.

Stem cell research and the rising numbers of stored IVF embryos in the world would become so inexorably intertwined that it pays to take a modern-day look at the system that has resulted in millions of stockpiled human embryos worldwide. (Currently, the numbers of IVF embryos that, upon transfer into a woman, make it to a live birth are around twenty-five percent, although estimates vary from clinic to clinic.) Douglas Powers, the scientific director at Boston IVF, which bears the distinction of being the largest fertility clinic in the United States due to its 3,000-plus IVF procedures each year, provides this description:

“Suppose fifteen eggs are retrieved” from a woman, postulates Powers, using realistic numbers. By exposing them to sperm from the male, “we try to fertilize all of them, but usually only about eighty percent—in this case, twelve—get fertilized.” The eggs that don’t get fertilized get discarded. Of those twelve, continues Powers, “often the three best embryos are transferred into the woman on day 3 of development.” Healthier embryos are distinguished by evaluating each embryo’s shape, texture, and other criteria through the microscope. With three embryos now in the uterus, that leaves nine others. “Maybe two of those nine will have failed by then,” says Powers, and therefore are discarded. “Of the remaining seven, perhaps four will be good enough to freeze;” in which case, the remaining three will also be discarded. With so many embryos disposed of along the way, IVF may seem like a wasteful procedure. Yet, according to fertility specialists, even in the natural setting of a woman’s

reproductive tract, the first few days of human development are fraught with losses every bit as severe. An estimated two-thirds or more of egg-and-sperm unions are so abnormal that either they don’t implant in the womb or, if they do, their development is short-lived.

At some future point many couples will use at least some of the embryos they’ve stockpiled. Many others won’t. At Boston IVF, of the approximately 5,000 IVF embryos that are cryopreserved each year, roughly ten percent are left unused by couples, according to Powers. A survey published in 2003 of more than 430 fertility clinics in the United States counted nearly 400,000 IVF human embryos lying in a frozen state, a figure that will likely keep rising, since unlike in England, where a five-year storage clause limits the numbers of stored IVF embryos, in the United States surplus embryos are stored indefinitely.

Many scientists would come to believe that the mounting numbers of IVF embryos in freezers, of which a sizable number were doomed waste, would be a valuable source of stem cells. As some saw it, these beginning conceptuses were not even full-fledged embryos and yet they had the gift of Life inside them.

In 1991, Ariff Bongso boosted the sustainability of IVF embryos by growing them on a bed of fallopian tube cells, just as if they were sequestered in the fallopian tube, their home before descending into the uterus. Fertility specialists were generally grasping the fact that if they could mimic an embryo’s natural environment in a petri dish, it would lead to healthier in vitro embryos. The particular approach that Bongso hit upon, he reports, gave IVF embryos a much better chance of reaching blastocyst stage on day 5, and, when transferred to the womb, of making it to a live birth.

In the process of growing fertilized eggs up to blastocysts,

Bongso’s thoughts would instinctively turn to the inestimable power of the few dozen cells that lay in a heap inside the fluid-filled cavity of an early blastocyst. From these specks, each one being hundreds of times smaller than an ant’s eye, entire civilizations had sprung. From these specks would sprout his clients’ sons and daughters, whom he would run into years later on the streets of Singapore. Bongso maintains that his mixed Christian-Muslim descent made him acutely aware of the remarkable events of early human development, particularly the fertilized egg’s power of procreation. “I grew up with the Bible and Koran side by side,” he relates, “and reading the teachings of these two books in English, I found revelation science. In the Old Testament and the Koran it is described very well that the sperm clings on to the egg; and it’s also stated that the embryo clings to the uterus. Clings—I was taken by that word,” and left with a deep regard for the potential brought about by the interlocking of egg and sperm, embryo and womb. Now, years later, confronted with pluripotent stem cells inside blastocysts and so aware of all they represented, he was left to wonder—what can I do with these special cells to make Life better?

One day in early 1993, says Bongso, “It came to me in a flash.” If he could isolate these unique human cells and sustain their division in culture, maybe he could control their fate. He would then have a gallery of cells with which to treat sick people. He raced to the library to do a search to see whether other groups had isolated human embryonic cells and also to collect clues about how to go inside the human blastocyst’s balloon-like inner cell mass. Bongso’s idea was “so powerful,” the magazine Asiaweek later described, “that it seemed to come from God’s own whisper.”

Overseas and in the scientific literature, there were signs that people were slowly waking up to this idea of using stem cells collected from early embryos to regenerate the body’s tissues. Having young embryos in dishes drew scientists’ attention to an embryo’s inner cell mass almost in a hypnotic way, making them ponder the pluripotent cells that sat there. For Bongso, because of the Seeds of

Life parables he was raised on and his study of animal reproduction ever since, “It was common sense that one should be able to direct stem cells into desirable lineages,” he recounts: kidney cells for kidney disease; brain cells for neurologic disorders; pancreas cells for pancreatic ills. His hospital’s active IVF program was a terrific resource. Thousands of embryos were stored away in liquid nitrogen tanks, many of them destined to languish in these cold beds and never be used. Bongso also had going for him the fact that Singapore’s Ministry of Health tolerated human embryo research so long as researchers followed stringent guidelines. This policy was the envy of scientists in other countries, such as Australia and the United States, where research on human embryos was hampered by funding restrictions.

One of the Ministry’s caveats was that if spare IVF embryos were to be used in research, they had to be less than fourteen days old. Up through day 13, it seemed safe to say, a human embryo has no capacity for feeling pain or any other sensation, for only on or around day 14 does the primitive streak start to form. Literally a narrow strip of migrating cells, this streak moves from one end of the embryo toward the other end like a creeping river, sending cells of the future skeleton and gut inside the embryo, and initiating the formation of the head and brain at the anterior end of the streak, explains Lewis Wolpert, a prominent British developmental biologist. Only at this stage, which is known as gastrulation, does the embryo begin to obtain the rudiments of a nervous system as well as begin to find shape. Some scientists believe that the primitive streak’s onset represents such an important starting point in the development of human life that prior to day 14 the growing clump of cells can’t be considered an embryo but instead should be looked upon as a “pre-embryo.”

As a famous saying by Lewis Wolpert goes, “It is not birth, marriage, or death, but gastrulation that is the most important event in your life.” Wolpert made this comment to “a doctor at a meeting in Belgium who clearly irritated me,” he mentions in an email. He

admits his remark may have been “a little exaggeration—but even so….” While Wolpert extols gastrulation’s importance in development, he’s of the opinion that the embryo still has miles to go before becoming a person. “The embryo is not a person until it can survive almost unaided outside the mother,” as he sees it.

After receiving approval for his project from National University Hospital’s ethics committee, Bongso obtained twenty-one spare IVF embryos from nine patients who, having no further plans for them, were glad to donate them to the Sri Lankan’s research. Bongso, meanwhile, was comfortable with the Singapore law he operated under; he, also, saw the primitive streak’s onset on day 14 as the very first hint of a human being. As he grew IVF embryos up to day-5 blastocyst stage, then bathed them in an enzyme that dissolved their thin outer coat so that their inner stem cells would be let loose, he worked with the knowledge that these spare embryos probably would have been incinerated at some point in time. Why not, therefore, try to squeeze some benefit from them? Once he put the coatless balls in culture together with feeder cells, he essentially had cells from both the blastocyst’s inner cell mass and outer cell layer in culture.

But all too soon came a big disappointment. The Singapore scientist had hoped that his culture broth would keep the cells multiplying, without differentiating into mature cells. To succeed for medicine’s sake, he had to establish a stem cell line that would grow and grow. Explains Bongso, “You want those cells to produce millions and billions of cells, to increase their numbers for later differentiation into desirable tissues for treating patients.” Yet quite maddeningly, although many of his cells lasted for two passages, they would spontaneously differentiate into mature fibroblasts. He and his labmates tried this, and they tried that, yet they couldn’t hold the cells in check. Differentiation spelled the end of the line for a stem cell; it stopped being a stem cell and lost the magic to churn out specialized cell types.

To the best of their knowledge, nonetheless, Bongso and his crew had carved a new benchmark. Reporting their work in the journal

Human Reproduction later that year, they stated that this was the first time anyone had successfully isolated cells from the inner cell mass of “late preimplantation human embryos”—day 6 and day 7 human embryos that are about to implant in the uterus—and kept them alive in vitro for two divisions.

Had they really isolated bona fide embryonic stem cells, as Bongso today claims? Other researchers would say no; the cells probably were not stem cells, but more likely cells that had begun to differentiate and take a more specialized form. As other scientists were also finding out, it was relatively simple to separate cells from human blastocysts, but whether they were at peak pluripotency was another matter. Bongso ran a battery of tests on the cells but was unable to verify that they were supremely pluripotent, which is why he referred to them as “stem-cell-like” in his paper. Now he wishes that instead of rushing to publish his account, he had “patiently worked out the obstacles to crossing the second passage,” he conveys.

Bongso, nevertheless—by keeping human embryonic stem cells dividing in a dish for any time at all—had managed to cross The Great Divide.