The Morph Man

David Gardiner doesn’t want to come off like a nerdy guy at a science fiction-fantasy convention, even though, as he sits in his UC Irvine office surrounded by dozens of frog tchotchkes and pictures of salamanders, there is a distinctly Harry Potter-esque atmosphere around him. But when he says that biology is about to give human beings the power to grow limbs the way some amphibians can generate new legs, isn’t he crossing over into the realm of science fiction?

As an academic biologist, Gardiner knows how ridiculous this sounds. So he points out that scientists have been contemplating human regeneration for about 200 years: “Somebody will figure this out, but I suspect it will not be us.”

But this isn’t what he really thinks. He thinks it inevitable that before his career is over, humans will regrow limbs, and he wants to make it happen. “Maybe my parents read too many fairy tales to me when I was a kid,” he says, adding later: “Deep down inside I want it to be me.”

Gardiner’s optimism symbolizes the growing sense among a small band of scientists working in a formerly obscure branch of biology that for the first time since 1768, when an Italian monk named Lazzaro Spallanzani observed that amphibians could regrow body parts, there are strong reasons to believe that humans can do it too.

The U.S. Department of Defense has in part validated Gardiner’s view by injecting money into regeneration research. The Defense Advanced Research Projects Agency (DARPA), the government agency that has recently received a wave of attention with its offbeat experiments, has mounted a preliminary “seedling” project called ReGenesis to investigate the possibilities. Starting in early 2003, DARPA doled out about half a million dollars to scientists. Gardiner, who was unaware of the project, was not one of them, but he has since been working with grant recipients and hopes to receive funding if a full-blown program is approved. He and other leading regeneration scientists will attend a DARPA meeting in Virginia this week to further explore such a program.

DARPA, concerned that growing numbers in the military are surviving wounds that once killed and are reentering civilian life without arms and legs, isn’t the only outfit giving regeneration a boost. Eli Lilly and Company, the Indianapolis-based drug giant, has been funding one of the world’s leading regeneration labs at Indiana University-Purdue University Indianapolis. Lilly figures that even if regrowing a leg proves to be a fairy tale, the effort could lead to new wound-healing drugs for those, like many diabetics, who suffer chronic, non-healing sores.

This momentum and some scientific breakthroughs have led Gardiner to think that human beings may be able to grow new body parts soon. If Gardiner is right, growing a finger or an arm may turn out to be as simple as taking a pill, applying a drug-coated bandage or spraying a liquid onto a freshly damaged stump.

If Gardiner were alone in thinking so, he’d be easy to dismiss. But he’s not. Ellen Heber-Katz, a biologist at Philadelphia’s prestigious Wistar Institute, famed for its basic biology, vaccine and cancer research, has seen regeneration in her own lab, and, she insists, “This is doable. I believe it is inevitable that we will regenerate an entire human limb.”

Swiss scientist Abraham Trembley created the field of experimental biology in 1744 when he observed that dissected freshwater hydras could regenerate their missing parts. Then when Spallanzani discovered vertebrates could regenerate, science began speculating that people might regrow body parts, too.

In the late-19th century, the German biologist August Weismann thought he had it figured out. Weismann theorized that something he called “ids,” the rough equivalent of genes, contained information to direct the development of the body. Ids, Weismann thought, built animals by starting from the midline of the embryo and working out. By the time the fingertips were created, the cells had run out of information and could build no more. Cells at the elbow, for example, could construct a forearm, a hand and fingers but could not make an upper arm, shoulder or back. So, according to Weismann’s theory, cells at the site of a stump should contain enough information to build an arm or leg.

In the late 1890s, American biologist Thomas Hunt Morgan tried tackling regeneration. He spent hours slicing flatworms called planaria into smaller and smaller pieces and then watching as the slices grew into whole planaria. But he eventually quit in frustration, deciding that technology had not advanced far enough to distill whatever was stimulating the process. He turned his attention to fruit fly heredity instead, and went on to become the father of modern genetics.

Over the next decades, new theories arose. There was speculation about “morphogenic fields,” complex chemical factors dictated by genes that told cells to regenerate. Other experiments showed that nerve transplants could help animals grow new legs. But every time scientists thought the wall was about to crumble, they realized the barrier was actually a complex labyrinth.

Regeneration would also pale against the glamorous findings of James Watson and Francis Crick in deducing the structure of DNA in 1953. This new molecular biology allowed scientists to conduct “hard science” chemistry experiments that yielded unambiguous results. Experiments on salamanders and planaria were regarded as fuzzy, old-fashioned “naturalism.”

One breakthrough came in 1976, when biologists Vernon French and Peter and Sue Bryant revealed their “polar coordinate model” of development, in which cells know an anatomical “address.” That address then determines how the cells will perform. For example, when biologists amputated a salamander’s leg, a new bud, called a blastema, formed over the site. If scientists removed that blastema and transplanted it to, say, an incision on the salamander’s side, a new limb would grow there. University of Michigan scientist Bruce Carlson, the dean of American regeneration research, once created an animal with 17 toes.

Again, biologists thought they had the answer. But while the model provided a valuable road map, it still did not explain the cellular signals. During the 1980s, “regeneration fell off the radar screen,” says David Stocum, dean of the School of Science at Indiana-Purdue. By the early 1990s, only about 10 scientists in the world were mounting significant efforts to understand it.

David gardiner was a most unlikely apostle. Most of the scientists in his field are the products of a tweedy tradition of biology for the sake of science. But Gardiner, now 55, comes from a very different pedigree that’s all about practical application. He’s a blond-haired former surfer who got a degree in biology from Occidental College and went on to graduate school at UC San Diego’s Scripps Institution of Oceanography partly so he could live at the beach, scuba dive and play volleyball. Graduating, he says, was kind of a bummer because it meant he had to get a job.

He took one with his undergraduate alma mater, which had a contract from Southern California Edison to do field surveys of the environmental impact of its coastal generating plants. He studied fish eggs and larvae in Redondo Beach, work he found unsatisfying. Worse, his marriage was breaking down.

“So at age 30 I woke up one day and said, ‘This is not what I had in mind when I went into science.’ Things did not work out personally or professionally.” He answered a recruiting ad in the back of Science magazine and was hired by UC Davis to work in developmental biology--the study of how animals grow from cells into complete creatures. He specialized in frogs.

About six months later, in 1980, he attended a scientific conference and met Sue Bryant, who, as it happened, was divorced from Peter. When Sue Bryant was offered a temporary post with the National Science Foundation, she asked Gardiner’s advice about leaving UC Irvine and taking the job. “I said it sounded good, and she said she was going to drive her Fiat Spider [to Virginia]. I said, ‘So, anybody going with you?’ ” He smiles. “We got to know each other on the trip.”

Even after he married Bryant and she had returned to UC Irvine, where he joined her, Gardiner had no desire to work on regeneration. Sue Bryant was a star. He had taught her work to his Davis undergraduates. He had no intention of fading into her shadow. So he became an independent researcher pushing his frog studies. “It was a disaster,” he says, laughing. Being independent sounded good, but he had no idea how to strike out on his own, get research grants, handle himself in the academic rough-and-tumble.

Finally, a member of Bryant’s lab “took me aside one day and he said, ‘You are being stupid. You don’t need to be independent. You’re in a position to work with Sue. You could learn so much about regeneration and take it from there.’ I did not take that well.” But he joined the lab.

Maybe it was because he came from outside, maybe it was because he didn’t know much about regeneration, maybe it was just his go-for-it style. But Gardiner saw long-standing problems in new ways. Led by Bryant, he and the other members of her lab became iconoclasts, contradicting long-held theories.

One epiphany came at the Costa Mesa International House of Pancakes during breakfast. “I’ve been thinking . . . “ Bryant began, wondering about a certain chemical that plays a part in regeneration. Then Gardiner chimed in, and dozens of napkins scrawled with diagrams and arrows and symbols later, they had developed a new theory.

They formulated that the “addresses” predicted back in 1976 by the polar coordinate model could be altered to change a cell’s function. By giving the cell a new address and therefore changing the genes it expressed, you could alter the signals the genes sent to other cells. Those cells could then be made to perform new functions, like, say, build. But without the ability to screen for genes, they weren’t going to learn how to do that any time soon. Then everything changed in the 1990s.

Thanks to the human genome Project, by the end of the century computers had placed genetic databases in the hands of students. “In 1990 I cloned some 20-odd homeobox genes [which play a crucial role in development] and it took forever,” Gardiner recalls. “Now we do 10,000 sequences per month.”

Even more important, biology was rocked by two earthquakes. The first was Dolly, the cloning of an adult sheep, in 1997.

Dolly proved that August Weismann had made a good guess, but that cells did not run out of information after all. Almost every cell (red blood cells, platelets and sperm cells are exceptions) still contained the information it needed to make an entire body. “Dolly was the ‘Aha!’ moment,” says Gerald Schatten, a developmental biologist at the University of Pittsburgh. Until Dolly, Schatten says, “All of us had a sense that regeneration was a backwater field. Most medical research viewed newt and salamander regeneration as esoteric, but Dolly has changed the entire thinking about the plasticity of mammals, including humans.”

The second earthquake hit in 1998 with the isolation of the first human embryonic stem cell lines in a study led by James Thomson, a former student of David Stocum. These were the most plastic of all cells, the modeling clay of the human body, and they promised breakthroughs in healing damage from grievous injuries such as the one that paralyzed actor Christopher Reeve and in treating diseases such as Alzheimer’s and Parkinson’s. Biologists now foresaw that an adult mammal cell could go back in time and make stem cells that would then recapitulate development. Cells got a do-over.

This was old news to regeneration specialists such as Gardiner, who had watched salamander cells re-create development. Equipped with new lab technology, spurred by growing speculation that what worked for salamanders might work for people, biology began talking about “regenerative medicine.” Suddenly, Gardiner and the other regeneration experts were in vogue.

Bruce Carlson has seen all this before. Carlson, now semi-retired from the University of Michigan, was once where Gardiner is now, so he understands the younger man’s optimism. But despite the current buzz, he’s not buying it.

“The late 1950s was a period of great excitement,” Carlson recalls of his graduate school days. “When I was doing my PhD, I was afraid the problem of human limb regeneration would be solved by the time I got out of school.” It wasn’t, of course, and Carlson doesn’t think the labyrinth is about to be solved now.

He and Schatten say there may be good reasons why humans make scars instead of regrowing limbs. Evolution may have placed a higher value on our not bleeding to death in the aftermath of an amputating injury. So our bodies make clots to stop bleeding, then scars to seal the wound, and they form an Iron Curtain that prohibits regeneration. Nobody knows what will happen if this mechanism is bypassed. (Wistar’s Heber-Katz, for one, thinks we could stop bleeding, then quickly repopulate the wound with growth cells so a scar need never form.)

Regeneration also means that cells would have to proliferate, divide and grow, and recruit blood vessels to nourish them. This sounds suspiciously like cancer. It may be that on the long road of evolution we traded much longer lives for the regeneration powers of our slimy and short-lived forebears. Because our bodies tend to recognize proliferating cells as an enemy, we often mount an immune response to them. Maybe our immune systems simply prohibit limb regeneration.

“We have to separate the extremely exciting science from the unrealistic, impossible direct applications,” Schatten says. Biologists ought to be wary about promising anything, he argues, in light of the embryonic stem cell political battles in which some scientists think the practical benefits of research were oversold. “We should not promise things. With cancer, with wound repair and regeneration, we are still in the early discovery stages.”

Jeremy Brockes, a London-based scientist who has spent 20 years studying regeneration in newts, is only slightly more optimistic. “The idea that you could make a complex structure like a limb seems at present quite fanciful,” he says. On the other hand, “I genuinely believe it is too difficult to say if it is impossible. There is no clear reason why, in principle, it should not happen.”

Gardiner understands the caution but can’t help thinking that 100 years after Thomas Hunt Morgan gave up, science may triumph. Evolution? Schmevolution. “There’s no doubt in my mind there will be a day when we can regenerate anything.”

A sign on the wall near the entrance to Gardiner’s lab declares, “It’s All Growth, Stupid!” Gardiner’s view is that regeneration happens in three stages. Because of the limitations of past technology, scientists spent most of their time looking at stage 3, after the process is well along. To Gardiner, this is like trying to jump the Mississippi at New Orleans. He’d prefer to jump the small stream way up in Minnesota, at the very beginning of stage 1.

Gardiner ushers a visitor into a closet-sized room that smells a little like the muddy Mississippi and explains how he plans to make the leap. His lab, in cooperation with others, is building an “infrastructure of gene sequences,” a library of salamander genes. With these references in hand, they should be able to tease out the signals that begin the earliest steps in regeneration.

He asks Mathieu Rondet, a French graduate student, to locate a newly de-limbed salamander from among stacks of Tupperware containers on tall metal shelves. Inside is an unhappy-looking 6-inch pink salamander who is missing a hind leg. There’s a tiny white bud, the blastema, in its place. Rondet is trying to unravel the events of the first six hours following an amputation that led to the presence of that blastema.

Even without the infrastructure in place, Gardiner already has a strong theory. He is increasingly convinced that “there is a signal and response to wounding.” When tissues are damaged in an amputation, cells at the site release signals to call in reinforcements. He thinks that fibroblast cells (a kind of catch-all term for a class of cells found in many tissues, including lower layers of skin) provide raw materials. These cells migrate into the wound and switch jobs. “That transition is the critical thing that lets everything proceed,” he says. The cells make a blastema, which in turn makes cartilage, connective tissue and bone.

He’s certain that science can make this happen for people because we already do it as embryos, when cells receive signals on their receptors and begin to specialize, some becoming tiny buds that later grow into arms, legs, fingers, toes. It stands to reason, Gardiner says, that cells are still equipped with the genes that would make those receptors.

The mammary cell that made Dolly, after all, became every kind of cell in a sheep’s body. And after Dolly, cloning scientists experimented with switching the fate of cells. A few, like Tanja Dominko, have turned fibroblasts into other types of body cells, such as nerves. Dominko has applied for patents on the process and started her own company, CellThera, to pursue it. She also has been part of DARPA’s seedling program and has been cooperating with Gardiner.

So for Gardiner, regeneration is a matter of sending the right signal to the right address. “The signals are information passed between cells at their surface, a binding of a ligand [a cell signal] to a receptor.” If scientists can decide on the signals to use, understanding subsequent events might not be important. “Maybe there’s no need to explain it,” Gardiner says. “Maybe you can just stimulate the fibroblasts to make a blastema.” Then it’s all growth. The blastema knows what to do.

Several years ago, Ellen Heber-Katz was working with mice bred to display symptoms of autoimmune disease. She wanted to test a possible new drug, so her lab crew injected the drug into four mice and punched holes in their ears to tell them apart from four control mice who did not receive the drug. All eight mice were placed in the same cage. Three weeks later Heber-Katz was mystified to find eight mice with no ear holes. She repeated the experiment, and, she says, “we watched the mice and their ear holes closed. It was amazing. No scarring. You could not see anything. The hole was just gone.”

The mice with autoimmune disease had powers of regeneration. But since a crippling disease was a high price to pay for new fingers, Heber-Katz used her DARPA seedling money to apply what she learned to non-regenerating mice. “We got regeneration,” she says. “It’s not extreme. We did not get a whole tail, but we did get regrowth.”

It was simple, too. No genetic engineering, no complex manipulations. “We just painted stuff on the ear.” She refuses to say exactly what the “stuff” was--there’s intellectual property at stake--only that it was “a small molecule chemical compound,” science lingo for a drug you don’t have to inject. This lends support to Gardiner’s theory that a drug administered as a pill, ointment or spray could target the cellular receptors at the site of a wound to form a blastema instead of making a scar. That’s exactly what DARPA wants--to deploy medication in a soldier’s pack that can be slapped onto a wound to prevent scarring and help initiate the blastema process.

With the ability to screen for “regeneration genes” in salamanders, they argue, they should be able to find similar genes in people. Finally, after all this time, the elusive signals those genes make ought to bubble up from the data. Turn that signal into a drug, administer the drug to make a blastema and then stand back and let nature take its course. Sounds simple, so simple that some scientists say they expect it within 10 years. Certainly in David Gardiner’s lifetime. And if he were the scientist to author it, then maybe fairy tales do come true.