Heal thyself

When Wistar Institute scientists announced last month they could endow mice with enhanced healing powers by altering just one gene, it raised a question: Why didn't evolution lead to mice - and people - with this power in the first place? Were we mammals cheated by evolution?

Scientists know that a few amphibians and fish can regrow lost limbs, fins, organs, and even the lenses of their eyes. Among vertebrates, the champion healers are salamanders, who can generate a whole new leg, if needed. For years, scientists have been looking into the salamander's cellular structure and genetic code, trying to divine its secret.

The recent discovery in mice is helping scientists finally start to put the pieces together, offering new clues to regeneration and even raising hopes that one day medicine may make up for what evolution denied us.

Wistar scientists in Philadelphia discovered super-healing mice by accident in 1996, but until now they didn't know exactly what separated them from their ordinary rodent counterparts. Now they've been able to show tissue regeneration in a strain of mice that lack a single gene, called p21. That gene was previously known for its role in preventing cancer.

So far, these mice don't seem unusually prone to cancer, said biologist Ellen Heber-Katz, the leader of the Wistar team behind the discovery. But they are prone to autoimmune diseases, in which antibodies attack normal cells and tissue. That gives them symptoms such as the skin irritations typical of lupus and other diseases.

So scientists may not want to start using genetic engineering to remove human p21 just yet.

That these mice get autoimmune diseases isn't a total surprise. The first super-healing mice, discovered 14 years ago, were bred to study such diseases. Called MRL mice - an abbreviation for Murphy Roths Large - they get a disease similar to lupus.

That's why Heber-Katz was working with them. An immunologist, she studies autoimmune diseases and was busy testing possible new drugs for treating multiple sclerosis. A colleague mentioned that these MRL mice might be useful because they get an autoimmune disease.

So she ordered some MRL mice. She asked a postdoctoral fellow to punch 2-millimeter holes in the ears of mice getting the treatment - a standard procedure that's often used to separate treated mice from untreated controls.

Then something weird happened.

Several weeks later, Heber-Katz discovered that none of the mice had ear punches. She thought the postdoc had forgotten and ruined the experiment. "I was really upset," she said. But the postdoc insisted she had indeed punched holes in the ears.

Heber-Katz realized that if the postdoc was correct, then an extraordinary thing happened to their mice.

"We went upstairs and re-punched them and watched them closely," she said. The holes healed within two weeks. "We didn't know if this had anything to do with immunology, but it certainly was an amazing form of wound healing," she said.

So she went to a meeting of specialists in wound healing. There she met an expert on salamanders - David Stocum from Indiana University. He thought there was a connection between the salamander and her mice.

Stocum says there's a fundamental difference between healing and regeneration. When we heal, we form scar tissue like a patch over a wound. Salamanders don't make scar tissue but instead reprogram groups of cells, turning back some molecular clock to make them act like embryonic stem cells that can generate all kinds of tissues.

He thought these mice were not healing but regenerating.

For the next 14 years, the Wistar team tried to figure out what gave the MRL mice this ability.

Eventually they found that cells near the sites of injuries were dividing in unusual ways - often developing a double nucleus, for example. A colleague of Heber-Katz noticed that a gene called p21 was inactive in cells at these healing sites.

Cancer researchers study p21 because it appears to act as part of a cancer-prevention mechanism. When cells divide, various "checkpoints" stop the process to identify and repair copying errors in the DNA. Left unchecked, such errors can render a cell malignant.

Other researchers had also identified p21 as important in distinguishing ordinary cells from so-called pluripotent cells - those that make up embryos and retain flexibility to become different parts of the body.

With so much interest in p21, other scientists had already made genetically altered mice without a p21 gene. Heber-Katz ordered some of these "knockout mice" and found that they possessed the same enhanced healing powers as the MRL mice. An ear punch closed up without a scar. These mice were also prone to autoimmune disease.

When regeneration expert Anthony Mescher heard about Heber-Katz and her healing mice, he started thinking about the immune system and its role in the power of the salamander.

Salamanders have what's called an innate immune system - a primitive one they share with mammals. But they lack the more complex type of immunity that produces antibodies to specific viruses and bacteria, said Mescher, who works at Indiana University.

"The development of a more sophisticated immune system in evolution seems to have brought about the loss of regeneration," he said.

Meanwhile, his Indiana colleague David Stocum is looking for the connection between salamanders and the super-healing mice. At the site of injury, he said, a salamander will make lots of a protein called evi 5. That appears to help switch the cells into a more embryo-like state.

He's planning to look at p21 in the salamander, and the Wistar group is now investigating evi 5 in their mice.

Stocum and Mescher also study frogs to understand why these amphibians can't regenerate limbs while salamanders can - except when the frogs are still tadpoles.

Perhaps, Mescher said, evolution didn't get rid of regeneration in frogs and mice and people. "Maybe it's still there - we just have to find out how to tap into it," he said.

As embryos, even mammals can regenerate organs and limbs much the way the salamander can. That suggests "that we have the genetic equipment to do it," he said, "But the decision has been made evolutionarily to use the scarring equipment instead."

Other experts in regeneration find the healing mice intriguing but aren't sure that it's true regeneration.

When a salamander loses a leg, it forms a nub of stem cells called a blastema, said Jeremy Brockes of the University College, London. He said it's not yet clear whether the MRL or the p21 mice are making blastemas as well. "It's not something where there's any agreed-about definition," he said.

One key property of a blastema is you can take one that's starting to form a new salamander leg, cut it off, transplant it to another part of the salamander's body, and it will continue to grow a leg.

Brockes also questions whether regeneration is the ancestral state of mammals, as has long been assumed because amphibians have been around much longer. "The challenge was to unlock the newt within," he said.

He started to doubt that conventional wisdom after discovering some regeneration-related genes that are unique to the salamander.

Brockes said these salamander genes may have been overlooked because scientists pay more attention to those genes that we and the salamander share.

His view now is that salamanders need a number of genes to regenerate their limbs, some that they share with us and some that they don't. "I don't think there's some ancestral option that's lost in animals that don't regenerate," he said. "I think in this case it's the salamander that solved the problem of getting a regenerative response to amputation."

In other words, there may be no newt within us.

But we do share p21 with the mouse and this gives Heber-Katz hope that something useful will come of it.

While it might not be wise to go destroy our own p21 genes in the quest for enhanced healing, she is investigating the chance that a drug could temporarily block the action of p21, transforming patients into super-healers for a month or so, just long enough to get over some severe injury without scarring.

She said they're starting to work with other scientists, experimenting with drugs that block p21 in mice. While tapping into regeneration might be a long-term project, she said, "there are a lot of potentially simple things to do that may help."