How to grow a new leg? Ask the axolotl!

Green squirrels have revealed the secret of the salamander's rebirthMembrane based on the materials of Wired Sciences and Technology Review

What a twist. Everything turned out to be completely different from what scientists thought before. Long–term studies of salamanders – champions of tissue regeneration – passed by an important feature of the recovery process and, moreover, they were looking for keys to it in the wrong place. And the main conclusion from the new work is optimistic: people have every chance to learn the amphibian trick.

The Mexican axolotl (Ambystoma mexicanum) taught scientists new lessons in growing limbs. This creature is one of the most outstanding examples of self–healing in the animal world.

That there is a tail, like some lizards: the axolotl successfully grows a new fully functional leg, instead of the severed one, scrupulously recreating all the bones, all the muscles, skin, vessels, nerve fibers... It takes only three weeks. Damaged lungs or spinal cord also in these salamanders (and in a number of others) are reconstructed remarkably. And there are no scars left. That would be so for us.

It has long been known that the restoration of an organ begins with the appearance of a blastema – a tumor-like cluster of cells that then multiply and turn into various tissues. These new cells are generated by ordinary cells at the site of injury, but the blastema cells are undifferentiated. However, they somehow find out what they will then turn into. So there is no way a tail can appear instead of a lost leg. This control system is a real mystery, the veil of mystery over which has been lifted by new experiments.

The general scheme of experiments: a transgenic animal synthesizes a fluorescent protein in all its tissues. The specific type of cells of interest to scientists from this specimen is transplanted to an ordinary animal, from which a limb is then cut off. After passing the blastema stage, the limb grows back, and fluorescent markers allow you to determine exactly what type of cells the transplanted "aliens" have turned into (photo by Martin Kragl et.al .).

Previously, biologists assumed that blastema cells are pluripotent, that is, each of them can become a cell of skin, muscle tissue, and so on, giving rise to the growth of a particular tissue. A group of researchers from Germany and the USA, who published an article in Nature, showed that this idea is wrong.

To find out the subtleties of regeneration, scientists have introduced a gene responsible for the synthesis of green fluorescent protein (the famous "Nobel GFP", so important for underwater organisms) into the axolotl.

Next, biologists alternately took certain cells from transgenic animals and introduced them to unmodified salamanders in the area of future trauma. And since the genetic information about GFP was preserved in all the cells – descendants of the borrowed cells, their position in the body, including in the newly grown organ, could be tracked with high accuracy.

So an amazing secret was revealed. Blastema cells are not identical, although they are not yet cells of certain tissues. But they remember from which cells they originated and thus the former muscle tissue cells produce only muscles, nerve fiber cells – new nerves, skin cells – skin and so on (with rare exceptions, about them – a little below).

"Many people had the impression that these very blastema cells were all the same," says one of the lead authors of the new work, Elly Tanaka, professor at the Center for Regenerative Therapy in Dresden. The answer surprised not only other researchers, but also her. It turned out that a blastema is not a homogeneous cell mass, but a cluster of progenitors generated by various tissues, which, in turn, generate each its own specific tissue.

That is, despite the blastema stage, these stem cells remain essentially different throughout the regeneration process.

In particular, experiments have shown that Schwann cells (which make up the protective shell of peripheral nerve fibers) obtained by an experimental amphibian from a transgenic specimen (and therefore glowing green), after the stage of supposedly pluripotent stem cells (as previously believed), generate only Schwann cells in the newly grown leg. After the limb has grown anew, the previously borrowed Schwann cells carrying the GFP gene spread and multiply exactly along the nerve fibers, creating their shells. And at the same time, no other cells of the original tissues turn into Schwann cells.

At the top of the figure: the spread of Schwann cells on the 3rd, 7th, 18th and 25th days after amputation. Below: various markers and imaging techniques have allowed scientists to clearly separate some cell types from others (photo by Martin Kragl et.al .).

The researchers also found that some cells remember not only their identity, but also their position in the body. Cartilage cells, for example, remember that they previously formed a shoulder, or the lower part of a limb, and migrate there in a new organ, while all Schwann cells indiscriminately simply move to any place where they are needed.

Tanaka says this work will trigger a major shift in thinking about regeneration requirements. Explaining why salamanders can repair limbs, but humans can't, she explains: "The hypothesis was that salamanders can greatly change the identity of cells."

But in fact, their cells never lose their "identity", – it follows from the new experiments, – on the contrary, salamanders seem to use "limited" in their capabilities stem cells from specific tissues, capable of generating only a certain part of a new limb.

Tanaka points out that humans also possess stem cells of specific tissues that replace the corresponding types of tissues. Only with us such healing is slow. "Salamanders don't do something much more complex than human stem cells can do," the researcher rejoices.

This means that pushing human cells to regenerate may not require such a radical step as turning cells into pluripotent cells, which many universities and institutes are vying with each other, sometimes demonstrating amazing results.

But now it turns out that nature's patent does not provide for the return of the "cell clock" for so many steps back, but only a small "rollback" occurs. And this process is more like experiments with direct reprogramming of some cell types into others, bypassing the pluripotent stage.

Alejandro Sanchez Alvarado, a researcher at the University of Utah and the Howard Hughes Medical Institute, who also studies the phenomenon of limb regeneration in amphibians, recognizes that the method of genetic "tattooing" of transplanted cells is a wonderful new way to study tissue regeneration. And he, they say, shows that the hypothesis about the heterogeneity of blastema cells is correct. (By the way, Alvarado published his own work on this topic in the same journal Nature).

But he warns that scientists still need to determine whether this principle is true for both adult axolotls and newts? If the same mechanism underlies other examples of regeneration, it will radically change scientists' ideas about the conditions of this remarkable process," says Alejandro.

But the main question remains unanswered: if people already have stem cells from different differentiated tissues, what is the difference between our cells and salamander cells? And wider – other animals?

Tanaka and her colleagues intend to deal with this issue: they will still conduct experiments in the hope of finding out exactly which genes are turned on at certain stages of the process and which molecular signals prompt cells at the wound site to form a blastema and follow the entire further path to limb reconstruction.

Portal "Eternal youth" http://vechnayamolodost.ru03.07.2009