Building a Regeneration Factory: New Discoveries in Limb Regrowth

Biotechnology

Building a Regeneration Factory: New Discoveries in Limb Regrowth

• April 11, 2025

The axolotl, a rare Mexican salamander, can regrow limbs, tail, heart, and even parts of its brain-abilities often described as "magical" or a biological "superpower."

At MDI Biological Laboratory, researchers see something less mythical: a marvel of nature and evolution. They are methodically uncovering the precise biological processes that drive this regenerative ability, down to the molecular level.

"Axolotl limb regeneration is one of the most fascinating phenomena in the animal kingdom," says Prayag Murawala, Ph.D. "The ultimate question is, 'How do they do this, and why can't we?'"

Murawala's group recently made a major advance: identifying specific cell populations and genetic activity that are essential to the salamander's ability to regrow a complete, functional limb.

Damián García-García, a Ph.D. candidate in Murawala's lab, led the benchwork over two and a half years, using MDI Bio Lab's powerful imaging and data analysis tools, while tending to a specialized population of transgenic axolotls.

"This is a huge step for us," García-García says.

(Re)building From the Ground Up

For humans, limb loss is usually permanent. Our immune systems prioritize sealing wounds over reconstruction, forming scars instead of new body parts. But axolotls take a different approach: their immune cells swiftly clean up the injury, and instead of scabbing over, a conical cluster of regenerative cells called a "blastema" forms at the wound site.

Blastemas are common in regenerative species like salamanders, zebrafish, but not humans. In axolotls, blastemas over time can transform into a new, fully functioning limb.

It's like reconstructing a high-tech factory that's been destroyed in an earthquake. First, debris is cleared (immune-system cleanup). Then specialized teams (regenerative cells) arrive: some will construct bone, others muscle, blood vessels, and nerves. Each step unfolds in order, until a fully functioning replica is complete.

As the blastema matures, immune cells give way to connective tissue (CT) cells, which typically function as structural "glue" among organs and tissues in many species. In the axolotl, adult CT cells do something extraordinary: they "dedifferentiate" and start acting like stem cells in a developing embryo.

As Murawala and colleagues demonstrated in Science (2018), during the blastema stage, as certain genes turn on or off, existing CT cells revert to this primitive, flexible state, then re-specialize into bone, tendon, and other cells that make up the body's scaffolding.

Getting Granular

In their newest work, Murawala's team used a next-gen tool called XENIUM to track more closely how varied cell types emerge in the blastema, and shift roles. The high-

resolution imaging allowed them to map individual cells, their positions within the blastema, and to observe the activity of certain involved genes.

"And what we found was that connective tissue cells play a far greater role in this process than was thought," Murawala says.

For decades, CT cells were thought to make up less than half the blastema. The new work shows they rise to as high as 75 percent of the blastema's makeup.

"So every three out of four cells are connective tissue cells," he explains. "This reveals that these cells are not just secondary, passive players. They are primary architects of limb reconstruction."

To prove it definitively, the team devised a novel method to selectively remove CT cells without harming the animal's overall health. The result: regeneration either failed or was severely impaired.

"Often in developmental biology or regeneration biology, what happens is, when one cell type is not present, another cell type can assume its role," Murawala says. "It was important for us to prove that if you don't have connective tissue cells, then you cannot build the limb. These are essential populations. It's not that somebody else is going to come and do the job for you."

From Salamanders to Humans

The ultimate challenge remains: How can we harness this biology for human healing?

Murawala says the new findings will guide future research in his lab:

• Immune System Differences: How do axolotl immune cells support regeneration, while human immune responses lead to scarring?
• Cellular Behavior: Why can axolotl connective tissue cells dedifferentiate while human cells cannot?
• Communication Mechanisms: How do these cell types in the blastema communicate with each other and what molecular signals orchestrate regeneration in axolotls, and could these pathways be unlocked in human limb repair?

"This research doesn't provide an overnight solution," Murawala emphasizes. "But it lays the foundation for tomorrow's therapies. By identifying where the roadblocks are in mammals, we can begin to chart a path toward overcoming them."