Limb Regeneration in Humans: New Research Challenges Long-Held Beliefs

The outcomes point to a shift in thinking about how regeneration might operate in human medicine.

In 2019, Ken Muneoka, a professor at Texas A&M University's College of Veterinary Medicine & Biomedical Sciences (CVMBS), published a ground-breaking article in Nature that demonstrated for the first time the feasibility of joint regeneration in mammals. Muneoka has a history of revolutionizing the field of regeneration.

His team has already begun to challenge further ingrained beliefs about the fundamental science of the matter, this time in relation to how mammals might repair bodily injury.

Only a few tissues, such the epidermis, the top layer of skin, and a few organs, like the liver, can normally regenerate in humans.

The ability to regenerate complex elements like bones, joints, and even entire limbs is possessed by other species, salamanders being the most notable. In order to comprehend the mechanisms underlying limb regeneration, researchers have researched these animals for more than 200 years with the goal of one day using these concepts to promote more thorough regeneration in people.

It is now commonly acknowledged as a result of this research that the nerves' presence is the single most crucial element in limb regeneration.

It might be true for salamanders and other animals, but not for mammals, according to two recent investigations by Muneoka. The first study, published in the Journal of Bone and Mineral Research in 2021, established that animals require mechanical loading, or the ability to apply force to or with an affected region. The second study, which was just released in Developmental Biology, demonstrated that nerve deficiency does not prevent regeneration.

These discoveries represent a significant shift in how regeneration might function in human medicine.

The two-century-old notion that you require nerves to regenerate is refuted by what these two investigations demonstrate, according to Muneoka. "Mechanical loading, not nerves, is what replaces it in mammals."

In order to stimulate regeneration in animals, two conditions need to be present in the damaged area, according to scientists. The first are molecules called growth factors, which can encourage cells to regenerate and rebuild damaged body parts.

These growth factors, which differ between species and depending on the area that has to regenerate, are created by the body during natural regeneration. These growth factors must be supplied to the area in order for regeneration to be caused by humans.

Nerves were regarded as the second ingredient that was required. This assumption was supported by a number of earlier investigations on human-induced mammal regeneration that focused on nerveless regions, typically digit tips, in which the entire limbs also lost their use.

According to the results of those tests, regeneration did not occur when growth factors were added, which led researchers to the conclusion that nerves, as in other animals, were necessary for regeneration.

However, the mechanical load factor was disregarded.

Is it really the nerves, or is absence of mechanical load also a factor? is the issue Muneoka and colleagues decided to ask themselves in their investigations.

A method to evaluate the denervation requirement in mammals was developed by Connor Dolan, a former graduate student in Muneoka's lab and the initial author of both recent research (who currently works at the Walter Reed National Military Medical Center). Dolan was motivated by astronauts.

NASA and other researchers have been experimenting with the method, known as hindlimb suspension, to see how mammals respond to zero gravity conditions for many years. When performing medical procedures on the legs of huge animals, a similar approach is utilized to stop the animals from putting weight on the injured limbs.

Despite having many nerves and being able to move, Dolan discovered that when the limbs were suspended, the animals couldn't truly apply pressure to their limbs, which prevented the digit tips from regenerating. It just prevented regeneration in any way.

But as soon as mechanical load comes back, regeneration is saved.

Nothing happens during the suspension, according to Muneoka. However, there will be a delay of a few weeks until the load is restored before they start to renew.

That initial step demonstrated that, despite the possibility that nerves were needed, mechanical stress was a crucial aspect of regeneration.

Dolan's second paper took the research a step further by proving that nerves are not necessary by demonstrating that a mouse can still regenerate a denervated digit even if it lacks nerves in one of its digits but has them in the others.

He discovered that they renew somewhat more slowly, but otherwise they did so normally, according to Muneoka.

Muneoka is careful to clarify that their studies are not refuting earlier research; rather, they are merely stating that it does not directly relate to humans.

"Numerous studies on salamanders have demonstrated that when the nerves are removed, they do not regrow," Muneoka added. Researchers have also been able to revive regeneration by introducing growth factors into the cells that they are aware are produced by nerves.

So, he concluded, "Salamanders presumably do need nerves to regenerate. But if we want to regenerate limbs in people, it will work much more similarly to how it does in mice.

Numerous of Muneoka's thoughts have challenged the accepted regeneration theories since he started studying the topic more than 20 years ago. He claimed that because they initially attempted to submit the two papers together, it took approximately three years to get them published.

Many scientists disagree with this concept, he claimed. "Studies of nerves and how they influence regeneration are incredibly important to a lot of people's professions. The whole biomedical usefulness of what researchers are doing in salamanders and fish kind of goes out the window when a study says that it's unlikely that humans would require the nerves.

It may seem like an intellectual point to say that mammals can regenerate without the need for nerves. After all, if a limb could not be felt or controlled because it lacked nerves, what good would it serve to regenerate it? In that regard, nerves will continue to be a crucial piece of the jigsaw.

According to Muneoka, the change is that nerves are now seen as a component of what has to be renewed rather than as a prerequisite for regeneration.

The problem, according to Larry Suva, chairman of the CVMBS Department of Veterinary Physiology & Pharmacology (VTPP), is that no one has previously considered the load factor.

Suva stated, "Imagine an explosion injury where a soldier is left with a stump. "Nobody was even considering a demand from mechanical factors before this study was published. Nobody was looking into the mechanical load component, but you had folks notice that a denervated animal doesn't regenerate and they think it's because the nerve was cut.                                                                    

Since I work with bones, he explained, "when I perceive a problem, I look at the bone problem." "People who work with nerves just focus on nerves. Therefore, it's extremely uncommon for someone to stand back and adopt a more comprehensive perspective like Dr. Muneoka.

That is what he brought to this concept, to this information that is 200 years old, Suva stated. Now that we are aware of how crucial the mechanical influences are, we must see regeneration from a different perspective.

One benefit of studying nerves is that researchers have been able to mimic the growth hormones that nerves secrete, enabling them to initiate regeneration in salamanders even in the absence of nerves. With these new discoveries, according to Suva, researchers will know they must take the mechanical stress factor into consideration if they wish to begin mammalian regeneration.

Scientists have already managed to deceive the body into believing that nerves are still present, he added. However, they now understand that they'll also need to mislead it into believing there is a mechanical load, which has never been done before.

Because cells respond differentially to mechanical stress, that stress is somehow translated into the cell's biochemistry.

Few laboratories are investigating the biochemical underpinnings of what mechanical load causes to a cell, according to Muneoka. If we could comprehend that biochemical signal, perhaps we might substitute the mechanical load's physical force with a molecular cocktail that would produce the identical signals in the cells.

Full human regeneration may still be a ways off, but according to Suva, this kind of fundamental shift in perspective is a significant milestone on the path.

It may still be science fiction, but we do know some facts about human limb regeneration. For example, we now know that mechanical load is necessary in addition to growth hormones. "That alters the approach that upcoming engineers and scientists will take to solving this issue.

Before it is possible to regenerate full human limbs, a number of challenging issues must be resolved, but Dr. Muneoka's results represent a crucial next step in making sure the proper issues are being addressed.

Connor P. Dolan, Felisha Imholt, Tae-Jung Yang, Rihana Bokhari, Joshua Gregory, Mingquan Yan, Osama Qureshi, Katherine Zimmel, Kirby M. Sherman, Alyssa Falck, Ling Yu, Eric Leininger, Regina Brunauer, Larry J. Suva, Dana Gaddy, Lindsay A. Dawson, and Ken Muneoka published "Mouse Digit Tip Regeneration Is Mechanical

The study, "Digit specific denervation does not inhibit mouse digit tip regeneration," was published in Developmental Biology on March 31, 2022 by Connor P. Dolan, Felisha Imholt, Mingquan Yan, Tae-Jung Yang, Joshua Gregory, Osama Qureshi, Katherine Zimmel, Kirby M. Sherman, Hannah M. Smith, Alyssa Falck, Eric Leininger, Ling Yu, Regina Brunauer, Larry J. Suva

By TEXAS A&M UNIVERSITY