Low-oxygen environments can activate limb regeneration programs in mammals.

Biologists have long been puzzled by the fact that salamanders and frog tadpoles can regrow complete limbs after amputation, while mammals, including humans, cannot. A recent study from the Swiss Federal Institute of Technology in Lausanne (EPFL) reveals that the key to this difference may lie in oxygen. 

The study has been published in the top international journal Science.

Regeneration and Scarring: Two Forked Paths to Wound Healing

The first step in limb regeneration is wound healing. When a limb is amputated, the cells at the wound site must quickly close the wound and transform into cell types capable of regeneration. In amphibians, this process proceeds smoothly; however, in mammals, wound closure is slow, and scar tissue forms rapidly, thus blocking the possibility of regeneration. 

The research team noticed a long-overlooked environmental difference: amphibian larvae typically develop in aquatic environments, where oxygen levels are much lower than in air. In contrast, mammalian tissues are often exposed to higher concentrations of oxygen after injury. Whether this difference is a direct cause of regeneration or merely a side effect of different lifestyles has remained inconclusive. 

The oxygen sensing pathway determines whether regeneration can be initiated.

A research team led by Kan Aztekin, a scientist at the Swiss Federal Institute of Technology in Lausanne (EPFL) and now working in the Friedrich Michel laboratory at the Max Planck Society, discovered, by comparing amputated limbs of frog tadpoles and mouse embryos, that the way cells sense oxygen directly determines whether the regeneration process can be initiated.

Researchers removed developing limbs from frog tadpoles and mouse embryos and cultured them in vitro under controlled oxygen conditions. They lowered the oxygen level to a low concentration comparable to that in aquatic environments or raised it to a high concentration close to that in air, and systematically tracked wound healing rate, cell motility, gene activity, metabolic state, and epigenetic changes.

The research focuses on a protein called HIF1A, a key oxygen sensor in cells. When oxygen levels are low, HIF1A tends to stabilize, thereby activating a series of genetic programs that prepare for wound healing and regeneration. 

Hypoxia conditions awaken the potential regenerative capacity of mammals

Lowering oxygen levels had a significant impact on the limbs of mouse embryos. Under hypoxic conditions, wound healing in mouse cells was significantly accelerated, and early signs of entering the regeneration process began to emerge. Similar effects could be achieved even when oxygen levels were maintained at higher levels, provided that the HIF1A protein was stabilized through other means. 

Further analysis revealed that the hypoxic environment also altered several cellular behavioral characteristics: skin cells became more active, and their mechanical properties changed; cellular metabolism shifted to glycolysis, an energy metabolism mode unique to hypoxia; and chemical markers on DNA-related proteins also showed changes that were conducive to the activation of regeneration genes.

Amphibians are naturally endowed with the advantage of regeneration under low oxygen conditions.

In stark contrast, frog tadpoles exhibit highly efficient limb regeneration under various oxygen concentrations, even those far exceeding normal atmospheric oxygen levels. Molecular analysis reveals that frog tadpole cells have low gene expression levels responsible for shutting down the HIF1A pathway, thus maintaining stable HIF1A activity even at elevated oxygen concentrations.

By further comparing datasets from frogs, axolotls, mice, and humans, the research team discovered a consistent pattern: amphibians with regenerative abilities exhibit a lower level of oxygen sensing, which allows the regeneration process to be initiated and continue running smoothly; while mammals, on the contrary, have cells that respond strongly to oxygen and actively shut down the regeneration process shortly after injury.

Providing a fresh perspective on a century-old problem

These results indicate that mammalian limbs retain potential regenerative capabilities during early development, and whether this capability is activated depends on how cells respond to environmental signals such as oxygen. This suggests that by regulating oxygen sensing pathways, it may be possible to improve the quality of wound healing in humans and even induce a certain degree of regenerative response in the future.

To be clear, this study does not claim that human limb regeneration is about to become a reality. However, it does demonstrate that the differences in regenerative capacity previously thought to be inherent between different species depend to a considerable extent on how cells respond to microenvironmental conditions.

Aztkin stated that the research team was very excited about the results. He pointed out that by directly comparing species capable of regeneration with those that cannot, this study provides a completely new perspective on a problem that has plagued the scientific community for centuries. The research confirms that regenerative processes in mammalian tissues can be activated and has begun to outline a clear, verifiable pathway to promote limb regeneration in adult mammals.

The experiment was conducted at the Swiss Federal Institute of Technology in Lausanne (EPFL), in accordance with Swiss animal welfare regulations, and had been approved by the relevant veterinary authorities. The animal experiments were based on the judgment that the potential scientific benefits outweighed the possible suffering inflicted on the animals.

Original source: EPFL (École Polytechnique Fédérale de Lausanne)

Published in: Science magazine