Every year, African lungfish survive to the dry seasons in Africa by creating a cocoon that allows them to live on land for months or even years, until water returns following long periods of heat and drought. This cocoon, in which the lungfish is suspended in a prolonged state of aestivation, or torpor or dormancy, is a great way to prevent water loss, as many others have reported while studying aestivating frogs and other amphibians.
But researchers at The University of New Mexico are trying to find out how these animals survive pathogen and predator attacks when they are in this dormant, vulnerable state. The research could shed light on treating inflammatory diseases and investigating immune systems.
A study titled The lungfish cocoon is a living tissue with antimicrobial functions, recently published in Science Advances, was led by Associate Professor of Biology Irene Salinas and her team. The study was led by Ph.D. candidate at the Salinas lab Ryan Heimroth, who graduated in late 2020 and is now a postdoctoral researcher at Emory University. Key contributions were made by postdoctoral researchers in the Salinas lab, Elisa Casadei and Ottavia Benedicenti. Collaborators outside UNM include Chris Amemiya at University of California, Merced, and Pilar Muñoz at Universidad de Murcia, Spain.
This study reveals the extraordinary adaptations of the immune system of African lungfish which allow this species to survive the harsh aestivation periods every year. Salinas said this is the most fascinating and fun project she has ever worked on in her entire career as an evolutionary immunologist.
"We started this project in 2017 and we knew it was going to be exciting but we never thought that the results were going to be so astounding," she said. "When we started this project, we thought the cocoon is formed by mucosal secretions that dry up around the lungfish body, but when we looked closer we realized the cocoon was actually full of cells and the cells were alive. Further experiments revealed that the cocoon is formed and shedding of layer after layer of skin epidermis, thanks to the large numbers of dermal stem cells that lungfish."
We know very little about the immune system of lungfish, Salinas continued, but very old studies from the 1930s gave the researchers some hints. These pioneer studies told them that the lungfish produces unusually large numbers of immune cells called granulocytes. Granulocytes are very important first lines of defense against pathogens and the first cells to migrate to sites of inflammation.
But why do lungfish have so many? Why is that so important? The team looked carefully at how these granulocytes changed when they aestivated lungfish in the laboratory. Granulocytes left their tissue reservoirs, traveled in the blood, and flooded the skin of aestivated animals. This is a hallmark of inflammation, very similar to what happens in the human gut and skin when it is inflamed. Lungfish do it to themselves as soon as they sense that the environment is unfavorable.
"So what do these cells do when they get to the skin? Well, they leave the skin and become part of the cocoon," Salinas said. "Yes, the cocoon is now not only a layer of mucus that prevents water loss but an immunological shield, full of granulocytes, potent antimicrobial soldiers that can trap and kill pathogens."
Once the researchers saw the granulocytes in the cocoon they were sure the cocoon had immunological functions, Salinas said. Following a series of investigative procedures, they concluded that the cocoon acts as an extracorporeal bacterial trapping device, the lungfish body staying healthy during aestivation, the cocoon fighting bacteria outside the body.
"The key to the cocoon antimicrobial function is that granulocytes have fancy magic tricks under their hats," Salinas said. In a process known as extracellular trap formation, granulocyte extrude their DNA along with many antimicrobial compounds that decorate the DNA forming these traps. The lungfish cocoon was full of granulocytes that were caught in the process of making extracellular traps, explaining why bacteria did not penetrate into the aestivating lungfish body.
The team set experiments to answer what happens if a lungfish cocoon cannot make extracellular traps and found that extracellular DNA was essential for lungfish to stay healthy during the aestivation process.
While these findings reveal very fundamental immunological adaptation of a vertebrate animal with extreme biology, they may also illuminate key aspects of maladaptive immune responses that occur during inflammatory diseases at mucosal barriers.
Clearly, lungfish are very good at injuring their skin and sef-inflicting inflammation, Salinas said, yet when water returns, they are able to regenerate their tissues and swim back in the water as if nothing ever happened.
"These animals, therefore, may keep many secrets that we could use in the future to treat inflammatory diseases, Salinas said, adding that she also wants to advocate for the investigation of immune systems in non-traditional models, such as the lungfish.
This work was generously supported by the National Science Foundation award #1938816. The team hopes to continue to investigate the immune system of this fascinating animal in the next few years, Salinas noted.
