Researchers in Japan have created the first line of coral cells that can live and proliferate long enough to be useful for lengthy research projects in the lab, solving a well-known problem in coral cell biology.

The method, documented in a study published April 26 in Marine Biotechnology, has the potential to massively simplify lab-based coral research, which is critical for conservation as the populations of wild corals continue to decline around the globe.

"Okinawa is surrounded by very beautiful seas, previously full of very beautiful coral reefs," said senior author Noriyuki Satoh, a marine genomics professor at Okinawa Institute of Science. "But since [the] 1998 El Nino, the worldwide coral reef almost disappeared." 

Satoh is referring to the particularly harsh El Nino event of 1998 that triggered the first global coral bleaching event. 

"So many kinds of people [are] very much interested in some kind of preservation or restoration of coral reefs in the future," he added.

Coral researchers are more frequently relying on laboratory methods to study coral as the reefs themselves are becoming scarcer worldwide. Nearly half of the Great Barrier Reef off the coast of Australia has died over the last quarter-century.

Farther north, almost three-quarters of the coral around Okinawa has died due to coral bleaching, which occurs when photosynthetic algae living within coral are expelled into the water. This causes the coral to turn white and eventually die from malnutrition.

Laboratory experiments on coral can provide insights into the natural processes that sustain these complex creatures, particularly the symbiosis between the coral and the algae living within them. And while there are many examples of successful cell lines for other animals, including humans, coral cells tend to die when cultured in seawater or conventional culture media.

According to Satoh, the initial stage of cell culture, when the cells are isolated from tissue, is "not so difficult" for coral. However, it has proven a major challenge for researchers to create stable lines of cells that can be used over and over, continuing to proliferate after the initial culture stage.

"This is the main contribution of our recent research," he said.

Satoh explained that this project built upon previous work with sea squirts undertaken by first author Kaz Kawamura, an "expert in establishing stable cell lines." The team attempted to adapt Kawamura's methods to work with cells from Acropora tenuis, one of the most abundant species of coral in Pacific reefs. 

The researchers used existing methods to successfully dissociate samples of coral larvae into three colors of cells: brown, translucent and pale blue. The group then tried many different combinations of culture media to encourage these cells to continue to grow. While blue and translucent cells died after just a week, the team discovered that the brown cells did quite well with the addition of plasmin, an enzyme that is found in blood and degrades certain proteins.

With the plasmin, the cells have proliferated in clumps for more than 10 months, continuing to double approximately every four or five days. The cells can even remain viable after being frozen in liquid nitrogen, which Satoh states is "crucial for being able to successfully supply the coral cell lines to research laboratories across the globe."

However, proliferation is not always a cause for celebration in cell line experiments, because it may also be a sign of contamination.

To verify that they had actually produced coral cells, the researchers used immunohistochemistry staining, a process in which cells are targeted with antibodies and then stained to detect the presence of antigens that bind to these antibodies, confirming the identity of the cells. Using antibodies specific to the coral cells, the researchers were able to detect coral-specific antigens, indicating the cells that proliferated must be coral cells. The team also looked to the cells' DNA for extra confirmation.

"We are very sure," Satoh said. "We did transcriptome analysis of these cultural cells, and the genes expressed there are nearly 100% A. tenuis."

The researchers hope that their method of producing coral cells will allow the team to study coral more closely than is possible when working in the field or on short-lived cell lines. The researchers hope they may even be able to observe the interaction between coral cells and the microscopic algae that live within them. This relationship has remained mysterious to researchers, specifically because it is so difficult to maintain coral cells in the lab.

"Nobody has observed under the microscope the symbiosis of coral cells and photosynthetic dinoflagellates," Satoh said. "It's kind of a dream."

In the meantime, the researchers hope that their methods will be applied to other coral species and be used by scientists across disciplines to study and conserve global coral reefs.

"Many kinds of coral researchers each have different interests," Satoh said. "If everything goes well, this kind of stable cell line [can] provide many kinds of opportunities to study the coral bio symbiosis with a different point of view."

The study, "Establishing Sustainable Cell Lines of a Coral, Acropora tenuis," published April 26 in Marine Biotechnology, was authored by Kaz Kawamura and Shigeki Fujiwara, Kochi University; and Koki Nishitsuji, Eiichi Shoguchi and Noriyuki Satoh, Okinawa Institute of Science and Technology.