In a bamboo forest, a giant panda chomps on some bamboo leaves—but for how long? While these cuddly looking bears have few natural predators, they remain vulnerable.1 Today, less than 2,000 giant pandas live in the wild, and another 600 survive in zoos and protected habitats around the world. As loss of this umbrella species will have a ripple effect on the entire ecosystem, scientists are working to conserve these gentle bears and increase their numbers. Inbreeding within a small population can lead to loss of genetic diversity, potentially decreasing resistance to diseases and parasites, so researchers are on the lookout for novel conservation approaches.
In 2011, scientists speculated that a revolutionary technology, induced pluripotent stem cells (iPSCs), could be the key to conserving endangered and vulnerable species and have since generated iPSCs from the northern white rhinoceros, the Tasmanian devil, and Grevy’s zebra.2–4 But although scientists had some success generating stem cells from the cheek mucosal cells of giant pandas, truly pluripotent cells eluded them.5
So when Jing Liu, a stem cell biologist at the Chinese Academy of Sciences, got a request in 2019 from the Chengdu Research Base of Giant Panda Breeding to create giant panda iPSCs from fibroblasts, he was up for the challenge. Now, in a study published in Science Advances, Liu and his team reported that iPSCs can be generated from the skin cells of giant pandas.6 Their findings open new avenues for researchers to study panda biology in greater detail and devise new measures to protect them.
The journey from panda fibroblasts to iPSCs was not easy. When Liu and his team first tried reprogramming conditions that worked for other species, they met their first hurdle. “Surprisingly, when we used the mouse and human conditions, they didn’t work for the panda [cells],” Liu said.
“The recipe from the mouse is not necessarily directly applicable to other species, even within mammalian species,” said Pierre Comizzoli, a gamete biologist at the Smithsonian’s National Zoo and Conservation Biology Institute, who was not involved in the study. “So, every time you have to go back to the basics to really understand what factors can influence the reprogramming of the cells.”
After some trial and error, Liu and his team finally found out that introducing a specific microRNA cluster was key in transforming the fibroblasts to iPSCs. Once they modified the growing conditions to include additional molecules, such as transcription factors, that are specific to pandas, the team successfully obtained iPSC clones. “The clones were very beautiful. We were so excited,” Liu reminisced.
The team then confirmed that the iPSCs that they had identified based on physical characteristics exhibited the associated genetic traits of the stem cells they were after, such as a decrease in the expression of genes associated with somatic cells and an abundance of pluripotency-related genes.
Once Liu and his team fine-tuned the reprogramming steps, they looked to shorten the process and make it more efficient. With a few tweaks to the cell culture medium to include certain signaling pathway modulators, epigenetic inhibitors, and kinase blockers, they reduced the total experimental duration from over three months to less than a month and increased efficiency by five-fold as compared to an unaltered medium.
The true mark of a pluripotent stem cell is its ability to divide and form the three germ layers— endoderm, mesoderm, and ectoderm—which are crucial for the development of the body’s tissue and organs. To put the giant panda iPSCs to the test, the team observed the formation of embryoid bodies, a collection of pluripotent stem cells that recapitulate some aspects of early embryogenesis. During this developmental stage, they observed an increase in ectodermal markers while later stages showed an increase in mesodermal and endodermal markers. When they injected the giant panda iPSCs into mice, the cells formed a mass that exhibited the three germ layers with elements of neural, muscular, and epithelial tissues.
“You can generate the iPSCs, but then after that when you culture them, they try to go back to something much more specialized and that’s really the challenge,” Comizzoli said. “But the paper is describing very interesting culture conditions to maintain the iPSCs in the same state for longer period of time, which is very inspiring for the field.” However, Comizzoli emphasizes that just because these conditions worked for the giant panda, it doesn’t mean that they’ll automatically work for other species.
Liu hopes to someday use the giant panda stem cells to create sperm and oocytes, which can potentially be used to create giant panda embryos. “We want to use these stem cells to create an animal,” Liu said. “This is a challenging thing in the field.”
According to Comizzoli, there’s a long way to go before scientists can generate functional gametes from iPSCs. He said, “The most immediate applications are in regenerative medicine to treat sick pandas and to better understand the embryology or fetal development of these animals.”.
