Though we’ve now been living with SARS-CoV-2, the virus that causes COVID-19, for close to five years, many aspects of its biology and pathology remain mysterious. This is not helped by the fact that new variants of the virus are constantly evolving, tweaking their features to wreak havoc on the human body.
Primarily a respiratory virus, our understanding of SARS-CoV-2 has been hampered by an inability to study the pathogen in its natural habitat—a living, breathing human lung. Thankfully, lung biologist Sandra Leibel, M.D., has been working with “mini-lungs” derived from human stem cells since 2012.
Once COVID-19 emerged in early 2020, Leibel teamed up with immunologist Ben Croker, M.D., to infect her balloon-like organoids with variants of the virus to understand its behavior and how our lungs respond to it. Doing so revealed the virus’s hidden infectious capability, a lung-specific immune response, and potential avenues for preventing and maybe even treating COVID-19. The duo, both from the University of California, San Diego, and colleagues reported the findings on July 19 in Proceedings of the National Academy of Sciences.
“This paper highlights the exciting time we live in, in which patient-derived induced pluripotent stem cells can be utilized to build mini lungs in the lab,” virologist Mart Lamers, of Duke-NUS Medical School in Singapore, told Fierce Biotech in an email. Lamers was not involved with the study.
Leibel originally started nurturing miniature lungs while working in the pediatric lung transplant center at Washington University in St. Louis.
“There are some babies that are born with a genetic deficiency in the surfactant protein A gene, and those babies will not survive unless they get a lung transplant,” Leibel told Fierce Biotech in an interview. Her work at the time was to take stem cells from her young patients, turn them into lung cells, fix the genetic glitch causing them not to make surfactant, and then put those cells back into the babies.
Over time, the clusters of lung cells Leibel was making in the lab became more complex and three-dimensional. Once Croker joined UCSD, the two got to chatting and saw the potential for using Leibel’s mini-lungs to understand how certain proteins drive inflammation in the lung during a viral infection. And then, COVID arrived.
“We decided we needed to put that model to work, to study the response to SARS-CoV-2,” Croker said in an interview.
Leibel has amassed stem cell lines from diverse patients, including men and women and people of different racial backgrounds. She and her colleagues grew these stem cells into mini-lungs consisting of multiple different types of lung cells. They then infected the lab lungs with COVID variants and tracked how the virus spread and how the cells responded.
“We pretty quickly realized how efficiently the virus could infect different types of lung cells,” Croker said. “Which didn't really sit exactly with what the literature was telling us at the time.” SARS-CoV-2 was known to infect cells by binding to receptors, like ACE2, on the cell’s surface, but Leibel and Croker saw that the virus could also get into cells lacking those receptors through a “back door.” When cells take in large amounts of fluid, a process called macropinocytosis, the virus can be gulped up too.
“The authors suggest that macropinocytosis might lead to ACE2-independent entry, but more work is needed to prove this,” Lamers said. “Cells may express ACE2 at levels below detection limits, but low amounts of ACE2 may still allow virus entry as was observed before in intestinal organoids.”
Normally, contracting a viral infection would lead to an immune response led by specialized immune cells. Even though the mini-lungs lacked any immune cells, the lung cells themselves still showed an immune response of their own. Analyzing gene activity revealed that after being infected, lung epithelial cells responded by producing pro-inflammatory proteins like chemokines and interferons. This response was controlled by a protein called surfactant B, which is related to other surfactants but had no previously recognized role in immunity.
“Surfactant protein A and D, those are the immune surfactants,” Leibel said. “B and C are usually the surfactants that help so our lungs don't collapse every time we exhale.” There had been some hints that surfactant B might help fight infections by the bacteria Pseudomonas aeruginosa, Leibel said, but this work is the first evidence that it defends against SARS-CoV-2 and contributes to a stand-alone lung immune response.
When Leibel and Croker added surfactant B to organoids before infecting them, they found that it protected the mini-lungs from the virus. “There was less viral infectivity with that prophylactic addition,” Leibel said. She and Croker next want to study the mechanism underlying this protective effect and see if surfactant B can be turned into a prophylactic for high-risk individuals, or potentially even a COVID treatment.
They also want to ratchet up the complexity of their mini-lungs by adding some immune cells, to see how lung cells infected by SARS-CoV-2 signal to the immune system that they’re in trouble.
“We're just showing that first response of the cells to the virus,” Leibel said. “But what about that second response when they're actually calling for help and they need those immune cells to come?”