Successful rodent chimera experiments, such as this mouse embryo harboring rat heart cells (red), have been hard to re-create with human cells. 

BELMONTE LAB, SALK INSTITUTE FOR BIOLOGICAL STUDIES

Embryo experiments take ‘baby steps’ toward growing human organs in livestock

The perpetual shortage of human organs for transplant has researchers turning to farm animals. Several biotech companies are genetically engineering pigs to make their organs more compatible with the human body. But some scientists are pursuing a different solution: growing fully human organs in pigs, sheep, or other animals, which could then be harvested for transplants.

The idea is biologically daunting and ethically fraught. But a few teams are chipping away at a key roadblock: getting stem cells of one species to thrive in the embryo of another. Last month, a U.S. group reported in a preprint that it had grown chimpanzee stem cells in monkey embryos. And newly loosened regulations in Japan have encouraged researchers to seek approval for experiments to boost the survival of human cells in the developing embryos of rodents and pigs. Insoo Hyun, a bioethicist at Case Western Reserve University in Cleveland, Ohio, says the work is being done responsibly. Efforts such as the new chimp-monkey chimeras represent "baby steps forward, gathering data as you go," he says. "And I think that's a wise approach."

Ultimately, the researchers envision reprogramming a person's cells to a primitive developmental state that can form most any tissue and injecting these induced pluripotent stem (IPS) cells into another species's embryo. The embryo would be implanted in the uterus of a surrogate, and allowed to grow to full size to serve as an organ donor. The IPS cells could come from the person awaiting transplant or, in a potentially faster and less costly approach, human organs could be grown in advance from cells from other donors, matched for key immune signaling proteins to prevent rejection.

So far, the feat has been modeled only in rodents. In 2010, stem cell biologist Hiromitsu Nakauchi and his team at the University of Tokyo reported growing rat pancreases in mice that couldn't form pancreases of their own. In 2017, Nakauchi and colleagues treated diabetes in mice by giving them transplants of insulin-producing mouse pancreas tissue grown in a rat.

But the success in rodents hasn't held up between larger and more evolutionarily distant animals. In 2017, cell biologist Jun Wu and colleagues in Juan Carlos Izpisua Belmonte’s lab at the Salk Institute for Biological Studies in San Diego, California, reported that when they injected pig embryos with human IPS cells and implanted the embryos into sows, about half of the resulting fetuses were stunted and slow growing. Those that were normal size had very few human cells after a month of gestation.

Wu, who is now at the University of Texas Southwestern Medical Center in Dallas, has since explored how human stem cells interact in a lab dish with stem cells from nonhuman primates, rats, mice, sheep, and cows. He's found what he calls "a very exciting phenomenon: a competition between cells of different species." Pitted against cells of distantly related animals, human cells tend to die off, and the team is now trying to understand the mechanism. "I think we are almost there," Wu says.

But competition isn't the only problem. Primate IPS cells are also more developmentally advanced, or "primed," than the "naïve" rodent stem cells used in the earlier successful chimera experiments. They are therefore less likely to survive in a chimeric embryo, says Nakauchi, who also has a lab at Stanford University in Palo Alto, California. To help primate IPS cells thrive, his Stanford team and collaborators endowed them with a gene that prevents cell death. In the experiments reported last month, they tested how the modified cells would fare in the embryo of a closely related primate species.

To avoid raising ethical concerns, the team decided not to use human IPS cells. If a nonhuman primate embryo with added human cells were allowed to develop in a surrogate and many human cells survived and proliferated, the result would be an unprecedented primate chimera. "People are concerned that the boundary between humans and animals could become blurred," says Misao Fujita, a bioethicist at Kyoto University in Japan who recently conducted a survey of attitudes toward animal-human chimeras in the Japanese public. Respondents were particularly worried that such animals could have enhanced intelligence or carry human sperm and egg cells.

Nakauchi's team instead modified IPS cells from the closest human relative, the chimpanzee, and put them into rhesus macaque embryos. They found that, compared with unmodified chimpanzee IPS cells, the cells with the survival-promoting gene were more likely to persist in the 2 days after they were inserted into a 5-day-old monkey embryo. It's hard to keep a monkey embryo alive in a dish for much longer than a week, Nakauchi says, but his team plans to grow its chimeras further by implanting them into the uteruses of female macaques "in the near future."

Nakauchi also has submitted proposals to a government committee in Japan to put the survival-promoting gene into human stem cells and inject them into mouse, rat, and pig embryos—but not nonhuman primates—that lack a gene critical to pancreas development. The researchers hope that, as in the earlier rodent experiments, the human cells will begin to form the missing pancreas. His team would implant the embryos in surrogate animals but remove them for study before they reach full term. The proposals are an initial test for new legal guidelines in Japan, which in March lifted an outright ban on culturing human-animal chimeras past 14 days or putting them into a uterus.

Other groups are honing different recipes for chimera-friendly stem cells. In January, a team from Yale University and the Axion Research Foundation in Hamden, Connecticut, described culturing monkey IPS cells with chemicals that prompted gene expression patterns like those of mouse embryonic stem cells, which are more likely to survive in a chimera. In April, Yale University stem cell biologist Alejandro De Los Angeles reported that the technique prompted similar gene expression changes in human IPS cells. He's now considering testing how these cells hold up in a mouse or other nonhuman embryo.

Such work faces hurdles in the United States. There is no outright ban, but in 2015 the U.S. National Institutes of Health (NIH) in Bethesda, Maryland, froze its review of grant applications for research that involves putting human pluripotent stem cells—whether IPS cells or cells from human embryos—into early embryos of nonhuman vertebrates. After protest from some researchers, the agency in 2016 proposed lifting the broad prohibition while keeping a funding ban on specific chimera experiments, including inserting human stem cells into early nonhuman primate embryos and breeding chimeric animals that may have human egg or sperm cells. The proposal is "still under consideration," according to an NIH spokesperson.

The moratorium "has had a very significant impact on the progress of this field," says Pablo Ross, a reproductive biologist at the University of California, Davis, who does chimera research. "Some of the concerns that are raised are to be taken seriously, but I think we have the tools to do that, and [these concerns] shouldn't prevent us from pursuing this goal."

Because of the slow pace of chimera research, even some of its proponents predict that xenotransplantation—the use of nonhuman tissue, such as modified pig organs, for transplants—will beat their approach to the clinic. "Xenotransplantation is close to prime time now," Wu says, and "we are lagging behind." But the possibility of creating organs that are a better match for their human recipients keeps his lab and others poring over stem cells and embryos, hoping to narrow the species divide.