Liver organoid generates organ-specific blood vessels for the first time

Scientists from Cincinnati Children’s and colleagues based in Japan report achieving a major step forward in organoid technology: producing liver tissue that grows its own internal blood vessels.

This significant advance could lead to new ways to help people living with hemophilia and other coagulation disorders while also taking another step closer to producing transplantable repair tissues for people with damaged livers.

The study, led by Takanori Takebe, MD, Ph.D., director for commercial innovation at the Cincinnati Children’s Center for Stem Cell and Organoid Research and Medicine (CuSTOM), was published in Nature Biomedical Engineering.

Co-authors included researchers from the Institute of Science Tokyo, the Icahn School of Medicine at Mount Sinai, and Takeda Pharmaceutical Company.

“Our research represents a significant step forward in understanding and replicating the complex cellular interactions that occur in liver development. The ability to generate functional sinusoidal vessels opens up new possibilities for modeling a wide range of human biology and disease, and treating coagulation disorders and beyond,” Takebe says.

What are organoids?

For more than 15 years, researchers at Cincinnati Children’s and many other institutions have been working to grow human organ tissue in the laboratory. Such tissues have already become important tools for medical research and may soon become sophisticated enough to be used directly to help repair damaged organs.

The complex process involves placing induced pluripotent stem cells (iPSCs) in special gels designed to prompt the stem cells to grow into specific tissue types. The stem cells can be generic or come from specific individuals with health conditions and can be gene-edited before beginning the process.

Cincinnati Children’s has been a leader in organoid research since 2010 when experts here developed the first functional intestinal organoid grown from iPSCs. Since then, CuSTOM has grown and evolved to include 37 labs across 16 research divisions, where teams are improving organoid technology and using organoids to shed new light on a wide range of diseases and conditions.

Overcoming a challenge

Until recently, the size of lab-grown organoids has been fundamentally limited because they have not included important tissues that connect organs to the rest of the body, such as nerves and blood vessels.

This study recounts how the research team overcame the blood vessel obstacle. The experiments involved required nearly a decade to complete.

Ultimately, the project succeeded at differentiating human pluripotent stem cells into CD32b+ liver sinusoidal endothelial progenitors (iLSEP). Then the team used an inverted multilayered air-liquid interface (IMALI) culture system to support the iLSEP cells as they self-organized into hepatic endoderm, septum mesenchyme, arterial, and sinusoidal quadruple progenitors.

The advantage of using the iLSEP progenitor cells as building blocks is that they are specific to the liver. Some other studies seeking to add vascularization to organoids have depended upon “fully committed” arterial endothelial cells. These vessels may not function inside an organ as well as progenitor cells from that organ.

Location and timing were also crucial to achieving the initial vessel formation.

“The success occurred in part because the different cell types were grown as neighbors that naturally communicated with each other to take their next development steps,” says the study’s first author, Norikazu Saiki, Ph.D., of the Institute of Science Tokyo.

The new method produced “perfused blood vessels with functional sinusoid-like features,” which means the vessels were fully open and included the pulsing cell types needed to help blood move through. The advanced organoids also generated the correct cell types needed to produce four types of blood coagulation factors, including Factor VIII, which is missing among people with hemophilia A. In mice that mimic hemophilia, the study showed that organoid-derived Factor VIII rescued them from severe bleeding.

By developing IMALI culture methods for allowing multiple cell types to self-organize naturally, the new technology may open a possibility to grow organ-specific vesselsin other types of organoids.

Big step closer to improved treatments for hemophilia, liver failure

In the U.S. an estimated 33,000 males live with hemophilia. Most have hemophilia A (factor VIII deficiency), while a smaller group has hemophilia B (factor IX deficiency).

The condition can cause repeated bleeding within joints that can lead to chronic pain and mobility limitations. Hemophilia makes surgery risky and other wounds harder to heal. It can also lead to seizures and paralysis when bleeding affects the brain.

Hemophilia is treated by injecting commercially prepared concentrates to replace the missing coagulation factors. However, human blood contains a dozen different clotting factors and there are no available human protein sources for missing coagulation factors V or XI. Also, about 20% of people with hemophilia A develop inhibitors to standard treatment products.

“These advanced liver organoids can secrete these coagulation factors. If they can be produced at scale, they could become a viable treatment source that would benefit people who have developed inhibitors or are not indicated for gene therapy,” Takebe says.

Meanwhile, people experiencing acute or chronic liver failure also do not produce adequate supplies of coagulation factors, placing them at higher risk of bleeding complications during surgery. A factor-secreting organoid “factory” could also help these patients.

Longer-term, increasingly sophisticated liver organoids may eventually supply repair tissues that can help diseased livers heal themselves.

Cincinnati Children’s co-authors on this study included Kentaro Iwasawa, MD, Ph.D., and Wendy Thompson, Ph.D. The Integrative Morphology Core and Pluripotent Stem Cell and Organoid Core at Cincinnati Children’s contributed.