Scientists have discovered a method to induce human endothelial cells from a small biopsy sample to multiply in the laboratory, producing more than enough cells to replace damaged blood vessels or nourish organs for transplantation, according to a preclinical study by Weill Cornell Medicine investigators. Endothelial cells form the inner lining of blood vessels and regulate blood flow, inflammation and healing. Traditional approaches for growing these cells in the lab have yielded only limited numbers before they lose their ability to function. The new method involves treating adult endothelial cells with a small molecule that triggers the hibernating cells to wake up and divide hundreds of times without signs of aging, mutation or loss of function.
The findings, published Oct. 14 in Nature Cardiovascular Research, may provide a reliable way to generate an enormous number of a patient's own endothelial cells, enabling vascular grafts for heart disease, diabetes treatments and organ transplants and strategies to target abnormal tumor blood vessels.
"Despite being able to isolate human endothelial cells in the lab for more than 50 years—a breakthrough that was originally reported by scientists at Weill Cornell in 1973—their use for human therapy has been cumbersome. It has been difficult to cultivate them in clinical quantities sufficient for vascularizing human organs," said Dr. Shahin Rafii , director of the Hartman Institute for Therapeutic Organ Regeneration and the Ansary Stem Cell Institute, chief of the division of regenerative medicine and the Arthur B. Belfer Professor in Genetic Medicine at Weill Cornell, who led the research. "Our technology allows clinical labs to use a small biopsy taken in a doctor's office and ultimately produce a trillion or more functional endothelial cells, without acquiring any aberrant features."
Dr. Yang Lin , instructor of regenerative medicine at Weill Cornell, and Dr. Fuqiang Geng , research associate in the Rafii lab, also co-led the research.
Rise and Shine
Human organs require a healthy network of blood vessels to heal and regenerate but producing the endothelial cells necessary for this has been tricky. Even young, healthy endothelial cells from discarded human umbilical cords become unstable and senescent after about eight cell divisions.
Adult endothelial cells are even less efficient at multiplying because a few signaling pathways keep a large subset of the cells dormant. One of these is the aryl hydrocarbon (AH) receptor pathway that regulates gene expression in the nucleus. Recently, scientists showed that inhibiting the AH receptor with a class of small molecules triggered hematopoietic stem cells to divide. The researchers hypothesized that this approach may also work with adult endothelial cells.
The team discovered that three different small molecules could block the AH receptor and propagate endothelial cells from various human tissues, including adult fat tissue, an easily accessible harvest site. Culturing a biopsy sample in the presence of an AH receptor inhibitor produced up to 2 trillion endothelial cells, which was 100 times more than the control culture. Even after this substantial expansion, the treated cells had normal gene expression, genomic stability and the potential to form blood vessels. "It's like a fountain of youth—the cells don't age and don't become cancerous, which is critical for clinical therapy," Dr. Rafii said, who is also a member of the Englander Institute for Precision Medicine and the Sandra and Edward Meyer Cancer Center at Weill Cornell.
A Surprising New Pathway
When the team looked further into how the inhibitor-treated cells were able to expand so much, they noticed something unexpected. "We thought that if we knocked down AH receptor gene expression genetically, we should have more proliferation even without the inhibitor, but we found that was not true," Dr. Lin said. "This provided a clue that the small molecule was not inhibiting the known AH receptor pathway but working in a completely different way."
Dr. Lin and Dr. Geng determined that instead of blocking the AH receptor's usual activity, the inhibitors triggered an alternative pathway that changed how the receptor interacted with other proteins involved in metabolism, oxidation and inflammation. The inhibitor decreased levels of harmful reactive oxygen species and allowed the cells to use alternative processes for energy, aside from just using glucose. "That allows them to keep replicating, while safeguarding their genomic integrity," Dr. Rafii said. Further experiments found that the inhibitors activate the production of a polyamine—a molecule vital for normal cell growth and survival—which may be pushing endothelial cells to multiply.
The findings lay the groundwork for generating the building blocks of blood vessels at the scale needed for human therapies. Next, the researchers will dissect how the inhibitor binding to the AH receptor reshapes cell signaling and metabolism in endothelial cells. Most importantly, the investigators plan to leverage this transformative approach to cultivate clinical-scale quantities of tissue-specific blood vessels that are ideal for engineering durable replacement organs.
