07:58pm Wednesday 23 August 2017

Molecule Boosts Blood-Forming Stem Cells

  • Bone marrow or cord blood transplants can help restore damaged immune systems in patients whose blood cells have been depleted by cancer or other diseases.
  • The mortality rate for such transplants is still very high.
  • Scientists have been searching for chemicals or cell culture methods that can amplify donor stem cells prior to transplant.
  • HHMI scientists have identified a specific lipid molecule that improves the ability of blood-forming stem cells to replenish the immune system after a transplant in both zebrafish and in laboratory mice.

A conceptual view of the competitive transplantation system used in the study. Blood stem cells were taken from zebrafish whose marrow was colored either red or green. The green cells were treated with different test chemicals, while the red cells were untreated. A mixture of the two was injected into other groups of zebrafish to see which color grafted better. The yellow fish show marrow of equal parts red and green; had the chemical worked, the marrow would have been predominantly green. (Credit: Ellen van Rooijen and Pulin Li)

After screening hundreds of chemicals, Howard Hughes Medical Institute (HHMI) scientists have found that treating blood-forming stem cells with a small lipid molecule called EET improves the cells’ ability to replenish the immune system after a transplant in both zebrafish and in laboratory mice. The finding suggests that EETs might one day improve the outcomes of bone marrow or cord blood transplants for patients.

“These molecules are able to enhance the transplantability of the marrow,” says Leonard Zon, an HHMI investigator at Boston Children’s Hospital who led the research. “Our work illustrates that small inflammatory lipids can have prominent effects on stem-cell homing, self renewal, and engraftment.” Zon and his colleagues published their findings July 23, 2015, in the journal Nature.

Bone marrow or cord blood transplants are used to help restore damaged immune systems in patients whose blood cells have been depleted by cancer, chemotherapy, or other diseases. But the procedure has serious risks. Zon says as many as 25 percent of recipients do not survive the transplant. “There’s a great need to make this process safer,” he says. “Being able to deliver more stem cells and more effective stem cells would really make a dent in that mortality rate.”

Researchers have been searching for chemicals or cell culture methods that can help, focusing largely on ways to amplify donor stem cells prior to transplant. In 2007, Zon’s lab demonstrated that one such chemical currently being evaluated in clinical trials, prostaglandin E2, can boost production of blood-forming stem cells.

But there’s room for improvement after transplant, too, Zon says. It can take many weeks for transplanted stem cells to reach the bone marrow and begin to make healthy blood cells, a process called engraftment. Zon and his colleagues wanted to look for compounds that enhance this process.

In their earlier work, Zon’s team had discovered prostaglandin E2 by monitoring stem cell production in zebrafish embryos, which are a powerful tool for studying development and disease. Now, they have devised a strategy to monitor transplanted stem cells directly in adult fish.

Their plan was to transplant two sets of bone marrow cells into the same fish – one set of cells treated with a potential transplant-enhancing chemical, and one left untreated. They watched to see which population of cells engrafted best. The cells would be easy to follow because the scientists would take them from fish that had been genetically engineered to produce fluorescent proteins and transplant them into fish that were entirely transparent. After the transplant, the marrow cells – and any blood cells derived from them — would glow red and green inside the recipient fish.

The team screened 480 compounds for their effects on transplantation. Graduate student Pulin Li performed the zebrafish transplants, a monumental effort involving 220 transplants every day for many months, Zon says. For each test, Li treated marrow cells from a green fish with one of the compounds for four hours. She then mixed the treated green cells with untreated red marrow cells from another donor fish, and injected the mixture into a fish whose own marrow had been eliminated by radiation. Over time, the fluorescent cells found their way to the kidneys – the natural home for marrow cells in a fish.

Four weeks after the transplant, the scientists compared the amount of red and green fluorescence in the kidneys. “We can just shine a fluorescent light on the kidney and see how much red and green is there,” Zon explains. “What we were looking for is a chemical that increased the green-to-red ratio.”

Ten of the compounds in the screen significantly boosted that ratio, indicating that the treatment prior to transplant had improved the cells’ ability to engraft. Two compounds called epoxyeicosatrienoic acids, or EETs, had particularly prominent effects. Like prostaglandins, EETs are small lipids that can promote inflammation. In the body, both EETs and prostaglandins are derived from the same precursor molecule, arachidonic acid. The scientists decided to investigate the EETs further.

In a series of chemical and genetic experiments, Li, graduate student Jamie Lahvic, and postdoctoral fellow Vera Binder showed that EETs activate a signaling pathway that ultimately turns on a gene called runx, a master regulator of blood stem cells during development.

Finally, the researchers set up competitive bone marrow transplants in mice. EET treatment enhanced engraftment at four weeks post-transplant. What’s more, when the researchers analyzed four different types of blood cells 24 weeks after transplanting a mixture of EET-treated and untreated marrow, they found that the majority of those cells were derived from the EET-treated marrow. “We were able to demonstrate a long-term multi-lineage engraftment,” Zon explains.

Zon says EETs seem to aid the stem cells’ ability to travel to the marrow and engraft in a transplant recipient, and may also enhance their self renewal. A next step, he says, will be to test whether the compound has similar effects on human blood stem cells when transplanted into immunodeficient mice. This would be an important step before a clinical trial, he notes.

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Jim Keeley
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