The lung is a feathery web of blood vessels and air sacs that branch closely together, but never touch. They’re nestled near enough that oxygen can diffuse into the bloodstream, but far enough that blood and air flow through separate pipes.
If researchers are to have any hope of creating lab-grown lungs or other organs, they’ll have to find a way to recreate their complex architectures using 3-dimensional bioprinting. In a recent study in Science, researchers report they were able to overcome the limitations of conventional bioprinting techniques in order to 3-D print multivascular networks in soft hydrogels. The ability to fabricate complex biological structures should hasten ongoing efforts to bioprint organ transplants made of hydrogel and human cells.
Before this study, a widely used and rapid 3-D printing technique—stereolithography—couldn’t make complicated structures, explains bioengineer Jordan Miller, of Rice University in Houston Texas. Its limitation was baked in: stereolithography uses focused blue light to pattern biological forms into nontoxic gels. The light polymerizes the hydrogel layer-by-layer. But as the layers build up, light can still penetrate into deeper, finished regions, continuing to polymerize them and ruining past work.
“You need something to block the light,” explains mechanical engineer Michael McAlpine of the University of Minnesota in the Twin Cities, who was not involved in the new work. Most dyes known to do that are toxic to living cells, he says, ruling them out of efforts to recreate living tissues.
Blocking light penetration with a nontoxic dye is the major innovation of this new study, McAlpine says. To do it, Miller and bioengineer Kelly Stevens at the University of Washington in Seattle found a nontoxic photoabsorber. They tried several, including derivatives of turmeric and blueberries mixed into the gel, and found the most effective was surprisingly simple: tartrazine, or the synthetic yellow food coloring known as Yellow 5. Adding food coloring to the gel unlocked complex design possibilities, including structures inside a single vessel (such as valves) and multivascular networks (such as the airway and bloodstream in the lungs).
“If they just published a paper on that, that would be an amazing paper,” McAlpine explains. But that was just the start.
After identifying a nontoxic dye, the researchers used it to bioprint a 3-D air sac surrounded by a net of hollow tubes, mimicking the close-but-separate vessel networks in the lung. No bigger than a penny, the model looks like a raspberry with a “breathing” air sac at its core. “More or less for the first time, we’re able to construct these complexities in a bioprinted manner,” Stevens says. “In the past it’s been very hard to fabricate these networks like they are in the body, very close to each other but not touching.”
The lung model is proof-of-concept that bioengineers can print unprecedentedly-complex structures using gel and food dye. “They took it to a whole new level by creating a model lung,” McAlpine says. “It’s mind-blowing they were able to accomplish all these things in one paper.”
The team inflated and deflated the air sac while running human blood cells through the network of hollow hydrogel tubes surrounding it, and found that oxygen diffused out of the airway for uptake by red blood cells, just as in real lungs, showing these models can replicate features of organs in a soft gel. Finally, the researchers implanted gels containing living cells into mice. After two weeks, the cells were still alive and functional, suggesting implantation studies are viable with these materials.
The new study is a huge leap forward for tissue engineering in regenerative medicine, but “there’s still quite a ways to go before printing perfect vascular structures,” Stevens says. One next step: going even smaller, explains Cole DeForest of the chemical engineering and bioengineering departments at the University of Washington in Seattle, who was not involved in the new work. Vessels come in a range of sizes, including some that make a penny-sized lung model look huge. “An exciting future advance,” DeForest explains by email, “would be to ‘miniaturize’ these techniques such that smaller microvessels including capillaries (~10 micron diameter) could be printed with the same 3-D control.”