Every day an average of 18 people die waiting for an organ transplant in the United States. Donated organs are tough to come by, which is why many scientists have spent the last two decades trying to create new livers, kidneys, hearts or lungs from scratch. One potential way to craft such delicate structures is 3-D printing with biologically compatible materials, or bioprinting—which has now reportedly produced functional models of lung and liver tissues, with a little help from an unconventional ingredient: food dye.
Would-be organ printers previously have been stymied by the complexity of certain organs. Our lungs and livers, for example, contain physically and biochemically entangled networks of blood vessels and airways (in the lung) or bile ducts (in the liver). Being able to recreate this vasculature—and make the fluid dynamics work so blood and other fluids flow properly—has been an ongoing challenge.
Now, a team of researchers from the University of Washington and Rice University say they have produced functional tissue models using a 3-D printing technique called projection stereolithography. This method exposes thin layers of liquid resin to blue light, which solidifies them into intricate arrangements of hydrogels—gels made up of tangled strings of polymer molecules. These form a structural “scaffolding,” into which researchers can implant live cells that enable it do the work of a lung or liver. In the new study the implanted cells survived, and the resulting models of organ tissue demonstrated some functions of the real thing. The results were published last week in Science.
“This is definitely a major advance in our ability to create 3-D-printed structures that approximate normal tissue,” says Anthony Atala, the director of the Wake Forest Institute for Regenerative Medicine, who was not involved in the new study.
The basic technology of projection stereolithography has been around since the 1980s, but “it wasn’t designed with biology in mind; it was used to make plastic structures,” says Jordan Miller, assistant professor of bioengineering at Rice’s Brown School of Engineering and a co-author of the new paper. The technique can produce finer layers than standard 3-D printing, and is faster, too. “Instead of creating one layer in minutes by extrusion, we can do it in seconds” with stereolithography,” Miller says. That speed is crucial: since the printed structure ultimately channels oxygen and nutrients to the cells, faster work means fewer cells die in the process of making it.
But there was a challenge. This type of printing process relies on photoreactor chemicals (ones that respond to light), so that certain preprogrammed areas of the liquid will solidify while other areas remain soft and can later be washed away. Unfortunately, many of these chemicals are carcinogenic. For a 3-D printer to create the fine vasculature an organ requires for nutrient delivery and waste removal, it needs the precision offered by stereolithography; but for transplants it would need safe, water-soluble photoreactors.
So, the researchers had to find a replacement for the proven but toxic chemicals. When Miller and his team guessed food dye might do the trick—they knew it would absorb the right light wavelengths to make the 3-D printing process work, and is relatively biocompatible—they were too impatient to wait for a supplier to ship the ingredient. So, Miller says, “I went to the supermarket, and I bought a kit of food coloring dye that people use to make confectionery.”
It worked. First, the team colored liquid polymers with the food dye yellow no. 5, or tartrazine, and then had the printer’s projector shine blue light on it. This induced a local chemical reaction that solidified the liquid. Because the printer projected light in a preprogrammed pattern, it created a design that hardened into a thin but tough biological structure. “We were screaming with joy, because it was stunning how simple an idea it was; it immediately enabled us to make this dramatically more complex architecture,” Miller says.
Yellow no. 5, found in many snack foods, had another advantage: It easily rinsed off the bioprinted structures, leaving a clear framework ready to nurture whatever cells the scientists filled it with. The traces of the dye that remained were not expected to affect cell health. (Studies suggest yellow no. 5 does not affect sperm count, as has been rumored; it might, however, exacerbate preexisting hyperactive disorders in children.)
To the Test
Although researchers have bioprinted tissues before, they have been unable to keep cells alive long enough. The latest study had to test the newly printed scaffolding in this regard, and red blood cells were a simple way to start.
The team created a scale model of an air sac mimicking a crucial part of a lung’s complex vascular network. It included one passage for air and separate channels for blood cells. In a healthy human lung, these two structures exchange oxygen without ever touching. The model performed the same feat, keeping the blood cells alive. It also proved sturdy enough to retain its structure as a simulated “breath” expanded and contracted the printed tissues.
Next, the researchers tested a model of liver tissue. Part of the printing process here included injecting specialized liver cells called hepatocytes into the printed structure. The team implanted the artificial liver tissues into live mice with chronic liver injury, as well as non-injured mice, then tested them. A fully functioning liver has over 500 functions and in this case they examined just one, but it did prove successful—and the hepatocytes survived in the living mice.
The new printing method also produced working intravascular valves, which play a key role in the heart and leg veins. In tests the printed versions maintained their structure as fluid flowed through them, and they kept it from moving backward through the valves.
Printed Organs Open Up
So how long until bioprinted organs are available to those on transplant lists? Scientists still have a lot to figure out—starting with the basics, such as determining the optimal base hydrogel. What kind of protein works best? And should additives such as growth factors be used to speed the process? “Now we can start methodically varying these factors to see which are more important—and asking how this affects the functions of the cells,” says paper co-author Kelly Stevens, an assistant professor in the University of Washington’s bioengineering and pathology departments. Then there is the question of how best to build the scaffolding, and how much printed material could realistically replace tissue. “Those are questions that this new leap in tech enables us to ask for the first time,” Stevens says.
The researchers did not want to be the only ones trying out these possibilities—so they made their technology open-source, allowing other bioengineers to test their own applications. “Bioprinting being open-source really helps to accelerate this technology—it really advances the field faster,” Atala says. He plans on applying the findings to a number of organ tissue structures his team is working on.
Other would-be organ-builders can buy specialized printers and inks—Miller and some of the paper’s other collaborators have founded a start-up, called Volumetric, to sell these materials—or can replicate the work themselves. Miller says sharing the DIY option was important to him. “We really are excited,” he says, “about opening up a new set of design freedoms in bioprinting.”