A team of biomedical researchers at Wake Forest Institute for Regenerative Medicine has just completed an invention 10 years in the making. It's a 3D printer that can craft relatively simple tissues like cartilage into large complex shapes—like an infant's ear. Using cartridges that are brimming with biodegradable plastic and human cells bound up in gel, this new kind of 3D printer builds complex chunks of growing muscle, cartilage, and even bone. When implanted into animals, these simple fabricated tissues survive and thrive indefinitely.

The scientists led by Anthony Atala surmounted two particularly thorny challenges that have long impeded the futuristic goal of printing living human tissues. First, their new device manufactures large, stable chunks of printed tissue that don't fall apart. Second, it keeps those large structures alive and growing. The new 3D printer is unveiled and outlined today in the journal Nature Biotechnology.

"This is the first bioprinter that can print tissue at the large scales relevant for human implantation."

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"This is the first [bioprinter] that can print tissue at the large scales relevant for human implantation," Atala says. "Basically, once we've printed a structure, we can keep it alive for several weeks before we implant it. Now the next step is to test these [printed tissues] for safety so we can implant them in the future in patients."

Blood and Plastic

To those familiar with 3D printing, Atala's new device—named the Integrated Tissue and Organ Printing System, or ITOP—is straightforward. The programmed printer slowly squirts out layer upon layer of a rapidly hardening material in the form of tiny droplets. Like other 3D printers, this layered approach allows ITOP to print highly complex shapes in three dimensions with incredible detail. The materials ITOP uses, and the way it structures the tissues that it builds, are what make this machine revoutionary.

The Integrated Tissue-Organ Printing System at work printing a jaw bone structure. Wake Forest Institute for Regenerative Medicine

Atala's 3D printer can inject cells suspended in gel, and those cells can be anything from mouse muscle to rabbit cartilage to human stem cells filtered from amniotic fluid. The key to the machine's success, however, is that it combines those cells with another material, a biodegradable plastic called polycaprolactone. Like an itty-bitty scaffold, this plastic keeps the printed tissue around it structurally sound as it is being fabricated and as the growing cells take root. Later on, it degrades away.

"This is very important. This process allows the tissues we print to keep the structural integrity necessary to implant inside the body," Atala says. "Basically we're printing a thread of hard [plastic], then a bead of these soft cells intermittently. So: hard, soft, hard, soft."

Completed ear and jaw bone structures printed with the Integrated Tissue-Organ Printing System. Wake Forest Institute for Regenerative Medicine

Scaling Up

This isn't the first bio-3D printer, but previous devices have been quite limited by the size of the living tissue they can create. That's because most living tissues need an influx of blood and nutrients to stay healthy, but 3D tissue printers aren't nearly advanced enough to print complex features like arteries and blood vessels. So by printing, say, a thick sphere of cartilage, you'd be sure to starve the cells trapped inside. According to Atala, until today the largest printed structure that could be kept alive without blood-cells was only 0.007 inches thick. That's about twice the size of a grain of salt.

ITOP, though, gets around this size limitation by fabricating a latticework of microscopic valleys into the bone, muscle, and cartilage it prints. Supported by the biodegradable plastic scaffolding, these valleys allow nutrients and blood to flow in, keeping tissues alive for months before they're implanted, and "indefinitely," after implantation, Atala says.

"This process allows the tissues we print to keep the structural integrity necessary to implant inside the body."

The team has shown that ITOP can print a impressive suite of living materials. In their demonstrations, the scientists crafted infant-sized ears of cartilage, chunks of replacement jawbone and shards of skull bones, and strips of muscle. Although these tissues are one-dimensional in that they're all crafted from stem cells that develop into a variety of different cell types, "the tissues mature to the same level as normal tissue would," says Atala.

The team has now implanted some of this bone, cartilage, and muscle tissue to great success. But not into humans—yet. In a weird throwback to the Vacanti mouse, the researchers printed an infant's ear of rabbit cartilage under the skin of a mouse, and let it grow. Months after implantation, the cartilage still looks just like an ear, and the mouse's body had started to attach and grow blood-vessels on it. A similar process happened with rat-implanted bone printed from human stem cells: Veins started to take hold.

The Integrated Tissue-Organ Printing System at work printing a jaw bone structure. Wake Forest Institute for Regenerative Medicine

Two weeks after implanting a strip of printed muscle in a rat, Atala's team found that it likewise started to thrive. While it only weakly responded to electric prodding (the researchers liken it to immature, developing muscle) the muscle started to grow both blood vessels and nerves.

For now, Atala's team is testing the long-term safety of ITOP's approach as a run-up to developing tissues for human implantation. In the future, the researchers hope to work with stem cells taken directly from a patient. But Atala believes that for some simple tissues like bone, the theoretical barriers for 3D printing customized replacement parts (say, a new kneecap, hot of the press) may be barriers no more.

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