3D-printing blood vessels with laser-welded sugar

Bioengineers at Rice University showed they could keep densely packed cells alive for two weeks in relatively large constructs by creating complex blood vessel networks from templates of 3D-printed sugar.

“One of the biggest hurdles to engineering clinically relevant tissues is packing a large tissue structure with hundreds of millions of living cells,” says Ian Kinstlinger, a bioengineering graduate student in Rice's Brown School of Engineering. “Delivering enough oxygen and nutrients to all the cells across that large volume of tissue becomes a monumental challenge.”

Nature solved this problem through the evolution of complex vascular networks, which weave through our tissues and organs in patterns resembling tree limbs. The vessels simultaneously become smaller in thickness but greater in number as they branch away from a central trunk, allowing oxygen and nutrients to be efficiently delivered to cells throughout the body.

“By developing new technologies and materials to mimic naturally occurring vascular networks, we're getting closer to the point where we can provide oxygen and nutrients to a sufficient number of cells to get long-term therapeutic function,” Kinstlinger says.

The sugar templates were 3D-printed with an open-source, modified laser cutter in the lab of Jordan Miller, an assistant professor of bioengineering at Rice.

The complex, detailed structures are made possible by selective laser sintering, a 3D-printing process that fuses small grains of powder into solid 3D objects. In contrast to extrusion 3D printing, where melted strands of material are deposited through a nozzle, laser sintering works by gently melting and fusing small regions in a packed bed of dry powder. Both extrusion and laser sintering build 3D shapes one 2D layer at a time, but the laser method enables the generation of structures which would be prone to collapse if extruded.

“There are certain architectures – overhanging structures, branched networks, and multivascular networks – that you really can't do well with extrusion printing,” Miller says. He demonstrated the concept of sugar templating with a 3D extrusion printer during his postdoctoral studies at the University of Pennsylvania. Miller began work on the laser-sintering approach shortly after joining Rice in 2013.

“Selective laser sintering gives us far more control in all three dimensions, allowing us to easily access complex topologies while still preserving the utility of the sugar material,” he says.

© Rice University | https://www.rice.edu

Sugar is useful in creating blood vessel templates because it's durable when dry, and it rapidly dissolves in water without damaging nearby cells. To make tissues, Kinstlinger uses a special blend of sugars to print templates and then fills the volume around the printed sugar network with a mixture of cells in liquid gel. The gel quickly becomes semisolid, and the sugar is then dissolved and flushed away to leave an open passageway for nutrients and oxygen.

“A major benefit of this approach is the speed at which we can generate each tissue structure,” Kinstlinger says. “We can create some of the largest tissue models yet demonstrated in under five minutes.”

The computational algorithm that generated the treelike vascular architectures in the study was created in collaboration with Nervous System, a design studio that uses computer simulation to make unique art, jewelry, and housewares that are inspired by patterns found in nature.

“We're using algorithms inspired by nature to create functional networks for tissues,” says Jessica Rosenkrantz, co-founder and creative director of Nervous System. “Because our approach is algorithmic, it's possible to create customized networks for different uses.”

After creating tissues patterned with these computationally generated vascular architectures, the team demonstrated the seeding of endothelial cells inside the channels and focused on studying the survival and function of cells grown in the surrounding tissue, including rodent liver cells called hepatocytes. The hepatocyte experiments were conducted in collaboration with University of Washington (UW) bioengineer Kelly Stevens, whose research group specializes in studying the delicate cells, notoriously difficult to maintain outside the body.

“This method could be used with a much wider range of material cocktails than many other bioprinting technologies,” Stevens says. “This makes it incredibly versatile.”

“We showed that perfusion through 3D vascular networks allows us to sustain these large liver-like tissues,” Miller says. “While there are still long-standing challenges associated with maintaining hepatocyte function, the ability to both generate large volumes of tissue and sustain the cells in those volumes for sufficient time to assess their function is an exciting step forward.”