Choice of materials is essential to the success of replacement bone created in a 3-D printer and researchers at Johns Hopkins University believe they have come up with the ideal formula.
Their paper published in ACS Biomaterials Science & Engineering calls for a mix at least 30% pulverised natural bone mixed with plastic.
“Bone powder contains structural proteins native to the body plus pro-bone growth factors that help immature stem cells mature into bone cells,” says study senior author Warren Grayson.
“It also adds roughness … which helps the cells grip and reinforces the message of the growth factors.”
Grayson says the scaffold for replacing bone missing by birth defects, trauma or disease is another piece of bone.
“But natural bones can’t usually be reshaped very precisely.”
The aim of Grayson and his team’s study was to find a composite material that would combine the strength and printability of plastic with the biological “information” contained in natural bone.
They began with polycaprolactone, or PCL, a biodegradable polyester used in making polyurethane.
“PCL melts at 80 to 100 °C – a lot lower than most plastics – so it’s a good one to mix with biological materials that can be damaged at higher temperatures,” says Ethan Nyberg, a graduate student on Grayson’s team.
But, despite its strength, PCL does not support the formation of new bone well. So they mixed into the plastic increasing amounts of “bone powder”, made by pulverising the porous bone inside cow knees.
The problem was getting the proportions right. With too much bone powder, there was too little PCL “glue” to maintain a clear shape.
“It was like a chocolate chip cookie with too many chocolate chips,” says Nyberg.
On the other hand, if there was too little bone powder, the material did not encourage bone formation.
To determine that, researchers added human fat-derived stem cells to scaffolds immersed in a nutritional broth.
After three weeks, cells grown on 70%-bone powder scaffolds showed gene activity hundreds of times higher in three genes indicative of bone formation, compared to cells grown on pure PCL scaffolds.
Cells on 30%-bone powder scaffolds showed large but less impressive increases in the same genes.
The scientists also added beta-glycerophosphate to the cells’ broth to enable their enzymes to deposit calcium.
The cells on 30%-scaffolds produced about 30% more calcium per cell, while those on 70%-scaffolds produced more than twice as much calcium per cell, compared to those on pure PCL.
Finally, the team tested their scaffolds in mice with large holes in their skull bones. Mice that received scaffold implants laden with stem cells had new bone growth within the hole over the 12 weeks of the experiment.