Researchers at Nanyang Technological University, Singapore (NTU Singapore) have developed a new biomaterial made entirely from discarded bullfrog skin and fish scales that could help in bone repair.
The porous biomaterial, which contains the same compounds that are predominant in bones, acts as a scaffold for osteoblasts, or bone-forming cells, to adhere to and multiply, leading to new bone formation. Bone-forming cells successfully latched onto the biomaterial and started growing, and it was found to have a low inflammatory risk.
This kind of scaffold could help regenerate bone tissue lost to disease or injury, such as jaw defects from trauma or cancer surgery. It could also assist bone growth around surgical implants such as dental implants.
The current standard practice of using a patient’s own tissues means extra surgery is needed for bone extraction. The biomaterial used, frog skin and fish scales, are a significant waste stream produced by Singapore’s aquaculture industry and using them helps repurpose this waste.
‘Waste-to-resource’
“We took the ‘waste-to-resource’ approach in our study and turned discards into a high-value material with biomedical applications, closing the waste loop in the process,” said Dalton Tay, Assistant Professor, Nanyang Technological University. “Our lab studies showed that the biomaterial we have engineered could be a promising option that helps with bone repair. The potential for this biomaterial is very broad, ranging from repairing bone defects due to injury or ageing, to dental applications for aesthetics. Our research builds on NTU’s body of work in the area of sustainability and is in line with Singapore’s circular economy approach towards a zero-waste nation.”
To make the biomaterial, the team first extracted Type 1 tropocollagen (many molecules of which form collagen fibres) from the discarded skins of the American bullfrog and hydroxyapatite (a calcium-phosphate compound) from the scales of snakehead fish, commonly known as the Toman fish.
Collagen and hydroxyapatite (HA) are two predominant components found in bones, thus conferring on the biomaterial a structure, composition, and ability to promote cell attachment similar to bone, as well as toughness.
The scientists removed all impurities from the bullfrog skin, then blended it to form a thick collagenous paste that is diluted with water, from which collagen was extracted. “Using this approach, we were able to obtain the highest ever reported yield of collagen of approximately 70 per cent from frog skin, thus making this approach commercially viable,” said Asst Prof Tay, who is also from the NTU School of Biological Sciences (SBS).
HA was harvested from discarded fish scales through calcination – a purification process that requires high heat – to remove the organic matter, and then air-dried.
The biomaterial was synthesised by adding HA powder to the extracted collagen, then cast into a mould to make a 3D porous scaffold — a two-week process which the team believes can be shortened.
Testing the biomaterial
To assess the biological performance of the porous biomaterial scaffold for bone repair, the scientists seeded bone-forming cells onto the scaffold.
The cells proliferated, and after a week, the cells were uniformly distributed across the scaffold – an indicator that the scaffold could promote proper cellular activities and eventually lead to tissue formation. The scientists also found that the presence of HA in the biomaterial significantly enhanced bone formation.
The biomaterial was also tested for its tendency to cause an inflammatory response, which is common after a biomaterial is implanted in the body.
Using real-time polymerase chain reaction, the scientists found that the expression of pro-inflammatory genes in human immune cells exposed to the biomaterial stayed “relatively modest” compared to a control exposed to endotoxins, a compound known to stimulate immune response, said Asst Prof Tay.
For instance, the expression of the gene IL6 in the biomaterial group was negligible and at least 50 times lower than that of the endotoxins-exposed immune cells. This suggests that the risk of the NTU-developed biomaterial to trigger an excessive acute inflammatory response is low.
The team is now further evaluating the long-term safety and efficacy of the biomaterial as dental products. Further research would involve studying how the body responds to this biomaterial in the long term, as well its use in other applications such as skin wounds, along with further development of the waste-to-resource pipeline.
A preprint copy of the article is available as a PDF for download.
Source: Nanyang Technical University
