Characterization of recombinant amelogenin-integrated biomaterials using mammalian cells

Abstract

Amelogenin, the predominant protein in enamel formation, has been recognized not only for its role in guiding hydroxyapatite crystal growth but also for its therapeutic potential in periodontal regeneration.
In this study, we investigated the effects of different amelogenin assembly states, using modified amelogenin variants on the proliferation, metabolic activity, and osteogenic differentiation of the murine pre‑osteoblastic cell line M3CT3‑E1. Two distinct biomaterial platforms were employed: rigid surface coatings and alginate hydrogels, to assess how the three-dimensional microenvironment influences cell behavior. In vitro analyses using AlamarBlue assays, ALP activity measurements, and Alizarin Red
mineralization assays revealed that amelogenin assembly could markedly enhance osteoblast proliferation and mineral deposition compared to controls. Furthermore, the kinetic profiles observed
on rigid coatings differed significantly from those on hydrogels. Hydrogels exhibited a delayed yet robust cellular response, likely due to their three-dimensional architecture and diffusion properties that more closely mimic physiological tissue environments. These findings highlight the critical role of both
protein assembly state and scaffold microenvironment in regulating osteogenic differentiation.
Collectively, our results offer valuable insights for the design and development of next-generation biomimetic scaffolds aimed at improving periodontal and bone regenerative outcomes.

Popular Abstract

Nano-Engineered Amelogenin: Blueprint for Next-Gen Bone Regeneration
Imagine if the protein that built your tough tooth enamel could also help repair broken bones. My project
reveals how re-engineering this tiny protein into nanosized structures might be the secret recipe for
next-generation bone regeneration.
Bone healing isn’t always as straightforward as it sounds. Fractures or bone defects can lead to
prolonged pain, non-unions, and cumbersome recovery processes, often requiring invasive surgeries and
expensive treatments. These setbacks are especially common in aging populations or in cases of severe
trauma, where the body’s natural repair mechanisms sometimes fall short.
That’s where nature’s own design comes into play. Amelogenin is the protein responsible for forming
the hard, resilient enamel on our teeth. Inspired by its natural talent for guiding mineral crystal growth,
I set out to explore whether this protein could be re-engineered to kick-start bone repair. In my project,
I transformed recombinant amelogenin into different nanoformations: clumped microaggregates and
long, elegant nanoribbons. We even introduced creative tweaks into the protein to see which formulation
best fuels the bone healing process; think of it as testing various recipes to create the ultimate healing
cocktail!
To test these nano-creations, I grew bone precursor cells (pre‑osteoblasts) on two kinds of “scaffolds.”
One was a classic, hard surface coating, and the other was a squishy, three-dimensional alginate hydrogel
that mimics natural tissue. Simple tests, like checking cell vitality, measuring bone marker activity, and
detecting mineral deposits, told an exciting story. The nanoribbon versions, especially those with our
protein tweaks, orchestrated a symphony of cell growth and mineral buildup. On the rigid surfaces, cells
jumped into action quickly, while in the more life-like hydrogels, they took their time but ultimately put
on an impressive performance.
In essence, this work shows that tiny, well-designed protein structures can transform a simple scaffold
into a dynamic, bone-healing powerhouse. These insights not only deepen our understanding of nature’s
own building blocks but also pave the way for innovative, custom-made therapies that might one day
help our bodies regenerate damaged bone in a truly remarkable way.

• author: Granec, Zuzana
• supervisor: Johan Svensson Bonde
• organization: Pure and Applied Biochemistry
• course: KBKM01 20251
• year: 2025
• type: H2 - Master's Degree (Two Years)
• subject: Technology and Engineering
• keywords: Amelogenin, Biomaterials, Osteoblasts, Hydrogel, Applied biochemistry
• language: English
• id: 9193751
• date added to LUP: 2025-06-13 11:45:46
• date last changed: 2025-06-13 11:45:46