Advances in bone tissue engineering increasingly focus on biomaterials that enhance osteoblast function, which is essential for effective bone regeneration [1]. This study investigates the effect of varying surface-bound protein concentrations on osteoblast adhesion and proliferation using Ti-6Al-4V, a titanium alloy widely recognized for its biocompatibility and mechanical strength in orthopedic applications. 

Aiming to optimize conditions for cellular activity and matrix formation, we employed self-assembling human serum albumin to create a gradient of surface-bound protein concentrations on Ti-6Al-4V substrates. Surface characterization assessed roughness (AFM), morphology (SEM), hydrophilicity (water contact angle), and composition (XPS). Albumin adsorption was quantified via the Bicinchoninic Acid assay. Results showed that varying protein concentrations slightly influenced surface hydrophilicity while maintaining consistent surface roughness.

In vitro analysis revealed that osteoblasts respond distinctly to different levels of surface-bound protein concentration on the Ti-6Al-4V surface. Variation in protein concentrations led to different level of cell proliferation and the expression of osteogenic markers, suggesting an interacting effect between the protein modified surface and the cells. The morphology of the cells also varied at different surface-bound protein concentrations. Surface chemistry analysis and albumin adsorption suggest that differences in osteogenic marker expression and cellular viability were primarily attributable to protein concentration variations. These findings underscore the importance of tuning surface-bound protein concentration in the design of surface coatings to support bone regeneration and present a bioactive, adjustable method to optimize osteoblast response on metal-based substrates.

This research demonstrates the potential for precise surface-bound protein concentration adjustments on Ti-6Al-4V in developing orthopedic biomaterials, highlighting a sustainable strategy for enhancing cellular function and bone tissue formation. Future studies will focus on more detailed evaluations and the potential for combining these protein-functionalized Ti-6Al-4V surfaces with other biomolecules to amplify osteogenic effects.

Acknowledgement: The authors gratefully acknowledge support of the Deutsche Forschungs-gemeinschaft (DFG, German Research Foundation) – 444711651, RTG 2723 Materials-Microbes-Microenvironments (M-M-M: Projects B). JSMC Jena School for Microbial Communication 

References
[1] Manzini, B.M., Machado, L.M.R., Noritomi, P.Y. et al. Advances in Bone tissue engineering: A fundamental review. J Biosci 2021, 46, 17.