Brillouin spectroscopy and radiography for assessment of viscoelastic and regenerative properties of mammalian bones

Abstract

Biomechanical properties of mammalian bones, such as strength, toughness, and plasticity, are essential for understanding how microscopic-scale mechanical features can link to macroscale bones' strength and fracture resistance. We employ Brillouin light scattering (BLS) microspectroscopy for local assessment of elastic properties of bones under compression and the efficacy of the tissue engineering approach based on heparin-conjugated fibrin (HCF) hydrogels, bone morphogenic proteins, and osteogenic stem cells in the regeneration of the bone tissues. BLS is noninvasive and label-free modality for probing viscoelastic properties of tissues that can give information on structure-function properties of normal and pathological tissues. Results showed that MCS and BPMs are critically important for regeneration of elastic and viscous properties, respectively, HCF gels containing combination of all factors had the best effect with complete defect regeneration at week nine after the implantation of bone grafts and that the bones with fully consolidated fractures have higher values of elastic moduli compared with defective bones.

Keywords: Brillouin light scattering; biomechanical properties; bone morphogenic proteins; bones; critical-sized defect; elastic; heparin-conjugated fibrin gel; viscous.

Fig. 1

Photographs of the bone samples. Bovine, ovine, and chicken tibia bones.

Fig. 2

Photographs of the surgical procedures and ulna bones of the rabbits. (a) Section of the radius before resection, (b) resection of the bone with an oscillating saw, (c) created critical-sized defect, (d) implantation of the bone grafts with various combinations of the HCF gels, MSCs and BMPs into the defect area, (e) ulna bones of the rabbits after the sacrifice, and (f) location of the point-measurement spot on the ulna bone.

Fig. 3

Schematic representation of the confocal BLS spectroscopy setup. Tandem Fabry–Perot interferometry coupled to confocal microscopy for spontaneous Brillouin microspectroscopy.

Fig. 4

(a) Brillouin spectra acquired for the ovine tibia bone, (b) the Brillouin peak’ positions for bovine, ovine, chicken tibia, and ulna bones under the compression load. Error bars represent a 95% confidence interval based on the standard deviation. Corresponding breaking limits are shown with vertical dashed lines, and (c) FWHM obtained from the spectra as a function of compression load for bovine, ovine, chicken tibia, and rabbit ulna bones.

Fig. 5

Dynamics of the bone defect regeneration after HCF implantation with MSCs, BMP-2, and BMP-7.

Fig. 6

The Brillouin peaks obtained from the spectra as a function of compression load for the different bone grafts (a) HCF gel with MSCs, (b) HCF gel with MSCs, BMP-2, and BMP-7, (c) HCF gel with BMP-7, and (d) HCF gel with BMP-2 and BMP-7.

Fig. 7

FWHM obtained from the spectra as a function of compression load for the different bone grafts (a) HCF gel with MSCs, (b) HCF gel with MSCs, BMP-2, and BMP-7, (c) HCF gel with BMP-7, and (d) HCF gel with BMP-2 and BMP-7.