3D Raman mapping of the collagen fibril orientation in human osteonal lamellae

Abstract

Chemical composition and fibrillar organization are the major determinants of osteonal bone mechanics. However, prominent methodologies commonly applied to investigate mechanical properties of bone on the micro scale are usually not able to concurrently describe both factors. In this study, we used polarized Raman spectroscopy (PRS) to simultaneously analyze structural and chemical information of collagen fibrils in human osteonal bone in a single experiment. Specifically, the three-dimensional arrangement of collagen fibrils in osteonal lamellae was assessed. By analyzing the anisotropic intensity of the amide I Raman band of collagen as a function of the orientation of the incident laser polarization, different parameters related to the orientation of the collagen fibrils and the degree of alignment of the fibrils were derived. Based on the analysis of several osteons, two major fibrillar organization patterns were identified, one with a monotonic and another with a periodically changing twist direction. These results confirm earlier reported twisted and oscillating plywood arrangements, respectively. Furthermore, indicators of the degree of alignment suggested the presence of disordered collagen within the lamellar organization of the osteon. The results show the versatility of the analytical PRS approach and demonstrate its capability in providing not only compositional, but also 3D structural information in a complex hierarchically structured biological material. The concurrent assessment of chemical and structural features may contribute to a comprehensive characterization of the microstructure of bone and other collagen-based tissues.

A literature review of the spatial organization of lamellar bone

“…This effect would be even stronger if the cut is not exactly parallel to the growth front, which due to the curvature of the lamellae, the fact that bone is a living substance, and inaccuracies during preparations, it will almost never be. This explanation is also consistent with the loss of anisotropy observed in some lamellae by Schrof et al (2014), which they suggest might be due to the difference in lamellar thickness and their own measurement resolution.…”

“…D. Twisted plywood model with a smooth transition from one orientation of collagen fibers to the next. Lamellae, however, are not distinguished (based on Varga et al, 2013 andSchrof et al, 2014). E. Oscillating plywood model, which is similar to D, but with a slight difference in collagen fiber orientations.…”

“…D. Modèle de contreplaqué enroulé, avec transition floue d'une orientation des fibres de collagène à la suivante. Les lamellae ne sont cependant pas distinguées (d'après Varga et al, 2013et Schrof et al, 2014. E. Modèle de contreplaqué oscillant, similaire à D, mais avec une légère différence dans les orientations des fibres de collagène.…”

“…Consequently, it is not clear how the smooth changes in fibril orientation relate to the dark and light bands as seen in polarized light microscopy. Using the same definition as Varga et al (2013) (i.e., a continuous change of fibril orientations without backflip), Schrof et al (2014) found twisted and oscillating plywood in the same osteons.…”

Stochastic multiscale modelling of cortical bone elasticity based on high-resolution imaging

“…While on the one hand our hypothesis of cylindrical pores at the tissue scale seems reasonable (Granke et al 2015), our assumptions at the lower scales (aligned collagen fibres and spherical mineral particles) may lead to inaccurate predictions (Deuerling et al 2009). Other micro-and nanoscale features such as the morphology of the lacuno-canalicular network and the mineralisation at the nanoscale can affect bone elasticity, strength, and failure (Tai et al 2008;Langer et al 2012;Schrof et al 2014). All these microarchitectural features should be investigated and their effects on the tissue level properties assessed.…”