Mineral Distributions at the Developing Tendon Enthesis

Andrea G Schwartz,Tyrone L Daulton,Stavros Thomopoulos,Jill D Pasteris,Guy M Genin,Ryan K Roeder

doi:10.1371/journal.pone.0048630

Andrea G Schwartz, Tyrone L Daulton + Show 4 more

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https://doi.org/10.1371/journal.pone.0048630

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Abstract

Tendon attaches to bone across a functionally graded interface, “the enthesis”. A gradient of mineral content is believed to play an important role for dissipation of stress concentrations at mature fibrocartilaginous interfaces. Surgical repair of injured tendon to bone often fails, suggesting that the enthesis does not regenerate in a healing setting. Understanding the development and the micro/nano-meter structure of this unique interface may provide novel insights for the improvement of repair strategies. This study monitored the development of transitional tissue at the murine supraspinatus tendon enthesis, which begins postnatally and is completed by postnatal day 28. The micrometer-scale distribution of mineral across the developing enthesis was studied by X-ray micro-computed tomography and Raman microprobe spectroscopy. Analyzed regions were identified and further studied by histomorphometry. The nanometer-scale distribution of mineral and collagen fibrils at the developing interface was studied using transmission electron microscopy (TEM). A zone (∼20 µm) exhibiting a gradient in mineral relative to collagen was detected at the leading edge of the hard-soft tissue interface as early as postnatal day 7. Nanocharacterization by TEM suggested that this mineral gradient arose from intrinsic surface roughness on the scale of tens of nanometers at the mineralized front. Microcomputed tomography measurements indicated increases in bone mineral density with time. Raman spectroscopy measurements revealed that the mineral-to-collagen ratio on the mineralized side of the interface was constant throughout postnatal development. An increase in the carbonate concentration of the apatite mineral phase over time suggested possible matrix remodeling during postnatal development. Comparison of Raman-based observations of localized mineral content with histomorphological features indicated that development of the graded mineralized interface is linked to endochondral bone formation near the tendon insertion. These conserved and time-varying aspects of interface composition may have important implications for the growth and mechanical stability of the tendon-to-bone attachment throughout development.

Highlights

Tendons and ligaments attach to bone across transitional tissue interfaces that are several micrometers to millimeters in thickness
Microscale mineral gradients at the developing enthesis Raman microprobe analysis demonstrated an increase in mineral content across the murine supraspinatus tendon-to-bone interface at all time points studied
This study investigated the development of the mineralized interface at the tendon-to-bone insertion

Summary

Introduction

Tendons and ligaments attach to bone across transitional tissue interfaces that are several micrometers to millimeters in thickness. The fibrocartilaginous enthesis contains a functionally graded transitional tissue, with variations in extracellular matrix structure and composition giving rise to variations in mechanical properties across the interface [4,5,6,7,8]. Within the transitional zone of the rat supraspinatus tendon enthesis, an increase in mineral relative to collagen has been observed through the transition from tendon to bone [10]. These variations in structural and compositional properties result in graded mechanical behavior that contributes to an efficient transfer of muscle load from tendon to bone [11,12,13]. It is believed that the gradient in mineral content is important for limiting stress concentrations at the mineralized interface

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