Supplementary material from "Mineral phase changes during intervertebral disc degeneration"

Intervertebral disc disease is a common cause of pain and neurological deficits known to be associated with degeneration and calcification of discs in humans and animals. Here, we analysed samples of herniated disc material and compared it to material taken from non-herniated discs following surgical treatment in dogs. Our clinical approach to these cases allowed collection of approximately 100 samples providing a unique opportunity for a substantial case-controlled study, an opportunity not available to the human neurosurgeon. We analysed all samples using Fourier transform infrared (FTIR) spectroscopy, as well as subsets of 10 samples by X-ray diffraction (XRD) and two samples by transmission electron microscopy (TEM). We show the majority of herniated samples have FTIR spectra consistent with the presence of crystalline hydroxyapatite (HAp), whereas most non-herniated discs showed spectra consistent with amorphous phosphate. XRD and TEM confirmed these findings and the two samples examined with TEM identified the amorphous material as amorphous calcium phosphate (ACP) nanoparticle clusters of approximately 20 nm diameter and the crystalline HAp as polycrystalline needles up to 100 nm in length. Our data suggest that the intervertebral disc degenerative process involves conversion of ACP into crystalline Hap, which might predispose a disc to herniate.