BIOCHEMISTRY OF INTERVERTEBRAL DISC

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Most experts are convinced that degenerating intervertebral discs are at the root of many cases of low-back pain. The intervertebral disc has only just begun to be studied in detail at the molecular level and is still a poorly characterized connective tissue. We propose to characterize in more detail the collagen of human and animal intervertebral discs using the latest biochemical techniques. The emphasis will be an attempt to relate structure to function. An important aim is to understand the characteristic changes in human disc properties with age. The work on human discs will focus initially on autopsy specimens of lumbar discs carefully selected on the basis of histopathological examination to be typically normal for their age, with no gross lesions. Rabbit and cow lumbar discs will also be studied. Molecular species of collagen will be mapped quantitatively within the annulus fibrosus and nucleus pulposus, including the bulk types I and II collagens that are known to be present and the AlphaA, AlphaB, 1Alpha, 2Alpha, and 3Alpha species which may be present. Site-related changes in the distribution of these collagen species will be correlated with variations in the distribution and extractability of proteoglycans and non-collagenous proteins, and with swelling properties of the tissue. Crosslinking residues of collagen will be isolated and quantified, including hydroxypyridinium residues, the mature forms. Pepsin-solubilized collagen will be fractionated into individual species, type I, type II, etc., and the level and type of crosslink in each genetic species and other chemical properties will be measured. Using high performance liquid chromatography (HPLC) to fractionate collagen peptides, any alterations in the distribution of hydroxypyridinium crosslinks with age and location in the disc will be assessed. The molecular packing of annulus and nucleus collagen will be examined by low angle X-ray diffraction and electron microscopy seeking, for example, age-related changes in hydration. Discs removed at surgery to correct scoliosis and other conditions, including spondylolisthesis are on hand and will continue to be available and will also be studied. A collaboration will be initiated with the biomechanics laboratory at Beth Israel Hospital to relate the visco-elastic shear modulus of disc specimens with their composition. In summary, the objective is a better understanding of the relationship between molecular structure, age-related degeneration and mechanical failure.