Abstract
Alterations to the biochemical composition of the intervertebral disc (IVD) are hallmarks of aging and degeneration. Methods to assess biochemical content, such as histology, immunohistochemistry, and spectrophotometric assays, are limited in their ability to quantitatively analyze the spatial distribution of biochemical components. Fourier transform infrared (FTIR) microscopy is a biochemical analysis method that can yield both quantitative and high-resolution data about the spatial distribution of biochemical components. This technique has been largely unexplored for use with the IVD, and existing methods use complex analytical techniques that make results difficult to interpret. The objective of the present study is to describe an FTIR microscopy method that has been optimized for imaging the collagen and proteoglycan content of the IVD. The method was performed on intact and discectomized IVDs from the sheep lumbar spine after 6 weeks in vivo in order to validate FTIR microscopy in healthy and degenerated IVDs. FTIR microscopy quantified collagen and proteoglycan content across the entire IVD and showed local changes in biochemical content after discectomy that were not observed with traditional histological methods. Changes in collagen and proteoglycans content were found to have strong correlations with Pfirrmann grades of degeneration. This study demonstrates how FTIR microscopy is a valuable research tool that can be used to quantitatively assess the local biochemical composition of IVDs in development, degeneration, and repair.