Sodium MRI of Intervertebral Disc Degeneration

Objective: To establish sodium MRI as a non-invasive method for measuring intervertebral disc proteoglycan content Introduction: Intervertebral disc (IVD) degeneration is a common and sometimes debilitating condition, incurring tremendous financial cost on society. The initial stage of IVD degeneration involves the breakdown of proteoglycans (PG) in the nucleus pulposus (NP) region of the disc[1]. Currently, IVD degeneration is usually diagnosed using T2–weighted proton MRI, which is not sensitive to changes in PG content. Previous studies have demonstrated that sodium MRI can be used to non-invasively analyze PG content in the articular cartilage [2]. Sodium MRI directly measures the Na concentration ([Na]) of the tissue, which is the primary cation in the NP region of the IVD. The Na cations are attracted by the fixed–charge density on the PG. Thus sodium MRI can be used to estimate the PG concentration ([PG]) of IVD. The purpose of this study is to establish sodium MRI as a non-invasive method for measuring IVD [PG], by comparing [Na] map obtained using sodium MRI and 1,9-dimethylmethylene blue (DMMB) PG assay results. Materials and Methods: Two fresh veal lumbar spines were obtained from a local abattoir within a few hours of slaughter. From each spine specimen, the last two caudal discs on the posterior side were surgically harvested. MRI was performed on a 3T Siemens Trio clinical MRI scanner. Tissue samples were placed inside a custom-made low-pass quadrature birdcage RF coil tuned to sodium resonance frequency at 3T. The dimensions of the RF coil were 17cm in diameter and 12.5cm long, and contained 16 struts. The two receiver ports were inductively coupled to the coil and oriented 90° relative to each other. Five 10% agarose gel phantoms containing 100mM, 150mM, 200mM, 250mM, and 300mM [Na] were imaged alongside each specimen for later [Na] calibration. The vendor’s 3D FLASH pulse sequence was used to acquire all sodium images. Imaging parameters were as follows: TE/TR = 6/30 ms, flip angle = 90, FOV = 15 x 15 cm, matrix size = 128 x 128, slices = 128, slice thickness = 1.2 mm, BW = 60 Hz/Pixel, signal average = 75. These parameters were chosen to obtain a minimum signal-to-noise ratio (SNR) of 15:1 for an isotropic voxel size of 1.2mm. The isotropic voxel size allowed us to extract 2D slices from the 3D image in any orientation. The sodium signals from IVD and phantoms were corrected separately for T1 and T2 * decays. A calibration curve of phantom sodium MRI signal and their known [Na] was used to compute [Na] maps of the IVD. After imaging, IVDs were isolated via sharp dissection. A series of ordered 4-mm diameter punches were harvested from the NP for subsequent DMMB assay, the punch removal sites were indicated in Fig 1. The discs were photographed against a dark background to record the position of the removed punches. These photographs were used later to generate image masks for reporting [Na] values from the exact region where the punches were located. The wet weight of the punches was determined prior to digestion using papain. Digested solutions were diluted and determination of sulfated–glycosaminoglycan content was performed using 1, 9-dimethylmethylene blue (DMMB) in microplate reader assay. PG concentration was calculated from a standard curve of shark chondroitin sulfate C. All data processing and analysis were carried out using algorithms developed with MATLAB software (Mathworks, Natick, MA). Photographs of each dissected disc were interpolated to the same spatial resolution as the sodium MR images (1.2 mm). Intensity thresholding of the photographs resulted in binary masks showing the region of interest (ROI) where punches were removed. The location of each ROI was used to report average [Na] from the maps. An automatic co-registration routine applied step-wise in-plane rotation and translation while optimizing the linear regression fit of [Na] map ROIs and the PG assay results.