Quantitative magnetic resonance imaging of enzymatically induced degradation of the nucleus pulposus of intervertebral discs

STUDY DESIGN: The structural integrity of the nucleus pulposus (NP) of intervertebral discs was targeted by enzyme-specific degradations to correlate their effects to the magnetic resonance (MR) signal. OBJECTIVE: To develop quantitative MR imaging as an accurate and noninvasive diagnostic tool to better understand and treat disc degeneration. SUMMARY OF BACKGROUND DATA: Quantitative MR analysis has been previously shown to reflect not only the disc matrix composition, but also the structural integrity of the disc matrix. Further work is required to identify the contribution of the structural integrity versus the matrix composition to the MR signal. METHODS: The bovine coccygeal NPs were injected with either enzyme or buffer, incubated at 37 degrees C as static, unloaded and closed 3-disc segments, and analyzed by a 1.5-Tesla MR scanner to measure MR parameters. RESULTS: Collagenase degradation of the NP significantly decreased the relaxation times, slightly decreased the magnetization transfer ratio, and slightly increased the apparent diffusion coefficient. Targeting the proteoglycan and/or hyaluronan integrity by trypsin and hyaluronidase did not significantly affect the MR parameters, except for an increase in the apparent diffusion coefficient of the disc after trypsin treatment. CONCLUSIONS: Our results demonstrate that changes in the structural integrity of matrix proteins can be assessed by quantitative MR.

Insulin-like growth factor (IGF)-binding protein-3 (IGFBP-3) is closely associated with the chondrocyte nucleus in human articular cartilage

OBJECTIVE: Insulin-like growth factor-I (IGF-I) is critically involved in the control of cartilage matrix metabolism. It is well known that IGF-binding protein-3 (IGFBP-3) is increased during osteoarthritis (OA), but its function(s) is not known. In other cells, IGFBP-3 can regulate IGF-I action in the extracellular environment and can also act independently inside the cell; this includes transcriptional gene control in the nucleus. These studies were undertaken to localize IGFBP-3 in human articular cartilage, particularly within cells. DESIGN: Cartilage was dissected from human femoral heads derived from arthroplasty for OA, and OA grade assessed by histology. Tissue slices were further characterized by extraction and assay of IGFBPs by IGF ligand blot (LB) and by enzyme-linked immunosorbent assay (ELISA). Immunohistochemistry (IHC) for IGF-I and IGFBP-3 was performed on cartilage from donors with mild, moderate and severe OA. Indirect fluorescence and immunogold-labeling IHC studies were included. RESULTS: LBs of chondrocyte lysates showed a strong signal for IGFBP-3. IHC of femoral cartilage sections at all OA stages showed IGF-I and IGFBP-3 matrix stain particularly in the top zones, and closely associated with most cells. A prominent perinuclear/nuclear IGFBP-3 signal was seen. Controls using non-immune sera or antigen-blocked antibody showed negative or strongly reduced stain. In frozen sections of human ankle cartilage, immunofluorescent IGFBP-3 stain co-localized with the nuclear 4',6-diamidino-2-phenyl indole (DAPI) stain in greater than 90% of the cells. Immunogold IHC of thin sections and transmission electron immunogold microscopy of ultra-thin sections showed distinct intra-nuclear staining. CONCLUSIONS: IGFBP-3 in human cartilage is located in the matrix and within chondrocytes in the cytoplasm and nuclei. This new finding indicates that the range of IGFBP-3 actions in articular cartilage is likely to include IGF-independent roles and opens the door to studies of its nuclear actions, including the possible regulation of hormone receptors or transcriptional complexes to control gene action.