Study of subchondral bone adaptations in a rodent surgical model of OA using in vivo micro-computed tomography.

OBJECTIVE: To non-invasively investigate the changes to epiphyseal bone occurring in a longitudinal pre-clinical model of osteoarthritis (OA) using in vivo micro-computed tomography (micro-CT). DESIGN: In vivo micro-CT images were acquired using a bench-top micro-CT scanner, which produces three-dimensional data with isotropic voxel spacing of 0.046 mm. Male rodents were scanned prior to surgical destabilization, consisting of anterior cruciate ligament transection and partial medial menisectomy (ACLX). Subsequent scans were performed every 4 weeks post-ACLX, for up to 5 months. Volumetric bone mineral density (vBMD) was measured in specific, anatomically segmented regions within each image. The ACLX rodent data were compared with the contralateral non-operated hind limb of the same animal, as well as a sham-operated group (SHAM) of animals, for each time point. End-point histology compared changes to cartilage and bone between the ACLX and control animals. RESULTS: The micro-CT protocol produced sufficient spatial resolution and signal-to-noise ratio (SNR=19) to quantify subchondral bone pathology, with an acceptable entrance exposure to radiation (0.36 Gy). Significantly lower vBMD was measured in the ACLX group, vs SHAM rodents, at 1, 4, and 5 months post-surgery (P<0.05). Qualitative observations of ACLX joints revealed significant loss of cartilage, subchondral bone cysts, and calcification of tendon similar to changes found in humans. CONCLUSIONS: This study demonstrates in vivo micro-CT as an effective method for investigating the development of rodent knee OA longitudinally. This method can be applied, in future pre-clinical trials, to non-destructively monitor the efficacy of pharmacological interventions.

Global analyses of gene expression in early experimental osteoarthritis.

OBJECTIVE: To analyze genome-wide changes in chondrocyte gene expression in a surgically induced model of early osteoarthritis (OA) in rats, to assess the similarity of this model to human OA, and to identify genes and mechanisms leading to OA pathogenesis. METHODS: OA was surgically induced in 5 rats by anterior cruciate ligament transection and partial medial meniscectomy. Sham surgery was performed in 5 additional animals, which were used as controls. Both groups underwent 4 weeks of forced mobilization, 3 times per week. RNA was extracted directly from articular chondrocytes in the OA (operated), contralateral, and sham-operated knees. Affymetrix GeneChip expression arrays were used to assess genome-wide changes in gene expression. Expression patterns of selected dysregulated genes, including Col2a1, Mmp13, Adamts5, Ctsc, Ptges, and Cxcr4, were validated by real-time polymerase chain reaction, immunofluorescence, or immunohistochemistry 2, 4, and 8 weeks after surgery. RESULTS: After normalization, comparison of OA and sham-operated samples showed 1,619 differentially expressed probe sets with changes in their levels of expression > or = 1.5-fold, 722 with changes > or = 2-fold, 135 with changes > or = 4-fold, and 20 with changes of 8-fold. Dysregulated genes known to be involved in human OA included Mmp13, Adamts5, and Ptgs2, among others. Several dysregulated genes (e.g., Reln, Phex, and Ltbp2) had been identified in our earlier microarray study of hypertrophic chondrocyte differentiation. Other genes involved in cytokine and chemokine signaling, including Cxcr4 and Ccl2, were identified. Changes in gene expression were also observed in the contralateral knee, validating the sham operation as the appropriate control. CONCLUSION: Our results demonstrate that the animal model mimics gene expression changes seen in human OA, supporting the relevance of newly identified genes and pathways to early human OA. We propose new avenues for OA pathogenesis research and potential targets for novel OA treatments, including cathepsins and cytokine, chemokine, and growth factor signaling pathways, in addition to factors controlling the progression of chondrocyte differentiation.