Fig. 3

Excessive tissue and ectopic chondrocyte formation are key features of advanced disease. a A 12-week mouse IVD demonstrates both early (i) and intermediate (ii) stages of excessive tissue formation using H&E stain. Scale bar = 300 μm. Inflammatory infiltrate (yellow arrow) surrounds the affected joint, while fibroblast-like cells (arrowheads) and chondrocyte-like cells (arrows) are located adjacently to the residual disc (asterisks). Scale bar = 200 μm in (i) and (ii). CEP cartilaginous end plate. b Excessive tissue at the periphery of the joint is greatly increased in more advanced disease (arrows). Representative image from a 24-week mouse. Scale bar = 400 μm. c In addition to excess tissue (black arrow), columnar chondrocytes expand ectopically along the cortical bone between affected joints (yellow arrowheads). Representative image from a 24-week mouse. Scale bar = 300 μm. d Representative images taken from 24-week mice show an unaffected IVD and affected joints with excessive or ectopic chondrocyte formation. Cartilaginous tissue (arrows) is positive for toluidine blue (proteoglycan content) and type II collagen. Type X collagen stains for hypertrophic chondrocytes while type I collagen delineates mature bone. Scale bar = 100 μm. e Spinal gene expression profiles of cartilage markers Col2a1, Comp and Sox9 were analysed by qPCR reaction and normalized to β-actin. Black bars represent naïve and open bars represent PGISp mice. Expression levels are presented as mean ± standard deviation. ***P < 0.0001, **P < 0.001, *P < 0.01 compared with naïve at the same time point, Mann-Whitney test. Col2a1 type II collagen, α 1, COMP cartilage oligomeric matrix protein, Sox9 sex determining region Y- box 9