Figure 1. Loss of DOT1L disrupts chondrocyte homeostasis and triggers osteoarthritis.

(a) Immunohistochemistry showing reduced methylated H3K79 levels (H3K79me2) that reflect loss of DOT1L activity in damaged areas from osteoarthritic patients (OA) as compared to their corresponding preserved areas and to cartilage from non-OA patients. Images are representative of images from four different patients. Scale bar, 400 μm. (b) Heat maps of differential mRNA expression determined by quantitative PCR in chondrocytes treated with DOT1L inhibitor EPZ-5676 (EPZ) or vehicle (V) from passage 0 (P0) until P2, and from preserved versus damaged areas in OA cartilage. The colour code represents the mean expression level of six and four independent patient samples respectively. (c) Immunoblot analysis showing decreased methylated H3K79 levels in mouse articular chondrocytes after intra-articular injection of EPZ into C57Bl/6 wild-type mouse knees. The image is representative of one experiment with protein extracts pooled from two or three mice per condition. Unprocessed original scans of blots are shown in Supplementary Fig. 10. (d,e) C57/Bl6 wild-type mouse knees were injected with EPZ (5 mg kg^–1) or vehicle and killed after 2 or 4 weeks. Knees were sectioned and stained with Hematoxylin-Safranin O (d). Scale bar, 200 μm. Cartilage damage was scored (see Methods section) and is shown in (e). One experiment was performed with n=10 and 5. Representative images from the 4 week evaluation are shown. *P<0.05 (two-tailed t-test). Error bars indicate mean±s.e.m.