B. J. Kim, D. -W. Kim, S. H. Kim, J. H. Cho, H. J. Lee, D. Y. Park, S

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Presentation transcript:

Establishment of a reliable and reproducible murine osteoarthritis model B.J. Kim, D.-W. Kim, S.H. Kim, J.H. Cho, H.J. Lee, D.Y. Park, S.R. Park, B.H. Choi, B.-H. Min Osteoarthritis and Cartilage Volume 21, Issue 12, Pages 2013-2020 (December 2013) DOI: 10.1016/j.joca.2013.09.012 Copyright © 2013 Osteoarthritis Research Society International Terms and Conditions

Fig. 1 (a) Schematic drawing of the cages of the confined cage group. The area was (75L × 40W mm2) for each mouse. (b) Photographic image of the confined cage, (c) treadmill was designed to encourage exercise accordingly. Osteoarthritis and Cartilage 2013 21, 2013-2020DOI: (10.1016/j.joca.2013.09.012) Copyright © 2013 Osteoarthritis Research Society International Terms and Conditions

Fig. 2 (a) C57BL/6 mice visualized by Ethovision® XT in the standard cage. The size of red square is 198L × 258W mm2 per mouse, (b) C57BL/6 mice visualized by Ethovision® XT in the confine cage. The size of red square is 38L × 73W mm2 per mouse, (c) daily distance of C57BL/6 movement in the confine cage (n = 4) compared to the standard cage (n = 4) by Ethovision® XT (mean and 95% CIs, P < 0.001), (d) weight changes of C57BL/6 in the confine cage (n = 4) and standard cage (n = 4) (mean and 95% CIs). Osteoarthritis and Cartilage 2013 21, 2013-2020DOI: (10.1016/j.joca.2013.09.012) Copyright © 2013 Osteoarthritis Research Society International Terms and Conditions

Fig. 3 Histological analysis of OA legions of C57BL/6 mice in the confined cage and standard cage. Histological sections of the medial femoral condyle (MFC) and the MTP were obtained and stained with Safranin-O at 2, 4, and 8 weeks after OA surgery. Mice with sham surgery were used as a control (n = 3, each week). (a) Representative images from each group with the highest and lowest OA scores were presented from samples of 8 weeks. Scale bar = 100 μm. (b) Average OA scores in the standard cages (n = 5, each week) and confined cages (n = 5, each week) are presented as mean and 95% CIs, P = 0.0157. Osteoarthritis and Cartilage 2013 21, 2013-2020DOI: (10.1016/j.joca.2013.09.012) Copyright © 2013 Osteoarthritis Research Society International Terms and Conditions

Fig. 4 Effect of increasing the amount of treadmill exercise on the induction of OA in mice maintained in confined cage. (a) C57Bl/6 mice in the confined cage were subjected to treadmill exercise of 0 (n = 5, each week), 400 (n = 5, each week), 800 (n = 5, each week), and 1200 m/day (n = 5, each week) at 2, 4, and 8 weeks after OA surgery. Histological sections of MTP at 2, 4, and 8 weeks after OA surgery were presented after Safranin-O staining. Scale bar = 100 μm. (b) In the graphs, OA scores of samples are presented as mean and 95% CIs. Osteoarthritis and Cartilage 2013 21, 2013-2020DOI: (10.1016/j.joca.2013.09.012) Copyright © 2013 Osteoarthritis Research Society International Terms and Conditions

Fig. 5 Effect of intra-articular HA injection on the OA lesions of C57/Bl6 mice depending on the movement condition. (a) Mice were subjected to OA surgery and maintained either in the standard cage with free movement (n = 36) or confined cage with an exercise of 400 m/day (n = 36). Saline or HA (10 mg/ml) was injected into the synovial cavity of OA knee joints at 0, 2, 4, and 6 weeks after OA surgery. Histological sections of cartilages were obtained at 2, 4, and 8 weeks and stained with Safranin-O. Representative images of the MFC and the MTP showing the highest and lowest OA scores were presented. Scale bar = 100 μm. (b) The graphs show OA scores of samples presented as mean and 95% CIs. Osteoarthritis and Cartilage 2013 21, 2013-2020DOI: (10.1016/j.joca.2013.09.012) Copyright © 2013 Osteoarthritis Research Society International Terms and Conditions

Supplementary Fig. 1 Safranin-O staining of all quadrants in the joint observed in the standard cage (sham, free moving) and confined cages (treadmill exercise of 0, 400, 800, and 1200 m/day) at 8 weeks after DMM surgery. (a) A consistent feature of the DMM model was loss of Safranin-O, fibrillation and cartilage loss on the medial tibial plateau (arrowhead) and medial femoral condyle. MFC = medial femoral condyle; LFC = lateral femoral condyle; MTP = medial tibial plateau; LTP = lateral tibial plateau; MM indicates a medially displaced medial meniscus. Scale bar = 200 μm. (b) Safranin-O staining of all four quadrants in the joints was scored for OA development at 8 weeks. Comparison was performed between the free moving and four confined cage groups, and between 0 m/day and three exercise groups in the confined cages. P values <0.05 are provided as below. Medial femoral condyle: 0 m/day vs 400, 800, 1200 m/day (P < 0.001, P < 0.001, P = 0.025), Medial tibial plateau: free moving vs 0 m/day (P = 0.015), 0 m/day vs 400, 800 m/day (P < 0.001, P = 0.006), Lateral femoral condyle: free moving vs 800, 1200 m/day (P = 0.001, P = 0.045), 0 m/day vs 400, 800, 1200 m/day (P = 0.002, P < 0.001, P = 0.011), Lateral tibial plateau: free moving vs 400, 800, 1200 m/day (P = 0.004, P = 0.012, P = 0.014), 0 m/day vs 400, 800, 1200 m/day (P = 0.004, P = 0.012, P = 0.014). Statistical analysis was performed using the one-way analysis of variance (ANOVA) with the Tukey's post hoc test using SPSS 12.0.1 (SPSS, Inc.). (c) Osteophyte formation was observed in all groups and determined by an osteophyte formation score system34. Scoring was done by two independent researchers blinded to any other information and results were averaged. Statistical analysis was performed using the one-way analysis of variance (ANOVA) with the Tukey's post hoc test using SPSS 12.0.1 (SPSS, Inc.). Comparison was performed between the free moving and four confined cage groups, and between 0 m/day and three exercise groups in the confined cages. P values are presented in the figure. Osteoarthritis and Cartilage 2013 21, 2013-2020DOI: (10.1016/j.joca.2013.09.012) Copyright © 2013 Osteoarthritis Research Society International Terms and Conditions

Supplementary Fig. 2 Histological analysis of OA lesions of no exercise (free moving) and exercise (400 m/day) groups in the standard cage. (a) Histological sections of medial tibial plateau at 2, 4 weeks after OA surgery and treadmill exercise (400 m/day) after Safranin-O staining. Scale bar = 100 μm. In the graphs, OA scores of samples are presented as mean and 95% CIs. (b) Mean and SD, their P values, CV and 95% confidence intervals (CIs) of histologic scores in the free moving (no exercise, n = 5) and exercise (400 m/day, n = 4) groups raised in the standard cages. The diversity of variance was determined by an F-test. The statistical significance of differences between means was verified in two sample t tests. Osteoarthritis and Cartilage 2013 21, 2013-2020DOI: (10.1016/j.joca.2013.09.012) Copyright © 2013 Osteoarthritis Research Society International Terms and Conditions