Comparison of mouse and human ankles and establishment of mouse ankle osteoarthritis models by surgically-induced instability S.H. Chang, T. Yasui, S.

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Comparison of mouse and human ankles and establishment of mouse ankle osteoarthritis models by surgically-induced instability S.H. Chang, T. Yasui, S.

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Comparison of mouse and human ankles and establishment of mouse ankle osteoarthritis models by surgically-induced instability S.H. Chang, T. Yasui, S. Taketomi, T. Matsumoto, J.R. Kim-Kaneyama, T. Omiya, Y. Hosaka, H. Inui, Y. Omata, R. Yamagami, D. Mori, F. Yano, U. Chung, S. Tanaka, T. Saito Osteoarthritis and Cartilage Volume 24, Issue 4, Pages 688-697 (April 2016) DOI: 10.1016/j.joca.2015.11.008 Copyright © 2015 Osteoarthritis Research Society International Terms and Conditions

Fig. 1 Comparison of mouse knee and ankle joints. (A) Anatomy of mouse ankle joint. Tibialis posterior tendon (TP, black arrows) and deltoid ligament (DL, green arrows) are shown in the medial side (left panel). Anterior talofibular ligament (ATFL, blue arrows), calcaneofibular ligament (CFL, green arrows), and peroneus longus/brevis tendons (PL/PB, black arrows) are shown in the lateral side (right panel). Scale bars, 1 mm. (B) Safranin O stainings of mouse ankle and knee joints. Inset boxes indicate the regions shown in the enlarged images below. T: tibiotalar joint, S: subtalar joint, M: medial compartment of knee joint, L: lateral compartment of knee joint. Scale bars, 200 μm. (C) Enlarged Safranin O stainings of articular cartilage of tibiotalar and knee joints in mouse and human. Scale bars, 40 μm for mouse, and 300 μm for human. (D) Thickness of articular cartilage and cancellous bone volume (BV/TV) of subchondral bone of the talus in the tibiotalar joint and the tibia in the medial knee joint of mouse and human. Thickness of articular cartilage was measured at 15 points in three sections. BV/TV was measured at six areas in three sections. Data are expressed as scatter plots, and as means (bars) ± 95% confidence interval (CI) (error bars). Osteoarthritis and Cartilage 2016 24, 688-697DOI: (10.1016/j.joca.2015.11.008) Copyright © 2015 Osteoarthritis Research Society International Terms and Conditions

Fig. 2 Articular cartilage degradation of mouse ankle and knee joints with aging. (A). Safranin O stainings of ankle and knee joints of 25-month-old mice. Inset boxes indicate the regions shown in the enlarged images below. T: tibiotalar joint, S: subtalar joint, M: medial compartment of knee joint, L: lateral compartment of knee joint. Scale bars, 200 μm and 50 μm for low and high magnification images, respectively. (B) OARSI score of two joints in the ankle and both compartments in the knee. Data are expressed as scatter plots for five mice. Data are expressed as means (bars) ± 95% confidence interval (CI) (error bars). Osteoarthritis and Cartilage 2016 24, 688-697DOI: (10.1016/j.joca.2015.11.008) Copyright © 2015 Osteoarthritis Research Society International Terms and Conditions

Fig. 3 Medial ankle OA model. (A) Resection of TP (left panel) and DL (right panel). Red double lines indicate TP and DL, respectively. (B) Safranin O staining of the representative ankle joint in the medial OA model. Inset boxes indicate the regions shown in the enlarged images. T: tibiotalar joint, S: subtalar joint. Scale bars, 200 μm and 50 μm for low and high magnification images, respectively. (C) OARSI score of tibiotalar and subtalar joints in operated and sham limbs. Data are expressed as scatter plots for 12 mice. Data are expressed as means (bars) ± 95% CI (error bars). (D) Micro CT images of the medial side of ankle joints; before (left panel) and 8 weeks after surgery (right panel). Red arrow indicates a medial malleolar osteophyte. Osteoarthritis and Cartilage 2016 24, 688-697DOI: (10.1016/j.joca.2015.11.008) Copyright © 2015 Osteoarthritis Research Society International Terms and Conditions

Fig. 4 Lateral ankle OA model. (A) Resection of ATFL (left) and CFL (right). Red double lines indicate ATFL and CFL, respectively. (B) Safranin O staining of the representative ankle joint in the lateral OA model. Inset boxes indicate the regions shown in the enlarged images. T: tibiotalar joint, S: subtalar joint. Scale bars, 200 μm and 50 μm for low and high magnification images, respectively. (C) OARSI score of tibiotalar and subtalar joints in operated and sham limbs. Data are expressed as scatter plots for 12 mice. Data are expressed as means (bars) ± 95% CI (error bars). (D) Micro CT images of the lateral side of ankle joints; before (left panel) and 8 weeks after surgery (right panel). Red arrows indicate lateral malleolar osteophytes. Osteoarthritis and Cartilage 2016 24, 688-697DOI: (10.1016/j.joca.2015.11.008) Copyright © 2015 Osteoarthritis Research Society International Terms and Conditions

Fig. 5 Bilateral ankle OA model. (A) Resection of PL/PB. Bilateral model was prepared by resection of TP, DL, ATFL, CFL, and PL/PB. (B) Safranin O staining of the representative ankle joint in the bilateral OA model. Inset boxes indicate the regions shown in the enlarged images. T: tibiotalar joint, S: subtalar joint. Scale bars, 200 μm and 50 μm for low and high magnification images, respectively. (C) OARSI score of tibiotalar and subtalar joints in operated and sham limbs. Data are expressed as scatter plots for 12 mice. Data are expressed as means (bars) ± 95% CI (error bars). (D) Micro CT images of the medial (left panel) and the lateral side (right panel) of the mouse ankle joint 8 weeks after surgery. Red arrows indicate medial and lateral malleolar osteophytes, respectively. Osteoarthritis and Cartilage 2016 24, 688-697DOI: (10.1016/j.joca.2015.11.008) Copyright © 2015 Osteoarthritis Research Society International Terms and Conditions

Fig. 6 Sequential progression of the medial ankle OA model. Safranin O stainings, TUNEL stainings, and immunofluorescence of Adamts5 and Mmp13 in sequential sections of the medial model. Inset box indicates the regions shown in the enlarged images below. Scale bars, 200 μm and 100 μm for low and high magnification images, respectively. Osteoarthritis and Cartilage 2016 24, 688-697DOI: (10.1016/j.joca.2015.11.008) Copyright © 2015 Osteoarthritis Research Society International Terms and Conditions