Evidence for articular cartilage regeneration in MRL/MpJ mice J. Fitzgerald, Ph.D., C. Rich, M.S., D. Burkhardt, B.S., J. Allen, Ph.D., A.S. Herzka, M.D., C.B. Little, B.V.M.S., Ph.D.  Osteoarthritis and Cartilage  Volume 16, Issue 11, Pages 1319-1326 (November 2008) DOI: 10.1016/j.joca.2008.03.014 Copyright © 2008 Osteoarthritis Research Society International Terms and Conditions

Fig. 1 Anatomical location of full thickness lesion (circle) and partial thickness laceration (line) in the trochlear groove. The joint capsule of the right hindlimb was opened and the patella subluxated to the left to expose the trochlear groove. The full thickness defect was made on the proximal part of the trochlear and the partial thickness lesion distal to this. Osteoarthritis and Cartilage 2008 16, 1319-1326DOI: (10.1016/j.joca.2008.03.014) Copyright © 2008 Osteoarthritis Research Society International Terms and Conditions

Fig. 2 Analysis of articular cartilage lesions at 6 weeks timepoint. (A) Representative sagittal sections of full thickness lesions from four C57Bl/6 (left) and MRL/MpJ (right) mice stained with toluidine blue for proteoglycan. The MRL mice have significant proteoglycan deposition compared to the C57Bl/6 control strain. (B) Mean and standard deviation of total histological score for full thickness cartilage lesions in MRL/MpJ and C57Bl/6 male and female experimental groups. The worst area of the central part of the lesion that could be found in serial sections of each mouse was scored. Female MRL mice scored significantly better than MRL males (P=0.04, Mann–Whitney U test) and both male and female C57 mice. The scoring matrix used for assessment of the full thickness lesions is shown below Fig. 2(B). Osteoarthritis and Cartilage 2008 16, 1319-1326DOI: (10.1016/j.joca.2008.03.014) Copyright © 2008 Osteoarthritis Research Society International Terms and Conditions

Fig. 3 Analysis of full thickness cartilage lesions 12 weeks following surgery. (A) Representative sagittal sections of full thickness lesions from three C57Bl/6 (left) and MRL/MpJ (right) mice stained with toluidine blue. The MRL mice have extensive proteoglycan deposition compared with the C57Bl/6 control strain, which in the majority of mice have no metachromatic staining. (B) Representative sagittal sections of full thickness lesions from MRL/MpJ (panels b, e, i) and C57Bl/6 mice (panels c, f, j). Non-operated C57Bl/6 mice (panels a, d, h) and sections stained with secondary antibody only serve as controls (panels g, k). Sections were stained for the presence of proteoglycans (toluidine blue), collagen II and collagen VI. The MRL/MpJ mice have extensive proteoglycan, collagen II and collagen VI deposition in the lesion compared with the C57/Bl6 control strain. Panels a–g, k are 10× and panels h, i, j are 20×. Panel l is a higher power image (40×) of proteoglycan stained MRL/MpJ lesion. Note the abundance of chondrocytes in the lesion. (C) Mean and standard deviation of total histological score for full thickness cartilage lesions. (D) Median, maximum score, minimum score and 25th and 75th percentile of total histology data for each experimental group. Although there is variation within experimental groups the MRL male groups clearly heals better overall. (E) Individual parameter scores for full thickness cartilage lesion. MRL males are significantly better healers in four out of five parameters. Integration with native cartilage at wounded site was similar in all experimental groups. The P-values for each individual parameter for MRL males vs MRL females and C57 males are shown in the table below the graph. Osteoarthritis and Cartilage 2008 16, 1319-1326DOI: (10.1016/j.joca.2008.03.014) Copyright © 2008 Osteoarthritis Research Society International Terms and Conditions

Fig. 4 (A) Representative sagittal section of partial thickness cartilage lesions at 12 weeks after surgery stained with toluidine blue. (B) Mean and standard error of the mean (s.e.m.) of total histological score for partial thickness cartilage lesions 6 weeks and 12 weeks after surgery. There was poor healing of cartilage laceration in both strains and no change in healing between sexes or with post-operative time. Osteoarthritis and Cartilage 2008 16, 1319-1326DOI: (10.1016/j.joca.2008.03.014) Copyright © 2008 Osteoarthritis Research Society International Terms and Conditions

Fig. 5 Analysis of ear hole closure in MRL/MpJ and C57bl/6 mice 12 weeks post-surgery. (A) Representative examples of ear hole closure from three C57Bl/6 (above) and MRL/MpJ (below) mice. (B) Mean and standard deviation of ear hole closure of all experimental groups of operated mice (in mm). When compared by strain C57 ear holes did not close appreciably while MRL ear holes were significantly smaller (P=0.0002). Osteoarthritis and Cartilage 2008 16, 1319-1326DOI: (10.1016/j.joca.2008.03.014) Copyright © 2008 Osteoarthritis Research Society International Terms and Conditions