1. Changes in micro-architecture of subchondral bone in knee osteoarthritis Sam de Groen Supervisors: Prof Karen J Reynolds, Dr Egon Perilli Associates: Bryant C Roberts (Hons)

2. Introduction Osteoarthritis (OA): painful, disabling disease Estimated cost AU$1.2 billion dollars in 2004-2005 1 OA in the knee or hip is associated with altered walking gait patterns 2 1. Mehrotra C, Remington P, Naimi T, et al. (2005) 2. Astephen J.L, Deluzio K.J (2007) 2

3. Background OA: Bone and cartilage Pathogenesis of OA is complex Initially thought as a wear and tear disease of the cartilage 1 However, changes in subchondral bone structure were also found 2 Subchondral bone is an effective shock absorber 3 1. Radin E.L, Rose R.M (1986) 2. Lajeunesse D, Reboul P (2003) 3. Goldstein S.A (1983) 3 Jacquelyn Gilchrist (2003) http://www.interactive-biology.com/wp- content/uploads/2012/07/Osteoarthritis-of-Knee- Joint-1024x1024.jpg

4. Background Imaging of bone micro-architecture Traditionally subchondral bone micro-architecture was studied by histology (2D method) or micro-CT (3D method) on small biopsies from excised tibia Biopsies showed regional changes in subchondral bone and plate structure 4 Bobinac D, et al. (2003) 8mm Cartilage Sub. Plate Sub. Bone Milz, et al. (1994)

5. Aim To develop a method for 3D subregional analysis of the micro-architecture in the knee subchondral bone, on entire excised tibia plateau scanned with micro-CT Bigger project: To explore, on patients undergoing total knee arthroplasty, the relationships between the variations in subchondral bone micro-architecture measured in their excised tibial plateau, and the loading measured in vivo during walking prior to surgery. Project funded by Arthritis Australia (Grant in Aid, 2013 Perilli E, Thewlis D, Reynolds KJ) 5

6. Hypothesis The micro-architecture of the subchondral bone and thickness of subchondral plate, as measured by micro-CT, will differ among four anatomical sub-regions (anterior, posterior, medial, lateral) ACL PCL Posterior Lateral Medial Anterior 6

7. Method Tibial plateau harvested during TKA (Burnside Hospital)* Micro-CT imaging (Skyscan1076) at Adelaide Microscopy In the micro-CT cross-section images of entire plateau, subregions for analysis were created via software (sw CTAn, Skyscan), in two ways Bone volume and plate thickness were measured in these subregions (software CTAn, Skyscan) *Ethics Approval granted in July 2013 7

8. Subchondral Bone and Plate 8 2D Micro-CT cross-section images, 17 µ m/pixel

9. Selection of Regions of Interest 3 mm-thick ROIs (176 slides) in subchondral bone Irregular shaped ROIs 18mm diameter circular ROIs 9 Micro-CT cross-section images, 17 µ m/pixel 18mm 26mm

10. Results 10 26mm 18mm

11. Part II: Spatial Mapping Systematic mapping of the spatial variations in microarchitecture of both subchondral plate and subchondral bone, in 3D, without coring 11

12. Subchondral Bone and Plate 12 2D Micro-CT images, 17 µ m/pixel Subchondral Plate Subchondral Bone

13. Subchondral Bone and Plate 13 2D Micro-CT images, 17 µ m/pixel Subchondral Plate Subchondral Bone

14. 3D Analysis over VOI 5 x 5 x 21 mm 3 Separated subchondral bone and subchondral plate VOIs: 5 x 5 mm grid, 21 mm high 104 VOIs in total 14 3D Micro-CT images, 17 µ m/pixel

15. Subchondral bone and plate in 3D 15 3D Micro-CT images, 17 µ m/pixel

16. Subchondral bone and plate in 3D 16 3D Micro-CT images, 17 µ m/pixel

17. 17 Morphometric analysis over all VOIs (CTAn) Results into Excel spread sheet For graphic representation a MATLAB script was developed to generate ‘heat maps’ Analysis over VOI 5 x 5 x 21 mm 3

18. Subchondral bone (5x 5 x 21 mm 3 VOI) Bone Volume (mm 3 )Tb.Th (µm) 18 (µm) (mm 3 )

19. 19 Plate Th. (µm) Bone Volume (mm 3 ) (µm) (mm 3 ) Subchondral plate (5x 5 x 21 mm 3 VOI) Plate.Th (µm)

20. Summary 2 First micro-CT study that quantifies regional differences in entire human tibial plateau without coring the bone First method carrying out a 3D mapping of subchondral bone and subchondral plate microarchitecture Medial compartment showed highest bone volume in both subchondral bone as also in plate Medial compartment showed highest plate thickness 20

21. Future Directions Do the analysis on more samples (N=7 for pilot study) for looking at statistical significant differences among regions Select uniform VOI size for subchondral bone analysis –3x5x5mm ROIs across entire bone Automating processes –Edge detection for separation of subchondral bone and subchondral plate from micro-CT images –MATLAB script to generate results 21

22. Changes in micro-architecture of subchondral bone in knee osteoarthritis Sam de Groen Supervisors: Prof KJ Reynolds, Dr E Perilli, Assoc: BC Roberts (Hons) Thank you

23. Subchondral bone (5x 5 x 21 mm 3 VOI) Bone Volume (mm 3 ) Tb.Th (µm) 23 (µm) (mm 3 )

24. 24 Plate Th. (µm) Bone Volume (mm 3 ) (µm) (mm 3 ) Subchondral plate (5x 5 x 21 mm 3 VOI) Plate.Th (µm)

25. Future Directions -To look for statistical significant differences among 4 regions (or n x n): 1 ANOVA for repeated measurements 2 Post hoc analysis (Fishers, Scheffe or Bonferroni) -to look for correlations between joint moments and microarchitecture, in the 4 regions: correlation analysis. 25