Preliminary results confirmed that we could perform the MR imaging of the pig knee to detect as few as 100 simulated cells in the defect. According to our simulated labeled cell MRI and the results of the normal mini-pig knee MRI (figure 3.13), we chose the FSE PD FS (proton density fast spin echo sequence with fat saturation) sequence for a better anatomical description and 3D-GRE sequence as an iron sensitive sequence to visualize the presence of the iron labeled cells in the knee. Figure 3-13. MR sequence optimization. Different MR sequences performed to choose and optimize the best anatomical resolution for the mini-pig articular cartilage. FSE PD Fat Sat (A), 2D GRE (B), 3D FIESTA (C), 2D MERGE (D), SE MT (E), SE No MT (F). 3.4.7.2 MRI of the mini-pigs’ knee MRI was performed on normal knee before the surgery (no cartilage defect), one week after creating the defect (before cell injection), immediately after cell injection, weeks and weeks after autologous MSC injection (1 x 107). 70 Figure 3.14 showed the location of the bulk of MSC immediately after the injection. Figure 3-14. Post injection MR images. MR images of the mini-pig knee joint immediately after the 107 labeled MSCs injection in the knee capsule. FSE PD sequence (A) provided more anatomical data and 3D-GRE sequence (B) provided the labeled cell localization data. (Arrowheads show the distribution of the labeled cells in the knee joint) MSCs were distributed in different places of the knee (figure 3.14). The labeled cells migrated to different locations over time including cartilage defect, injection site, surgery scar, and supra-patellar fat and the cartilage defect site. It is likely that the healing process and the secretion of proinflammation factors associated with remodeling and repair of the injury provided the stimuli for MSC migration. MRI revealed that the injected cells were in defect site, para-patellar fat and injection site of the knee joint (figure 3.15 and 3.16). 71 Figure 3-15. MR imaging of the mini-pig's femoral condyle. MR imaging of the mini-pig’s femoral condyle before defect creation (normal), one week after making the defect (before injection), immediately after injection, two and six weeks after the cells injection. Upper panel (right knee) show the knee injected with HA plus labeled MSCs, however lower panel (left knee) shows the HA only injection (control knee). (Arrowheads indicate the defect site and arrows demonstrate the labeled stem cells) 72 Figure 3-16. MRI of the distribution of the labeled MSCs in the mini-pig knee joint. The labeled cells migrated to the para-patellar fat tissue (A) and surgical scar in the skin (B). 3.4.8 Postmortem analysis Histological grading scores of the repaired cartilage (calculated based on Wakitani scoring system at table 3.1) were compared between the scaffold only and scaffold plus MSCs groups and showed a significant difference in the cell morphology (hyaline cartilage production) (p=0.007) and matrix production staining (p=0.011). These data suggested that intra-articular injection of MSC could improve repair procedure by production of a higher quality cartilage (hyaline cartilage rather than fibrocartilage). However, histological grading analysis of the surface regularity (p=0.227), cartilage thickness (p=0.170) and integration to the adjacent cartilage (p=0.180) failed to reveal any significant improvement. The overall score confirmed that intraarticular injection of MSC enhanced cartilage repair (p=0.024) (figure 3.17). 73 . labeled cells in the knee joint) MSCs were distributed in different places of the knee (figure 3. 14) . The labeled cells migrated to different locations over time including cartilage defect, injection. difference in the cell morphology (hyaline cartilage production) (p=0.007) and matrix production staining (p=0.011). These data suggested that intra -articular injection of MSC could improve repair. revealed that the injected cells were in defect site, para-patellar fat and injection site of the knee joint (figure 3.15 and 3.16). 72! ! Figure 3-15. MR imaging of the mini-pig's femoral