In vivo high-resolution MRI (7T) of femoro-tibial cartilage changes in the rat anterior cruciate ligament transection model of osteoarthritis: a cross-sectional study.

OBJECTIVE: To assess OA-related changes in mean compartmental femorotibial cartilage thickness in rat knees by three-dimensional (3D) MRI (7T). METHODS: MRI was performed in vivo at 7T on OA and untouched contralateral knee joints. Gradient Echo Fast Imaging 3D MR images were acquired sequentially in surgically induced OA (D0) in 40 Wistar rats (anterior cruciate ligament transection). Mean femoral (trochlear, lateral and medial) and tibial (lateral and medial) cartilage thicknesses were quantified from a 2D MRI slide in weight-bearing areas and from a 3D MRI data set. At each time-point [Day (D)8, D14, D21, D40 and D60], eight animals (16 knees) were sacrificed for concomitant histomorphometry. RESULTS: As body weight dramatically increased throughout the experiment (+150%, baseline vs endpoint), all compartmental mean cartilage thicknesses noticeably decreased (D8, D14) and then remained relatively stable. Femoral compartments in OA knees were thinner at the end of the experiment than in contralateral age-matched knees. Conversely, lateral and medial tibial cartilages were thicker than controls. Histological correlation was significant only in untouched healthy cartilages (3D better than 2D). CONCLUSIONS: 3D MRI (7T) enables in vivo monitoring of compartmental changes in OA-related femorotibial rat cartilage thickness vs contralateral age-matched knees.

Phenotypic analysis of cell surface markers and gene expression of human mesenchymal stem cells and chondrocytes during monolayer expansion.

Both chondrocytes and mensenchymal stem cells (MSCs) are the most used cell sources for cartilage tissue engineering. However, monolayer expansion to obtain sufficient cells leads to a rapid chondrocyte dedifferentiation and a subsequent ancillary reduced ability of MSCs to differentiate into chondrocytes, thus limiting their application in cartilage repair. The aim of this study was to investigate the influence of the monolayer expansion on the immunophenotype and the gene expression profile of both cell types, and to find the appropriate compromise between monolayer expansion and the remaining chondrogenic characteristics. To this end, human chondrocytes, isolated enzymatically from femoral head slice, and human MSCs, derived from bone marrow, were maintained in monolayer culture up to passage 5. The respective expressions of cell surface markers (CD34, CD45, CD73, CD90, CD105, CD166) and several chondrogenic-related genes for each passage (P0-P5) of those cells were then analyzed using flow cytometry and quantitative real-time PCR, respectively. Flow cytometry analyses showed that, during the monolayer expansion, some qualitative and quantitative regulations occur for the expression of cell surface markers. A rapid increase in mRNA expression of type 1 collagen occurs whereas a significant decrease of type 2 collagen and Sox 9 was observed in chondrocytes through the successive passages. On the other hand, the expansion did not induced obvious change in MSCs gene expression. In conclusion, our results suggest that passage 1 might be the up-limit for chondrocytes in order to achieve their subsequent redifferentiation in 3D scaffold. Nevertheless, MSCs could be expanded in monolayer until passage 5 without loosing their undifferentiated phenotypes.

Quantitative dynamic contrast enhanced MRI of experimental synovitis in the rabbit knee: comparison of macromolecular blood pool agents vs. Gadolinium-DOTA.

The purpose of this study was to assess 2 Gd-based macromolecular intravascular contrast agents (P792, rapid clearance blood pool agent (rBPA) and P717, slow clearance blood pool agent (sBPA)) compared to Gd-DOTA (representative extracellular non specific agent) in MR imaging of knee rabbit experimental synovitis. Quantitative dynamic contrast enhanced MRI (qDCE-MRI) after intravascular injection of a low molecular weight contrast agent of 0.56 kDa (Gd-DOTA) and 2 high-molecular-weight contrast agents of 6.47 kDa (P792) and 52 kDa (P717) was performed in rabbits with carrageenan-induced synovitis of the right knee. P792 and P717 provided a progressive and persistent enhancement of arthritic synovial tissue while Gd-DOTA provided an early and rapidly declining enhancement with a concomitant diffusion in synovial fluid, thus limitating delineation of synovial pannus. P792 allowed acquisition of high-quality MR arthrograms, due to both a better diffusion in synovial pannus (vs. P717) and a concomitant restricted diffusion into the synovial fluid (vs. Gd-DOTA). In fact, experimental rabbit synovitis represent a specific entity that favors the T1 effect of high-molecular-weight agents, and especially rBPA P792, entrapped in synovial pannus, without diffusion in the synovial fluid. Due to this lack of arthrographic effect, P792 accumulation could be specifically sequentially analyzed by qDCE-MRI for detecting, characterizing and monitoring synovial vascular permeability changes during mono- or polysynovitis.

Age-related quantitative MRI changes in healthy cartilage: preliminary results.

OBJECTIVES: As the early form of OA is characterized by elevated water content in the cartilage tissue, the purpose of this study was to verify in vivo if age-related changes in patellar cartilage in healthy volunteers can be detected using quantitative MRI with T2 mapping and volume measurement MRI methods. DESIGN: Thirty healthy volunteers of various classes of age (18 to 65 years old) were enrolled in this study. MR images of the patellar cartilage were acquired at 1.5T. Patellar cartilage volume and T2 maps were determined. RESULTS: Despite non-significance, there was a trend in reducing cartilage volume with ageing (r: -0.25). In contrast global T2 slightly increased with ageing (r: 0.46). BMI (r: 0.51) and bone volume (r: 0.69) are well correlated to cartilage volume. CONCLUSION. Age-related physiologic changes in the water content of patellar cartilage can be detected using MRI. The proposed T2-mapping method, coupled with other non-invasive MR cartilage imaging techniques, could aid in the early diagnosis of OA.

Effect of proteoglycan depletion on T2 mapping in rat patellar cartilage.

PURPOSE: To evaluate experimentally the sensitivity of T2 mapping with magnetic resonance (MR) imaging at 8.5 T in depicting variations in proteoglycan content and concurrent extracellular matrix of rat patellar cartilage. MATERIALS AND METHODS: The study was performed in 36 immature (age, 5 weeks) and 36 mature (age, 10 weeks) Wistar rats. Maintenance and care of the rats were conducted in accordance with National Institutes of Health guidelines. Fifty-six rats underwent T2 mapping in 28 right patellae degraded with hyaluronidase for 1 and 6 hours and in 28 undegraded age-matched patellae that served as controls. After MR mapping, the rats were sacrificed, and the patellae were studied histologically to evaluate proteoglycan and collagen content and collagen network organization in cartilage. Biochemical analysis was performed in 88 patellae to quantify sulfated glycosaminoglycan and hydroxyproline content. Effects of age and/or degree of degradation were evaluated after rank transformation of continuous data by using rank analysis of variance (ANOVA). Associations between continuous variables were assessed with the Spearman rank correlation coefficient. RESULTS: Results of histologic analysis showed proteoglycan loss after hyaluronidase degradation without alteration of collagen network. No significant variation in hydroxyproline sulfate content was observed with depletion of proteoglycan. Proteoglycan losses of 19% and 13%, found after 1-hour degradation in immature and mature groups, respectively, were associated with significantly increased global T2 values (ANOVA, P < .001). Six-hour degradation resulted in more severe proteoglycan losses of 45% and 53% in immature and mature groups, respectively, inducing significant increases in global T2 values in immature and mature groups (ANOVA, P < .001). Variations in T2 values between superficial and deep cartilage zones were not affected by proteoglycan depletion. CONCLUSION: In rat patellar cartilage, T2 mapping permits detection of slight or severe proteoglycan depletion and concurrent changes of extracellular matrix when age-matched samples are compared.

Dose-response relationship for exercise on severity of experimental osteoarthritis in rats: a pilot study.

OBJECTIVE: To investigate the influence of a calibrated exercise on the progression of structural lesions in an experimental model of osteoarthritis (OA) in the rat, and to explore the effect of exercise on the level of chondrocyte caspase-dependent apoptosis and of Hsp70. METHODS: The OA model was induced by anterior cruciate ligament transection (ACLT). Rats were placed in 4 experimental groups: operated (ACLT) free moving rats, and 3 exercise groups (slight, moderate and intense) subjected to running training. Rats were killed 14 and 28 days after surgery. RESULTS: On D14 histological assessment demonstrated a beneficial influence of a slight and a moderate exercise vs control ACLT group. Hsp70 increased significantly in the moderate group vs controls. On D28, histological lesions strongly decreased in the slight and moderate exercise groups vs ACLT group, while an intense effort abolished this beneficial trend. Interestingly, the concomitant course of apoptotic events (caspase 3-positive cells) and the co-expression of Hsp70 in the various groups varied, when significant, in an inverse manner. In the intense group this overexpression was not noted, as a "burn out" appeared, thus leading to a loss of this protective effect. CONCLUSION: This study shows that a calibrated slight or moderate exercise exerts a beneficial influence on the severity of chondral lesions in ACLT rats. Conversely, a strong effort abolishes this chondroprotective effect. This effect could be related to a reduced level of chondrocyte apoptosis through anti-apoptotic capacities of stress-induced Hsp70 overexpression.