Differential recovery of glycosaminoglycan after IL-1-induced degradation of bovine articular cartilage depends on degree of degradation

Abstract

In the present study we examined cartilage matrix repair following IL-1-induced matrix depletion. Previous data indicated that, in some cases, chondrocytes can synthesize macromolecules to establish a functional extracellular matrix in response to a matrix-damaging insult or when placed in a three-dimensional environment with inadequate matrix. However, the conditions under which such 'repair' can occur are not entirely clear. Prior studies have shown that chondrocytes in trypsin-depleted young bovine articular cartilage can replenish tissue glycosaminoglycan (GAG) and that the rate of replenishment is relatively uniform throughout the tissue, suggesting that all chondrocytes have similar capacity for repair. In the present study we used the characteristic heterogeneous distribution of matrix depletion in response to IL-1 exposure in order to investigate whether the severity of depletion influenced the rate of GAG replenishment. We used the delayed Gadolinium-Enhanced Magnetic Resonance Imaging of Cartilage (dGEMRIC) method to monitor the spatial and temporal evolution of tissue GAG concentration ([GAG]). For both mild (n=4) and moderate (n=10) IL-1-induced GAG depletion, we observed partial recovery of GAG (80% and 50% of baseline values, respectively) over a 3-week recovery period. During the first 2 weeks of recovery, [GAG] increased homogeneously at 10-15 mg/ml per week. However, during the third week the regions most severely depleted following IL-1 exposure showed negligible [GAG] accumulation, whereas those regions affected the least by IL-1 demonstrated the greatest accumulation. This finding could suggest that the most severely degraded regions do not recover fully, possibly because of more severe collagen damage; this possibility requires further examination.

Figure 1

Representative glycosaminoglycan (GAG) map series derived from T1 maps measured on successive weeks. Initial GAG concentration ([GAG]) was substantially different for the two animals (one animal/series), and therefore each series is shown on its own color-scale. (A and C) Control [GAG] is stable (COV varied by ± 2–12%) throughout the recovery period for both series. (B and D) At the beginning of the recovery period (week 0), [GAG] for IL-1-exposed samples is lower than the initial [GAG] and steadily increases over the 3-week recovery period.

Figure 2

Mean glycosaminoglycan concentration ([GAG]) in IL-1-exposed samples measured at weekly intervals using delayed gadolinium enhanced magnetic resonance imaging of cartilage (dGEMRIC). The mean [GAG] increase with recovery time for samples subjected to (A) mild (3 and 6 days of 10 ng/ml IL-1, n = 4; P = 0.06) or (B) moderate (6 and 9 days of 20 ng/ml IL-1, n = 10; P = 0.0001) degradation, and then permitted to recover for 3 weeks in culture. The mean [GAG] for a given sample at a given time point was computed as the mean of [GAG] measured across all pixels of the image; error bars are ± SD between sample means. Shaded regions represent initial [GAG]; 92 ± 11 mg/ml for 'mild' series and 57 ± 6 mg/ml for 'moderate' series.

Figure 3

(A) Example of regional analysis scheme. For samples in the 'moderate' group, glycosaminoglycan concentration ([GAG]) maps measured after 3 weeks of recovery were segmented into 'low', 'medium', and 'high' regions, as specified in the Methods section under Image processing (so that the set of pixels defined as 'low' represented the regions of tissue that recovered the least during the 3-week recovery period and the set defined as 'high' represented tissue that recovered the most). The mean [GAG] of these three regions were followed in time. At each time point, segmented images were analyzed separately to assess whether GAG contents and recovery rates were comparable. (B) Weekly mean [GAG] ± SD of regions defined as 'low' (red), 'medium' (yellow), or 'high' (green), according to the process illustrated in panel A. (C) Weekly changes in mean [GAG] ± SD are shown for each region. Rate of [GAG] recovery is independent of absolute [GAG] for the first 2 weeks of culture after IL-1 exposure. 'Low', 'medium', and 'high' GAG regions recover at statistically different rates during the third week following IL-1 exposure (* P < 0.0001). All mean [GAG] values and recovery rates are derived from a total of 10 samples.