Many biomechanical and chemical properties of cartilage are dependent on the fixed charge density (FCD) of the extracellular matrix. In this study, nuclear magnetic resonance (NMR) spectroscopy was investigated as a nondestructive technique for determining FCD in cartilage. Sodium content was measured by NMR in cartilage explants and was compared with sodium content measured by inductively coupled plasma emission spectroscopy (ICP) in order to verify the total NMR visibility of sodium in cartilage. The ratio of NMR to ICP results was 1.02 +/- 0.04 (calf, mean +/- SD, n = 7) and 1.04 +/- 0.11 (adult bovine, n = 8). Sodium concentration as measured by NMR was then used with ideal Donnan theory to compute estimates of FCD. For calf articular cartilage (AC) near physiological conditions, calculated FCD was -0.28 +/- 0.03 M (n = 10). NMR measurements were then made for individual cartilage specimens sequentially equilibrated in baths of differing salt composition, pH, or ionic strength. For calf and adult AC, calculated FCD decreased dramatically between pH 3 and 2, with adult specimens becoming positively charged but calf tissue retaining a net negative charge. For calf AC equilibrated in 0.3-0.015 M NaCl, calculated FCD was observed to decrease slightly with decreasing bath ionic strength. For epiphyseal cartilage, FCD varied with the position of origin of the explant within the joint, ranging from -0.19 to -0.35 M in a manner that correlated with tissue glycosaminoglycan content. Preliminary NMR imaging experiments demonstrated similar variations of sodium concentration in intact ulnar epiphyseal cartilage. Collectively, these results demonstrate the ability of NMR to nondestructively follow FCD in cartilage. The technique is applicable to dynamic studies as well as to both in vitro and in vivo studies on living tissue.

The goal of this work was to investigate magnetization transfer (MT) in cartilage by measuring water proton signals Ms/Mo, as an indicator of MT, in (i) single-component systems of the tissue's constituent macromolecules and (ii) intact cartilage under control conditions and after two pathomimetic interventions. Ms/Mo was quantified with a 12-microT saturation pulse applied 6 kHz off resonance. Both glycosaminoglycans (GAG) and collagen exhibited concentration dependent effects on Ms/Mo, being approximately linear for GAG solutions (Ms/Mo = -0.0137[% GAG] + 1.02) and exponential for collagen suspensions (Ms/Mo = 0.80 x exp[-(%collagen)/6.66] + 0.20); the direct saturation of water could not account for the measured Ms/Mo. Although the effect of collagen on Ms/Mo is much stronger than for a corresponding concentration of GAG, Ms/Mo is not very sensitive to changes in collagen concentration in the physiological range. Tissue degradation with 25 mg/ml trypsin led to an increase in Ms/Mo from the baseline value of 0.2 (final/initial values = 1.15 +/- 0.13, n = 11, P < 0.001). In contrast, a 10-day treatment of cartilage with 100 ng/ml of interleukin-1 beta (IL-1 beta) caused a 19% decrease in Ms/Mo (final/initial values = 0.81 +/- 0.08, n = 3, P = 0.085). The changes in hydration and macromolecular content for the two treatments were comparable, suggesting that Ms/Mo is sensitive to macromolecular structure as well as concentration. In conclusion, whereas the baseline Ms/Mo value in cartilage may be primarily due to the tissue collagen concentration, changes in Ms/Mo may be due to physiological or pathophysiological changes in GAG concentration and tissue structure, and the measured Ms/Mo may differentiate between various pathomimetic degradative procedures.

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