NMR microscopic studies of articular cartilage at 7.1 T are presented. Using a special experimental design, T2-weighted spin-echo images of cartilage-bone plugs were taken under variable angles with respect to the static magnetic field B0 to visualize the angular-dependent representation of internal matrix structures mediated by the collagen network arrangement. To quantify the observed orientational effect in the MR images, exact measurements of the transverse relaxation time T2 were taken using the CPMG sequence. The NMR experiments show the strong influence of the cartilage orientation with respect to the static magnetic field on the inhomogeneous appearance of the articular cartilage in the MR image. Additionally performed polarization light microscopic investigations demonstrate the direct relation between the oriented collagenous structures and the anisotropic regions observed in the MR images. A simple cartilage matrix model derived from the experimental findings is proposed, and consequences for the clinical assessment of the articular joint are discussed.
In T 2 -weighted MRI images joint cartilage can appear laminated. The multilaminar appearance is visualized as zones of different intensity. This appearance is based on the dipolar interaction of water molecules within cartilage zones of different collageneous network structures. Therefore, the MR visualization of zones of anisotropic arrangement of the collagen fibers depends upon their orientation to the static magnetic field (magic-angle effect). The aim of this article is to demonstrate the potential of high-resolution MRI for characterizing cartilage network structuring and biomechanical properties. Information equivalent to that from polarization light microscopy can be derived noninvasively. Based on NMR microscopic (mMRI) data, potential new possibilities of MRI for quantitative assessment of collagen structuring and intracartilagenous load distribution are presented. These methods use MR intensity angle dependence and load influence on cartilage visualization. Alternatively to the determination of mechanical parameters from cartilage deformation, it is demonstrated that stress distribution and biomechanical properties can be derived in principle from the local intensity variation of anisotropic fiber orientation zones. The limitations with respect to a clinical application of the proposed methods are discussed.
In this paper, investigations are described on the influence of osmotic pressures and of varying cation concentrations on water relaxation times in cartilage (pig articular cartilage and bovine nasal cartilage). Both water content and relaxation times decrease strongly with increasing osmotic pressure. This relaxation behavior can be explained in terms of a fast chemical exchange between unbound and bound water. Na+ does not influence water content or relaxation times, whereas Ca2+ causes a small reduction in these parameters.
Transverse nuclear magnetic relaxation and self-diffusion of water were measured in hydrated collagen II. Self-diffusion measurements were conducted by pulsed field gradient NMR (PFG NMR) and weighting of the different species in the signal by variable T2 relaxation in the experiment. Two fractions of water protons were detected, one with a short T2 value but high diffusivity and one with a long T2 value and low, completely restricted diffusion. The distance of the diffusion barriers was determined to be 2.3 microns. Possible reasons for the restriction in the movement of the water molecules in comparison with structural models of collagen II are discussed.
The action of sodium hypochlorite on N-acetylglucosamine, N-acetylgalactosamine, chondroitinsulfate and hyaluronic acid was studied by 1H-nuclear magnetic resonance (1H-NMR) in order to model some aspects of degradation processes caused by neutrophils on carbohydrate polymers of cartilage in rheumatoid arthritis. N-Acetyl side groups of carbohydrate monomers and chondroitinsulfate yield a resonance at 2.01-2.04 ppm in proton NMR-spectra. This resonance is observed in hyaluronic acid solutions only after a prolonged incubation to yield shorter polymeric chains. Sodium hypochlorite causes a continuous decrease of the line for N-acetyl groups. Two new resonances appear in the 1H-NMR spectra. An intermediate product, assumed as a chlorinated product of N-acetyl side chains, shows a chemical shift of about 2.35 ppm. This intermediate is hydrolyzed to a carbohydrate ring and acetate (1.90 ppm). Sodium hypochlorite acts in all systems investigated mainly on N-acetyl groups. Only small effects on the carbohydrate ring were found under our experimental conditions.
Saline solutions of manganese ions (Mn2+) were used as articular contrast agents in magnetic resonance microscopy (9.4 T) of cartilage of chicken femoral condyles and pig temporomandibular joints. The diffusion of Mn2+ from the articular surface into the cartilage matrix led to a strong contrast enhancement in the cartilage. The combination of the high spatial resolution and the contrast enhancement allowed the visualization of fine structures (tissue types) in the cartilage, which correlate with the tissue zones in histological sections stained with cationic dyes. We assume that the electrostatic interactions between the negatively charged groups of the proteoglycans and Mn2+ are most important for the mechanism of contrast enhancement. Hence, the different signal intensities of the various zones of cartilage indicate differences in density of proteoglycans. The intraarticular injection of the cationic contrast agent could improve the possibility of an early diagnosis of cartilage dysfunction and degeneration.
A combination of solid-state NMR spectroscopy and MRI was used to evaluate the formation of extracellular matrix in poly(D,L-lactide-co-glycolide) (PLGA) bone implants. Porous PLGA scaffolds were implanted into rat tibiae and analysed after 2, 4 or 8 weeks. MRI clearly delineated the implants within the cancellous bone. Differences in the trabecular structure of the implanted material and native bone were demonstrated. In addition, implants were analyzed by solid-state NMR spectroscopy under magic angle spinning. (13)C NMR spectra showed the unambiguous signature of collagen formed in the scaffolds, but also the characteristic signals of the PLGA matrix, indicating that resorption was not complete after 8 weeks. Furthermore, (31)P NMR spectroscopy detected the inorganic component of the matrix, which is composed of bioapatite. (31)P NMR spectra were quantified and this analysis revealed that the amount of inorganic extracellular matrix formed de novo was significantly lower than in native bone. This demonstrates that solid-state NMR spectroscopy, in particular in combination with MRI, can provide useful information on the composition and structure of the extracellular matrix, and serve as a tool to evaluate the quality of tissue engineering strategies.
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