The mobility of chondroitin sulfate in articular and artificial cartilage characterized by ¹³C magic‐angle spinning NMR spectroscopy

Abstract

We have studied the molecular dynamics of one of the major macromolecules in articular cartilage, chondroitin sulfate. Applying (13)C high-resolution magic-angle spinning NMR techniques, the NMR signals of all rigid macromolecules in cartilage can be suppressed, allowing the exclusive detection of the highly mobile chondroitin sulfate. The technique is also used to detect the chondroitin sulfate in artificial tissue-engineered cartilage. The tissue-engineered material that is based on matrix producing chondrocytes cultured in a collagen gel should provide properties as close as possible to those of the natural cartilage. Nuclear relaxation times of the chondroitin sulfate were determined for both tissues. Although T(1) relaxation times are rather similar, the T(2) relaxation in tissue-engineered cartilage is significantly shorter. This suggests that the motions of chondroitin sulfate in natural and artificial cartilage are different. The nuclear relaxation times of chondroitin sulfate in natural and tissue-engineered cartilage were modeled using a broad distribution function for the motional correlation times. Although the description of the microscopic molecular dynamics of the chondroitin sulfate in natural and artificial cartilage required the identical broad distribution functions for the correlation times of motion, significant differences in the correlation times of motion that are extracted from the model indicate that the artificial tissue does not fully meet the standards of the natural ideal. This could also be confirmed by macroscopic biomechanical elasticity measurements. Nevertheless, these results suggest that NMR is a useful tool for the investigation of the quality of artificially engineered tissue.

Monitoring Cartilage Tissue Engineering Using Magnetic Resonance Spectroscopy, Imaging, and Elastography

“…Special NMR spectroscopy techniques such as highresolution magic angle spinning (HRMAS), cross-polarization magic angle spinning (CPMAS), and heteronuclear decoupling, which reduce line broadening significantly and enhance SNR, particularly for 13 C NMR, have been used for NMR investigations of natural cartilage. 27,28,[66][67][68][69] Water is the most abundant molecule in biological tissues; therefore, the water signal dominates the proton NMR spectra of cartilage. The diffusion of water provides a window into tissue composition and dynamics.…”

“…This article 71 also reported that the residual dipolar coupling derived from proton double-quantum coherence spectroscopy was much smaller for the tissue grown as chondrocyte pellets and chondrocytes seeded in alginate beads when compared with the native tissue, which suggests a lack of preferred orientation in the collagen produced in engineered tissues. 63,71 Scheidt et al 28 used 13 C HRMAS NMR to study the sitespecific motion of 13 C nuclei in chondrocytes seeded in a collagen hydrogel for 3 weeks of culture time. They found that the T 2 relaxation times of the carbon atoms in engineered tissue are shorter than those observed in the native tissue.…”

“…They found that the T 2 relaxation times of the carbon atoms in engineered tissue are shorter than those observed in the native tissue. Schulz et al 20,28 have used 13 C magic angle spinning solid state NMR spectroscopy to quantify cell proliferation, metabolic activity, ECM composition, GAG mobility, and nanoarchitecture of engineered cartilage. Clearly, 13 C NMR spectroscopy is a useful tool for probing cartilage tissue engineering, and it will play an important role in understanding the growth dynamics in vitro and in vivo.…”

“…[24][25][26] It is also noted that the analytical macromolecular content does not wholly represent the complex structural and functional aspects of cartilage, which are assessed by elasticity and molecular mobility in the tissue. 27,28 Magnetic resonance spectroscopy (MRS), MRI, and magnetic resonance elastography (MRE) are noninvasive methods that can provide biochemical, anatomical, and biomechanical measures of tissue. Figure 1 describes exemplary features of MR spectroscopy, imaging, and elastography techniques.…”

“…MRS and imaging are wellestablished techniques that have been used to probe the structure and dynamics of biochemical changes in engineered tissues. [28][29][30][31][32][33][34][35][36][37][38][39][40][41][42] MR elastography, while relatively new, is rapidly being employed to assess the mechanical properties of engineered tissues. 30,43 Using MR spectroscopy, imaging, and elastography together, it is possible to create three-dimensional (3D) maps of chemical shifts, relaxation times, diffusion coefficient, spectral couplings, and tissue stiffness.…”