Ultrasound has potential as a non-destructive analytical technique to provide real-time online assessments of matrix evolution in cell-hydrogel constructs used in tissue engineering. In these studies, chondrocytes were encapsulated in poly(ethylene glycol) hydrogels, and gel degradation was manipulated to provide conditions with varying distribution of the large cartilage extracellular matrix molecule, collagen. Mechanical properties and matrix accumulations were simultaneously measured for each condition during 9.5 weeks of in vitro culture. Ultrasound data were used to construct cross-sectional B-scan images for qualitative observations of evolving constructs. Ultrasound data were also analyzed to calculate the speed of sound (SoS) and slope of attenuation (SoA) in developing constructs and a non-degrading hydrogel control without encapsulated chondrocytes. SoS and SoA were calculated from 50 and 100MHz ultrasound data, and sample correlation coefficients were calculated to identify important relationships between these ultrasound parameters and mechanical/biochemical properties of the evolving matrix. SoA appears to be more sensitive to the density of accumulated matrix molecules than SoS, while SoS appeared to be more sensitive to mechanical modulus than SoA in measurements performed at 100MHz. Correlation analysis revealed that ultrasound measurements at 100MHz are more likely to be good predictors of matrix evolution and neotissue function than measurements at 50MHz. Rigorous studies of the relationships identified in this study will lead to non-destructive real-time monitoring that will be useful in combination with bioreactors used to promote and study cartilage regeneration.
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