Journal
JOURNAL OF BIOMECHANICS
Volume 39, Issue 14, Pages 2555-2565Publisher
ELSEVIER SCI LTD
DOI: 10.1016/j.jbiomech.2005.09.007
Keywords
cartilage; aggrecan; atomic force microscopy; electromechanical properties; Poisson-Boltzmann
Categories
Funding
- NIAMS NIH HHS [AR33236] Funding Source: Medline
- NATIONAL INSTITUTE OF ARTHRITIS AND MUSCULOSKELETAL AND SKIN DISEASES [R37AR033236, R01AR033236] Funding Source: NIH RePORTER
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In this study, we have measured the nanoscale compressive interactions between opposing aggrecan macromolecules in near-physiological conditions, in order to elucidate the molecular origins of tissue-level cartilage biomechanical behavior. Aggrecan molecules from fetal bovine epiphyseal cartilage were chemically end-grafted to planar substrates, standard nanosized atomic force microscopy (AFM) probe tips (R-tip similar to 50 nm), and larger colloidal probe tips (R-tip similar to 2.5 mu m). To assess normal nanomechanical interaction forces between opposing aggrecan layers, substrates with microcontact printed aggrecan were imaged using contact mode AFM, and aggrecan layer height (and hence deformation) was measured as a function of solution ionic strength (IS) and applied normal load. Then, using high-resolution force spectroscopy, nanoscale compressive forces between opposing aggrecan on the tip and substrate were measured versus tip-substrate separation distance in 0.001-1 M NaCl. Nanosized tips enabled measurement of the molecular stiffness of 2-4 aggrecan while colloidal tips probed the nanomechanical properties of larger assemblies (similar to 10(4) molecules). The compressive stiffness of aggrecan was much higher when using a densely packed colloidal tip than the stiffness measured for using the nanosized tip with a few aggrecan, demonstrating the importance of lateral interactions to the normal nanomechanical properties. The measured stress at 0.1 M NaCl (near-physiological ionic strength) increased sharply at aggrecan densities under the tip of similar to 40 mg/ml (physiological densities are similar to 20-80 mg/ml), corresponding to an average inter-GAG spacing of 4-5 Debye lengths (4 5 nm); this characteristic spacing is consistent with the onset of significant electrostatic interactions between GAG chains of opposing aggrecan molecules. Comparison of nanomechanical data to the predictions of Poisson-Boltzmann-based models further elucidated the regimes over which electrostatic and nonelectrostatic interactions affect aggrecan stiffness in compression. The most important aspects of this study include: the incorporation of experiments at two different length scales, the use of microcontact printing to enable quantification of aggrecan deformation and the corresponding nanoscale compressive stress vs. strain curve, the use of tips of differing functionality to provide insights into the molecular mechanisms of deformation, and the comparison of experimental data to the predictions of three increasingly refined Poisson-Boltzmann (P-B)-based theoretical models for the electrostatic double layer component of the interaction (c) 2005 Elsevier Ltd. All rights reserved.
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