Micromechanical analysis of native and cross-linked collagen type I fibrils supports the existence of microfibrils.

The mechanical properties of individual collagen fibrils of approximately 200 nm in diameter were determined using a slightly adapted AFM system. Single collagen fibrils immersed in PBS buffer were attached between an AFM cantilever and a glass surface to perform tensile tests at different strain rates and stress relaxation measurements. The stress-strain behavior of collagen fibrils immersed in PBS buffer comprises a toe region up to a stress of 5 MPa, followed by the heel and linear region at higher stresses. Hysteresis and strain-rate dependent stress-strain behavior of collagen fibrils were observed, which suggest that single collagen fibrils have viscoelastic properties. The stress relaxation process of individual collagen fibrils could be best fitted using a two-term Prony series. Furthermore, the influence of different cross-linking agents on the mechanical properties of single collagen fibrils was investigated. Based on these results, we propose that sliding of microfibrils with respect to each other plays a role in the viscoelastic behavior of collagen fibrils in addition to the sliding of collagen molecules with respect to each other. Our finding provides a better insight into the relationship between the structure and mechanical properties of collagen and the micro-mechanical behavior of tissues.

[1]  Kumbakonam R. Rajagopal,et al.  Mechanical Response of Polymers: An Introduction , 2000 .

[2]  Marc Hendriks,et al.  Quantification of carboxyl groups in carbodiimide cross-linked collagen sponges. , 2007, Journal of biomedical materials research. Part A.

[3]  J. Wang.  Mechanobiology of tendon. , 2006, Journal of biomechanics.

[4]  J.J. Wu.  Quantitative Constitutive Behaviour and Viscoelastic Properties of Fresh Flexor Tendons , 2006, The International journal of artificial organs.

[5]  Joseph W Freeman,et al.  Collagen self-assembly and the development of tendon mechanical properties. , 2003, Journal of biomechanics.

[6]  D L Butler,et al.  Comparison of material properties in fascicle-bone units from human patellar tendon and knee ligaments. , 1986, Journal of biomechanics.

[7]  C. Schönenberger,et al.  Nanomechanics of microtubules. , 2002, Physical review letters.

[8]  E. Mosler,et al.  Structural dynamic of native tendon collagen. , 1987, Journal of molecular biology.

[9]  U Ziese,et al.  Corneal collagen fibril structure in three dimensions: Structural insights into fibril assembly, mechanical properties, and tissue organization , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[10]  T. Irving,et al.  Microfibrillar structure of type I collagen in situ. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[11]  J. Revel,et al.  Subfibrillar structure of type I collagen observed by atomic force microscopy. , 1993, Biophysical journal.

[12]  Roberto Ballarini,et al.  Viscoelastic properties of isolated collagen fibrils. , 2011, Biophysical journal.

[13]  J. Feijen,et al.  Glutaraldehyde as a crosslinking agent for collagen-based biomaterials , 1995 .

[14]  P. Fratzl,et al.  A new molecular model for collagen elasticity based on synchrotron X-ray scattering evidence. , 1997, Biophysical journal.

[15]  S. Okuma,et al.  A method for determining the spring constant of cantilevers for atomic force microscopy , 1996 .

[16]  P. Fratzl,et al.  Viscoelastic properties of collagen: synchrotron radiation investigations and structural model. , 2002, Philosophical transactions of the Royal Society of London. Series B, Biological sciences.

[17]  Ren G Dong,et al.  Estimation of the viscous properties of skin and subcutaneous tissue in uniaxial stress relaxation tests. , 2006, Bio-medical materials and engineering.

[18]  M. Koch,et al.  Stress-induced molecular rearrangement in tendon collagen. , 1985, Journal of molecular biology.

[19]  M. R. Dodge,et al.  Stress-strain experiments on individual collagen fibrils. , 2008, Biophysical journal.

[20]  D. Elliott,et al.  Effect of fiber orientation and strain rate on the nonlinear uniaxial tensile material properties of tendon. , 2003, Journal of biomechanical engineering.

[21]  W. Landis,et al.  The role of mineral in the storage of elastic energy in turkey tendons. , 2000, Biomacromolecules.

[22]  P. Hansen,et al.  Viscoelastic behavior of discrete human collagen fibrils. , 2010, Journal of the mechanical behavior of biomedical materials.

[23]  Kai-Nan An,et al.  Flexibility of type I collagen and mechanical property of connective tissue. , 2004, Biorheology.

[24]  A. Oberhauser,et al.  Mechanical design of proteins studied by single-molecule force spectroscopy and protein engineering. , 2000, Progress in biophysics and molecular biology.

[25]  M. Buehler.  Nanomechanics of collagen fibrils under varying cross-link densities: atomistic and continuum studies. , 2008, Journal of the mechanical behavior of biomedical materials.

[26]  Vinod Subramaniam,et al.  Micromechanical bending of single collagen fibrils using atomic force microscopy. , 2007, Journal of biomedical materials research. Part A.

[27]  J. Graham,et al.  Structural changes in human type I collagen fibrils investigated by force spectroscopy. , 2004, Experimental cell research.

[28]  Jan Feijen,et al.  Micromechanical testing of individual collagen fibrils. , 2006, Macromolecular bioscience.

[29]  Alberto Redaelli,et al.  Hierarchical structure and nanomechanics of collagen microfibrils from the atomistic scale up. , 2011, Nano letters.

[30]  Paul K. Hansma,et al.  Bone indentation recovery time correlates with bond reforming time , 2001, Nature.

[31]  Markus J. Buehler,et al.  Nature designs tough collagen: Explaining the nanostructure of collagen fibrils , 2006, Proceedings of the National Academy of Sciences.

[32]  H. Kahn,et al.  Nano measurements with micro-devices: mechanical properties of hydrated collagen fibrils , 2006, Journal of The Royal Society Interface.

[33]  D. Hulmes,et al.  Building collagen molecules, fibrils, and suprafibrillar structures. , 2002, Journal of structural biology.

[34]  Jan Feijen,et al.  Mechanical properties of native and cross-linked type I collagen fibrils. , 2008, Biophysical journal.

[35]  A. Dart,et al.  Mechanical and functional properties of the equine superficial digital flexor tendon. , 2005, Veterinary journal.

[36]  A. Oberhauser,et al.  The study of protein mechanics with the atomic force microscope. , 1999, Trends in biochemical sciences.

[37]  G. Pharr,et al.  Measurement of hardness and elastic modulus by instrumented indentation: Advances in understanding and refinements to methodology , 2004 .

[38]  Christopher M Dobson,et al.  Characterization of the nanoscale properties of individual amyloid fibrils , 2006, Proceedings of the National Academy of Sciences.

[39]  A. Zink,et al.  Structural investigations on native collagen type I fibrils using AFM. , 2007, Biochemical and biophysical research communications.

[40]  N. Sasaki,et al.  Elongation mechanism of collagen fibrils and force-strain relations of tendon at each level of structural hierarchy. , 1996, Journal of biomechanics.

[41]  D. Bader,et al.  An investigation into the effects of the hierarchical structure of tendon fascicles on micromechanical properties , 2004, Proceedings of the Institution of Mechanical Engineers. Part H, Journal of engineering in medicine.

[42]  M. Kjaer,et al.  Tendon properties in relation to muscular activity and physical training , 2003, Scandinavian journal of medicine & science in sports.

[43]  Mhj Koch,et al.  Quantitative analysis of the molecular sliding mechanisms in native tendon collagen — time-resolved dynamic studies using synchrotron radiation , 1987 .

[44]  M Raspanti,et al.  Tapping-mode atomic force microscopy in fluid of hydrated extracellular matrix. , 2001, Matrix biology : journal of the International Society for Matrix Biology.

[45]  Mehdi Balooch,et al.  In situ atomic force microscopy of partially demineralized human dentin collagen fibrils. , 2002, Journal of structural biology.

[46]  J. W. SMITH,et al.  Molecular Pattern in Native Collagen , 1968, Nature.

[47]  T L Haut,et al.  The state of tissue hydration determines the strain-rate-sensitive stiffness of human patellar tendon. , 1997, Journal of biomechanics.

[48]  Laurent Bozec,et al.  Mechanical properties of collagen fibrils. , 2007, Biophysical journal.

[49]  M Raspanti,et al.  Hierarchical structures in fibrillar collagens. , 2002, Micron.

[50]  J. Feijen,et al.  Cross-linking of dermal sheep collagen using a water-soluble carbodiimide. , 1996, Biomaterials.

[51]  Louis J Soslowsky,et al.  Strain-rate sensitive mechanical properties of tendon fascicles from mice with genetically engineered alterations in collagen and decorin. , 2004, Journal of biomechanical engineering.

[52]  U. Aebi,et al.  Exploring the mechanical properties of single vimentin intermediate filaments by atomic force microscopy. , 2006, Journal of molecular biology.

[53]  S. Magnusson,et al.  Tensile properties of human collagen fibrils and fascicles are insensitive to environmental salts. , 2010, Biophysical journal.

[54]  A Viidik,et al.  Effects of age on the stress-strain and stress-relaxation properties of the rat molar periodontal ligament. , 2004, Archives of oral biology.

[55]  Jürgen Engel,et al.  Versatile Collagens in Invertebrates , 1997, Science.

[56]  F. Silver,et al.  A self-assembled collagen scaffold suitable for use in soft and hard tissue replacement , 1995 .