Collagen Mechanical Properties Research Articles

A Nonlinear Constituent Based Viscoelastic Model for Articular Cartilage and Analysis of Tissue Remodeling Due to Altered Glycosaminoglycan-Collagen Interactions

A constituent based nonlinear viscoelastic (VE) model was modified from a previous study (Vena, et al., 2006, "A Constituent-Based Model for the Nonlinear Viscoelastic Behavior of Ligaments," J. Biomech. Eng., 128, pp. 449-457) to incorporate a glycosaminoglycan (GAG)-collagen (COL) stress balance using compressible elastic stress constitutive equations specific to articular cartilage (AC). For uniaxial loading of a mixture of quasilinear VE constituents, time constant and relaxation ratio equations are derived to highlight how a mixture of constituents with distinct quasilinear VE properties is one mechanism that produces a nonlinear VE tissue. Uniaxial tension experiments were performed with newborn bovine AC specimens before and after approximately 55% and approximately 85% GAG depletion treatment with guanidine. Experimental tissue VE parameters were calculated directly from stress relaxation data, while intrinsic COL VE parameters were calculated by curve fitting the data with the nonlinear VE model with intrinsic GAG viscoelasticity neglected. Select tissue and intrinsic COL VE parameters were significantly different from control and experimental groups and correlated with GAG content, suggesting that GAG-COL interactions exist to modulate tissue and COL mechanical properties. Comparison of the results from this and other studies that subjected more mature AC tissue to GAG depletion treatment suggests that the GAGs interact with the COL network in a manner that may be beneficial for rapid volumetric expansion during developmental growth while protecting cells from excessive matrix strains. Furthermore, the underlying GAG-COL interactions appear to diminish as the tissue matures, indicating a distinctive remodeling response during developmental growth.

Image Correlation Spectroscopy of Multiphoton Images Correlates with Collagen Mechanical Properties

Multiphoton microscopy (MPM) holds promise as a noninvasive imaging technique for characterizing collagen structure, and thus mechanical properties, through imaging second harmonic generation (SHG) and two-photon fluorescence in engineered and real connective tissues. Controlling polymerization pH to manipulate collagen gel microstructure, we quantified pore and fiber dimensions using both standard methods and image correlation spectroscopy (ICS) on MPM, scanning electron, and darkfield microscopy images. The latter two techniques are used to confirm microstructural measurements made from MPM images. As polymerization pH increased from 5.5 to 8.5, mean fiber diameter decreased from 3.7 ± 0.7 μm to 1.6 ± 0.3 μm, the average pore size decreased from 81.7 ± 3.7 μm 2 to 7.8 ± 0.4 μm 2, and the pore area fraction decreased from 56.8% ± 0.8% to 18.0% ± 1.3% (measured from SHG images), whereas the storage modulus G′ and loss modulus G′, components of the shear modulus, increased ∼33-fold and ∼16-fold, respectively. A characteristic length scale measured using ICS, W ICS, correlates well with the mean fiber diameter from SHG images ( R 2 = 0.95). Semiflexible network theory predicts a scaling relationship of the collagen gel storage modulus ( G′) depending upon mesh size and fiber diameter, which are estimated from SHG images using ICS. We conclude that MPM and ICS are an effective combination to assess bulk mechanical properties of collagen hydrogels in a noninvasive, objective, and systematic fashion and may be useful for specific in vivo applications.

OS1107 コラーゲン薄膜の機械的特性評価(OS11-02 生体材料・医療材料の表面機能・構造と評価2,OS11 生体材料・医療材料の表面機能・構造と評価)

OS1107 コラーゲン薄膜の機械的特性評価(OS11-02 生体材料・医療材料の表面機能・構造と評価2,OS11 生体材料・医療材料の表面機能・構造と評価)

Atomic Force Microscopy Investigation of Vocal Fold Collagen

Collagen is an important constituent of the vocal fold extracellular matrix and is necessary for providing tensile strength and maintaining tissue geometry. Traditional investigations of vocal fold collagen using light and electron microscopy do not provide information on the organization and mechanical properties of collagen in native topographic state. The primary objective of this study was to use Atomic Force Microscopy (AFM) to examine the surface characteristics and organization of collagen in the deep layer of the lamina propria at nanoscale resolution. Experimental in vitro design. Freshly dissected porcine vocal folds were mounted on AFM discs and imaged under contact and tapping mode to obtain information on topographic distribution of collagen. AFM imaging of the deep layer of the lamina propria revealed dense, abundant collagen fibers with a characteristic banding pattern. The distribution of collagen was heterogeneous, with both hydrophilic and hydrophobic regions within a sample. AFM offers a useful tool to obtain topographic information about biologic samples at nanoscale resolution with minimal sample preparation. Mapping the topography and mechanical properties of vocal fold collagen is necessary for designing rheologically compatible bioimplants for the treatment of dysphonia resulting from vocal fold scarring and bowing.

Analysis of articular cartilage as a composite using nonlinear membrane elements for collagen fibrils

To develop a composite fibre-reinforced model of the cartilage, membrane shell elements were introduced to represent collagen fibrils reinforcing the isotropic porous solid matrix filled with fluid. Nonlinear stress–strain curve of pure collagen fibres and collagen volume fraction were explicitly presented in the formulation of these membrane elements. In this composite model, in accordance with tissue structure, the matrix and fibril membrane network experienced dissimilar stresses despite identical strains in the fibre directions. Different unconfined compression and indentation case studies were performed to determine the distinct role of membrane collagen fibrils in nonlinear poroelastic mechanics of articular cartilage. The importance of nonlinear fibril membrane elements in the tissue relaxation response as well as in temporal and spatial variations of pore pressure and solid matrix stresses was demonstrated. By individual adjustments of the collagen volume fraction and collagen mechanical properties, the model allows for the simulation of alterations in the fibril network structure of the tissue towards modelling damage processes or repair attempts. The current model, which is based on a physiological description of the tissue structure, is promising in improvement of our understanding of the cartilage pathomechanics.

Changes in mechanical properties and cellularity during long‐term culture of collagen fiber ACL reconstruction scaffolds

Resorbable scaffolds for anterior cruciate ligament (ACL) reconstruction should provide temporary mechanical function then gradually breakdown while promoting matrix synthesis by local cells. Crosslinking influences collagen's mechanical properties, degradation rate, and interactions with cells. Our objective was to compare the effects of different crosslinkers on cellularity and mechanical properties during long-term (8 week) culture of collagen fiber scaffolds. Fibers were fabricated from an acid-insoluble dispersion of bovine dermal collagen and crosslinked with either ultraviolet irradiation (UV; a physical crosslinker) or 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC; a chemical crosslinker). Scaffolds consisted of 50 fibers bundled in parallel. Initial attachment of fibroblasts was similar on both scaffolds; however, from 1 to 8 weeks in culture, UV-crosslinked scaffolds had significantly more cells attached than EDC-crosslinked scaffolds. The initial breaking load (3.50 N) and stiffness (2.23 N/mm) of EDC-crosslinked scaffolds were significantly greater than those of UV-crosslinked scaffolds (2.32 N; 1.21 N/mm) and were unaffected by long-term fibroblast culture. In contrast, the load-bearing capacity of fibroblast-seeded UV-crosslinked scaffolds decreased 60% to 0.91 N after 8 weeks in culture. EDC-crosslinked scaffolds maintained strength and moderate cellularity; UV-crosslinked scaffolds, in contrast, were highly cellular, but had poor mechanical properties that decreased during culture. These in vitro results suggest that collagen fiber scaffolds crosslinked with EDC may be more suitable for ACL reconstruction than those crosslinked with UV.

Influence of laboratory ware related changes in conformational and mechanical properties of collagen

AbstractEnzyme‐treated acid‐solubilized bovine collagen (ESBC Type I) prepared in laboratory‐grade glass and polypropylene ware under identical conditions have shown different conformational and mechanical properties. Circular dichroism (CD) studies on ESBC have shown higher triple‐helical contents in polypropylene laboratory‐grade ware compared to laboratory ware of glass. Fourier transform infrared spectroscopy (FTIR) and differential scanning calorimetric (DSC) studies have supported the conformational changes performed on both preparations. Both preparations, in membrane form, were evaluated for their mechanical properties—viz., shrinkage temperature (St), dissolution temperature (Dt), and tensile strength (Ts). The collagen membrane of ESBC prepared in polypropylene showed significantly higher values compared to glass ones. No difference was seen in shrinkage temperature data of the two membranes. Reduced viscosity of ESBC was also measured for both preparations, and it had a significantly higher value in polypropylene ware compared to ESBC prepared in glassware. We postulate that the presence of silicate ions (SiO3−2) in glassware has a role in all these changes. © 2003 Wiley Periodicals, Inc. J Appl Polym Sci 87: 2186–2192, 2003

Fibrillar Structure and Mechanical Properties of Collagen

Collagen type I is among the most important stress-carrying protein structures in mammals. Despite their importance for the outstanding mechanical properties of this tissue, there is still a lack of understanding of the processes that lead to the specific shape of the stress–strain curve of collagen. Recentin situsynchrotron X-ray scattering experiments suggest that several different processes could dominate depending on the amount of strain. While at small strains there is a straightening of kinks in the collagen structure, first at the fibrillar then at the molecular level, higher strains lead to molecular gliding within the fibrils and ultimately to a disruption of the fibril structure. Moreover, it was observed that the strain within collagen fibrils is always considerably smaller than in the whole tendon. This phenomenon is still very poorly understood but points toward the existence of additional gliding processes occurring at the interfibrillar level.

Stress-strain curve and young's modulus of a collagen molecule as determined by the X-ray diffraction technique

The purpose of the work described in this paper was to make a stress-strain curve for a collagen molecule and estimate Young's modulus of a molecule along the molecular axis. X-ray diffractometry was performed on bovine Achilles tendon in order to measure strain in the collagen molecule along the molecular axis as a response to a macroscopically applied force. By geometrical calculations and experiments, cross-sectional areas of a molecule and molecules in a tendon collagen fiber were determined. The applied force was translated to the stress and the stress-strain curve of the collagen molecule was constructed, which was found to be almost linear. Young's modulus of the molecule was determined to be slightly smaller than when determined by dynamic mechanical methods. The difference was considered to suggest the existence of a viscoelastic component within the molecule as well as the difference in the mechanical properties of collagen in different tissues. The expected viscoelasticity was speculated to be related to the hydrogen bond network in the collagen molecule.

The Effect of Two Types of Cross-Linking on Some Mechanical Properties of Collagen

Samples of collagen were cross-linked by two different methods: (a) glutaraldehyde and (b) a combination of dehydrothermal treatment and cyanamide. The elastic modulus, the ultimate tensile strength (fracture stress), strain to failure, work of fracture, and fracture toughness were measured before and after cross-linking in ambient laboratory conditions, and during immersion in water. These tests were all performed over a range of strain rates. For collagen tested in the wet condition, it was found that cross-linking increased the elastic modulus from approximately 25-30 MPa, to between 55 and 60 MPa, but there was little effect on fracture stress, and strain to failure was reduced. The work of fracture of the collagen decreased on cross-linking. Cross-linking had the same effect on the elastic modulus, fracture stress, and strain to failure of dry collagen, but the work of fracture was unaffected. In conclusion, cross-linking increased the elastic modulus, reduced the strain to failure, and had little effect on the fracture stress of collagen under the present experimental conditions.

Hierarchical structure and mechanical properties of collagen in the intervertebral disc

Hierarchical structure and mechanical properties of collagen in the intervertebral disc

Local effects of impaired mechanical properties of collagen on bone formation and resorption

To study the relationship between the mechanical properties of collagen and the bone turnover, 2-week-old Balb/C mice were fed on a diet containing 0.25% Beta aminopropionitrile (B-APN), a potent inhibitor of collagen crosslink formation, for 3 weeks. Mandibular incisor socket was selected for the analysis of bone formation and resorption parameters. Plastic embedded sections stained with toluidine blue and cut at 4 microns were used to analyze the average area of bone lamellae, bone-forming surface, and the number of osteoblasts/mm of forming surface. Similar sections were used to localize acid phosphatase on resorbing surfaces and within the osteoclasts, while bone alkaline phosphatase was determined by a colorimetric method. Morphometric analyses showed that the area of newly formed bone lamellae, total forming surface, number of osteoblasts and the Alk. Pase activity were significantly lower in B-APN-fed mice as compared to the controls. There was a concomitant smaller, but significant, reduction in total resorption surface, active resorption surface and the number of osteoclasts. These results suggest that the regulation of bone formation and resorption at this site, which is independent of systemic regulation, is influenced by the mechanical properties of the collagenous matrix, which in turn may have a significant effect on the existing pool of bone-forming cells, but may not influence the recruitment of new cells.

Medial Collagen Organization in Human Arteries of the Heart and Brain by Polarized Light Microscopy

The mechanical properties of collagen as a biopolymer ensures that collagen has a significant influence on the mechanical behavior of the host tissue. Structural organization is a key to that influence. We have assessed this relationship quantitatively in the tunica media of arteries from the heart and brain, using the polarizing light microscope and Universal stage. Arteries from 22 autopsies were isolated, cannulated and fixed with 10% buffered formalin, at a distending pressure spanning normal values in vivo. We prepared the tissue for light microscopy, with paraffin embedding, sectioning at 7 microns, and staining with picrosirius red to enhance the natural birefringence of medial collagen. Individual measurements, 30 to 50 per arterial section, referenced against the central axis of the vessel segment, revealed a coherent organization, with an average orientation which was within 1 to 2 degrees of being perfectly concentric for all artery segments. Analysis was done with Lambert projections and circular statistics. We calculated the circular standard deviation, which was 5.2 degrees for 27 brain arteries (S.D. 1.9 degrees) and 5.6 degrees (S.D. 2.1 degrees), for 5 coronary arteries sectioned at less than 15 degrees. Our interpretation is that medial collagen can be strained even though highly aligned, revealing a mechanical property which contrasts that of type I collagen.

Connecting incremental shear modulus and Poisson's ratio of lung tissue with morphology and rheology of microstructure.

The width and curvature of the collagen and elastin fiber bundles in the human pulmonary interalveolar septa and alveolar mouths are measured. The data, together with the known mechanical properties of collagen and elastin fibers, are used to derive the incremental elastic moduli of the lung tissue. The constitutive equation for small incremental stress and strain superposed on a homeostatic inflated lung is linear and isotropic, and characterized by two material constants.

The influence of non-enzymatic glycosylation and formation of fluorescent reaction products on the mechanical properties of rat tail tendons.

Mechanical stability was examined in rat tail tendons after in vitro incubation in glucose at pH 7.4 using buffer systems of either phosphate or tris(hydroxymethyl)aminomethan. In the phosphate buffer system glucose and fluorescent compounds were found to be attached to the collagen molecules and the maximum 'stress' of the tendons was increased. In the tris(hydroxymethyl)aminomethan buffer system glucose was attached to the collagen molecules, but only small amounts of fluorescent compounds were attached to the collagen molecules and no changes in mechanical parameters were recorded. Initial incubation of tendons in a high concentration of glucose followed by incubation in either phosphate or tris(hydroxymethyl)aminomethan buffer solutions resulted in equal attachment of glucose to the collagen, but only the collagen reincubated in phosphate buffer developed a relatively high amount of fluorescent compounds and an increase in maximum mechanical 'stress' compared to the collagen reincubated in tris(hydroxymethyl)aminomethan buffer. This shows that the non-enzymatic glycosylation per se does not necessarily influence the mechanical properties of collagen. Additional reactions like transformation of glucose into fluorescent compounds or browning reaction products by Maillard's reaction seem to be essential.

Mechanical properties of collagen from decalcified rat femur in relation to age and in vitro maturation.

Cortical bone collagen was obtained by decalcifying femoral bones from 2-, 5-, 15-, and 25-month-old male rats. Collagen specimens were cut longitudinal to the long axis of the femur and tested mechanically. The maximum load (ultimate strength) and maximum slope of the load-strain curve (maximum stiffness) were found to decrease with age. The age-related reduction in the mechanical parameters resulted from a change in the mechanical strength of the constituent collagen and a change in the morphology of the bone tissue. In vitro aging, produced by incubating for 0-5 months, did not change the mechanical strength of bone collagen specimens obtained from 2-month-old rats. The changes in the mechanical characteristics of bone collagen, accompanying in vivo and in vitro aging are the opposite of those observed in soft tissue collagen. In the latter there is an increase in the mechanical strength during in vivo and in vitro aging.

Effects of Relaxin on the Mechanical Properties of Collagen Rich Tissues

ABSTRACTA collagenous scar capsule forms around biomaterial implants and for deformable implants such as a breast prosthesis, this phenomenon of capsular scar contracture may lead to deformation and hardening of the implant. Relaxin, a peptide hormone of pregnancy which is known to affect collagen rich tissues, was given to mice in an attempt to eliminate or lessen this contracture. A short (8 day) study was performed to observe the effects of relaxin and estrogen on the mechanical properties of Skin and the interpubic ligament. A longer (27 day) study observed the effects of relaxin in estrogen primed mice containing implants. Changes in skin, interpubic ligament, scar capsule and contracture phenomenon were noted. Stress-strain characterization, dynamic mechanical spectroscopy and light microscopy were used to characterize the effects of relaxin and estradiol on the contracture phenomenon. Qualitatively, relaxin given to estrogen primed mice was shown to lessen implant contracture and deformability. Administration of relaxin produced the documented changes in the interpubic ligament, but differences were noted between the 8 and 27 day studies. Differences in the failure stress and strain and modulus for the long implant and short non-implant study were noted for the relaxin treated mice.

On fundamental and phenomenological models, structure and mechanical properties of collagen, elastin and glycosaminoglycan complexes.

This paper reviews the structure (from molecular to macroscopic level) of collagen, elastin and glycosaminoglycans, with special reference to their functional properties and to how their behavior can be elucidated from in vitro manipulations of the tissues by biochemical means. Modes to analyse the mechanical behavior are briefly discussed and exemplified by data from reconstituted collagenous "tissue". It is concluded that the basic equation for the "non-linear elasticity" component in Fung's law (and for the reconstituted "tissue" a modification of it) is a powerful tool for analysis of the physiological range of the stress-strain curve. Further, enzymatic "dissection" of one tissue element at a time provides a valuable method for the analysis of tissue element interactions. This approach is illustrated with data from aortic tissue. It is shown that the mechanical properties of the aorta depend on the interaction between elasticity and collagenous elements and that the strength of the tissue is not derived from its collagen fibers per se.

An ultrastructural and cytochemical analysis of the cellular basis for tyrosine-derived collagen crosslinks in Leptogorgia virgulata (Cnidaria: Gorgonacea).

Electron-microscopical autoradiography and cytochemical techniques have been used to identify the distinct and separate subcellular structures involved in the secretion of 1) procollagen, 2) dihydroxyphenylalanine (DOPA), which is a precursor of a collagen-crosslinking compound, and 3) DOPA oxidase, which converts DOPA to a putative crosslinking compound of collagen in the axial skeleton of the gorgonian coral Leptogorgia virulata. Some skeletal-protein hydrolysates contain material that co-elutes with DOPA. The data indicate that these skeletogenic cells, corticocytes, are capable of modifying the number of non-reducible, tyrosine-derived crosslinkages of collagen by the secretion of a crosslinking compound that acts extracellularly on collagen. A mechanism for a cell-mediated control of the mechanical properties of collagen is thereby presented.