Effects of decorin proteoglycan on fibrillogenesis, ultrastructure, and mechanics of type I collagen gels

Abstract

The proteoglycan decorin is known to affect both the fibrillogenesis and the resulting ultrastructure of in vitro polymerized collagen gels. However, little is known about its effects on mechanical properties. In this study, 3D collagen gels were polymerized into tensile test specimens in the presence of decorin proteoglycan, decorin core protein, or dermatan sulfate (DS). Collagen fibrillogenesis, ultrastructure, and mechanical properties were then quantified using a turbidity assay, 2 forms of microscopy (SEM and confocal), and tensile testing. The presence of decorin proteoglycan or core protein decreased the rate and ultimate turbidity during fibrillogenesis and decreased the number of fibril aggregates (fibers) compared to control gels. The addition of decorin and core protein increased the linear modulus by a factor of 2 compared to controls, while the addition of DS reduced the linear modulus by a factor of 3. Adding decorin after fibrillogenesis had no effect, suggesting that decorin must be present during fibrillogenesis to increase the mechanical properties of the resulting gels. These results show that the inclusion of decorin proteoglycan during fibrillogenesis of Type I collagen increases the modulus and tensile strength of resulting collagen gels. The increase in mechanical properties when polymerization occurs in the presence of the decorin proteoglycan is due to a reduction in the aggregation of fibrils into larger order structures such as fibers and fiber bundles.

Nanoscale Imaging of Collagen Gels with Focused Ion Beam Milling and Scanning Electron Microscopy

“…Altering these variables changes the rate of polymerization and the resulting diameter distributions and organization of fibrils within the collagen network. Such results are highly relevant, as numerous experimental and computational studies have demonstrated that the microstructural organization of collagen fibrils plays a central role in the resulting macroscopic properties of gels in vitro and tissues in vivo (15,(18)(19)(20)(21), as well as the emergent cellular behavior observed in culture.…”

“…In vitro polymerization of type I collagen has played a crucial role as both a cell culture scaffold and as a model system to study the process of fibrillogenesis (14)(15)(16). The fibril network of a collagen gel is tightly linked to the polymerization conditions, including collagen source, collagen concentration, pH, temperature, and salt content, presence of other proteins and molecules, and other physical and chemical factors (15)(16)(17).…”

“…The fibril network of a collagen gel is tightly linked to the polymerization conditions, including collagen source, collagen concentration, pH, temperature, and salt content, presence of other proteins and molecules, and other physical and chemical factors (15)(16)(17). Altering these variables changes the rate of polymerization and the resulting diameter distributions and organization of fibrils within the collagen network.…”

“…Macroscopic properties are assessed using turbidity assays to study the kinetics of fibrillogenesis, mechanical testing to study the elastic, plastic, and viscoelastic properties of gels and a number of other assays to examine the permeability and diffusion coefficients of collagen gels (15,16,(22)(23)(24). The microscopic organization of collagen gels has been studied using a number of different microscopy techniques, including optical methods such as confocal reflectance microscopy, confocal fluorescence microscopy, second harmonic generation, and electron microscopy methods such as scanning electron microscopy (SEM) and transmission electron microscopy (TEM) ( Fig.…”

“…The microscopic organization of collagen gels has been studied using a number of different microscopy techniques, including optical methods such as confocal reflectance microscopy, confocal fluorescence microscopy, second harmonic generation, and electron microscopy methods such as scanning electron microscopy (SEM) and transmission electron microscopy (TEM) ( Fig. 1) (15,(25)(26)(27)(28)(29). The major advantage of the aforementioned optical methods is that they are inherently three dimensional (3D).…”