Externally applied boundary conditions regulate neovessel sprouting and elongation during angiogenesis, affecting both neovessel growth characteristics and network morphometry. Furthermore, neovessels align parallel to the direction of stress/strain or internally generated traction, and this may be because of collagen fibril alignment induced by the growing neovessels themselves.

Angiogenesis is regulated by the local microenvironment, including the mechanical interactions between neovessel sprouts and the extracellular matrix (ECM). However, the mechanisms controlling the relationship of mechanical and biophysical properties of the ECM to neovessel growth during sprouting angiogenesis are just beginning to be understood. In this research, we characterized the relationship between matrix density and microvascular topology in an in vitro 3D organ culture model of sprouting angiogenesis. We used these results to design and calibrate a computational growth model to demonstrate how changes in individual neovessel behavior produce the changes in vascular topology that were observed experimentally. Vascularized gels with higher collagen densities produced neovasculatures with shorter vessel lengths, less branch points, and reduced network interconnectivity. The computational model was able to predict these experimental results by scaling the rates of neovessel growth and branching according to local matrix density. As a final demonstration of utility of the modeling framework, we used our growth model to predict several scenarios of practical interest that could not be investigated experimentally using the organ culture model. Increasing the density of the ECM significantly reduced angiogenesis and network formation within a 3D organ culture model of angiogenesis. Increasing the density of the matrix increases the stiffness of the ECM, changing how neovessels are able to deform and remodel their surroundings. The computational framework outlined in this study was capable of predicting this observed experimental behavior by adjusting neovessel growth rate and branching probability according to local ECM density, demonstrating that altering the stiffness of the ECM via increasing matrix density affects neovessel behavior, thereby regulated vascular topology during angiogenesis.

The glycosaminoglycan of decorin, dermatan sulfate (DS), has been suggested to contribute to the mechanical properties of soft connective tissues such as ligaments and tendons. This study investigated the mechanical function of DS in human medial collateral ligaments (MCL) using nondestructive shear and tensile material tests performed before and after targeted removal of DS with chondroitinase B (ChB). The quasi-static elastic material properties of human MCL were unchanged after DS removal. At peak deformation, tensile and shear stresses in ChB treated tissue were within 0.5% (p > 0.70) and 2.0% (p > 0.30) of pre-treatment values, respectively. From preto post-ChB treatment under tensile loading, the tensile tangent modulus went from 242 AE 64 to 233 AE 57 MPa (p ¼ 0.44), and tissue strain at peak deformation went from 4.3 AE 0.3% to 4.4 AE 0.3% (p ¼ 0.54). Tissue hysteresis was unaffected by DS removal for both tensile and shear loading. Biochemical analysis confirmed that 90% of DS was removed by ChB treatment when compared to control samples, and transmission electron microscopy (TEM) imaging further verified the degradation of DS by showing an 88% reduction (p < .001) of sulfated glycosaminoglycans in ChB treated tissue. These results demonstrate that DS in mature knee MCL tissue does not resist tensile or shear deformation under quasi-static loading conditions, challenging the theory that decorin proteoglycans contribute to the elastic material behavior of ligament. ß

The viscoelastic properties of human ligament potentially guard against structural failure, yet the microstructural origins of these transient behaviors are unknown. Glycosaminoglycans (GAGs) are widely suspected to affect ligament viscoelasticity by forming molecular bridges between neighboring collagen fibrils. This study investigated whether GAGs directly affect viscoelastic material behavior in human medial collateral ligament (MCL) by using nondestructive tensile tests before and after degradation of GAGs with chondroitinase ABC (ChABC). Control and ChABC treatment (83% GAG removal) produced similar alterations to ligament viscoelasticity. This finding was consistent at different levels of collagen fiber stretch and tissue hydration. On average, stress relaxation increased after incubation by 2.2% (control) and 2.1% (ChABC), dynamic modulus increased after incubation by 3.6% (control) and 3.8% (ChABC), and phase shift increased after incubation by 8.5% (control) and 8.4% (ChABC). The changes in viscoelastic behavior after treatment were significantly more pronounced at lower clamp-to-clamp strain levels. A 10% difference in the water content of tested specimens had minor influence on ligament viscoelastic properties. The major finding of this study is that mechanical interactions between collagen fibrils and GAGs are unrelated to tissue-level viscoelastic mechanics in mature human MCL. These findings narrow the possible number of extracellular matrix molecules that have a direct contribution to ligament viscoelasticity.