Functional properties of cell-seeded three-dimensionally woven poly(epsilon-caprolactone) scaffolds for cartilage tissue engineering

Abstract

Articular cartilage possesses complex mechanical properties that provide healthy joints the ability to bear repeated loads and maintain smooth articulating surfaces over an entire lifetime. In this study, we utilized a fiber-reinforced composite scaffold designed to mimic the anisotropic, nonlinear, and viscoelastic biomechanical characteristics of native cartilage as the basis for developing functional tissue-engineered constructs. Three-dimensionally woven poly(epsilon-caprolactone) (PCL) scaffolds were encapsulated with a fibrin hydrogel, seeded with human adipose-derived stem cells, and cultured for 28 days in chondrogenic culture conditions. Biomechanical testing showed that PCL-based constructs exhibited baseline compressive and shear properties similar to those of native cartilage and maintained these properties throughout the culture period, while supporting the synthesis of a collagen-rich extracellular matrix. Further, constructs displayed an equilibrium coefficient of friction similar to that of native articular cartilage (mu(eq) approximately 0.1-0.3) over the prescribed culture period. Our findings show that three-dimensionally woven PCL-fibrin composite scaffolds can be produced with cartilage-like mechanical properties, and that these engineered properties can be maintained in culture while seeded stem cells regenerate a new, functional tissue construct.

FIG. 1.

Fiber architecture of a three-dimensionally orthogonally woven poly(ɛ-caprolactone) (PCL) structure. Three-dimensional structures were woven by interlocking multiple layers of two perpendicularly oriented sets of in-plane fibers (x- or warp direction, and y- or weft direction) with a third set of fibers in the z-direction. (A) Surface view of the X–Y plane (standard error of the mean [SEM]), (B) cross-sectional view of the X–Z plane, and (C) cross-sectional view of the Y–Z plane.

FIG. 2.

The compressive stiffness of three-dimensional (3D) PCL scaffolds was increased when consolidated with fibrin. Aggregate modulus (H_A) and Young's modulus (E) at day 0 as determined by confined and unconfined compression, respectively. The 3D PCL–fibrin composite scaffolds had significantly higher H_A and E values than did naked 3D PCL scaffolds (analysis of variance [ANOVA], *p < 0.05, **p < 0.0001). Data presented as mean ± SEM.

FIG. 3.

Compressive biomechanical properties of scaffolds at days 0, 14, and 28. (A) Aggregate modulus (H_A) and (B) Young's modulus (E) as determined by confined and unconfined compression, respectively. The addition of fibrin to 3D PCL scaffolds significantly increased H_A and E for both cellular and acellular groups (ANOVA, *p < 0.05, **p < 0.0001). (C) Hydraulic permeability (k) as determined by curve-fitting creep tests using a numerical least-squares regression procedure. Acellular 3D PCL scaffolds displayed significantly higher k-values than all other groups at days 14 and 28 (ANOVA, *p < 0.0001). Data represented as mean ± SEM.

FIG. 4.

Shear biomechanical properties of cultured scaffolds at days 14 and 28. (A) Complex shear modulus (G*) and (B) loss angle (δ) measured at ω = 10 rad/s and γ_o = 0.05. Adipose-derived stem cell–synthesized extracellular matrix significantly increased the shear stiffness of both 3D PCL and 3D PCL–fibrin composite scaffolds (ANOVA, *p < 0.05, **p < 0.0001). Data represented as mean ± SEM.

FIG. 5.

Equilibrium coefficient of friction measured under steady frictional shear. Cellular constructs displayed significantly higher coefficients of friction against a rotating stainless steel platen than did acellular scaffolds (ANOVA, *p < 0.05, **p < 0.001). Data represented as mean ± SEM.

FIG. 6.

Histology of cellular 3D PCL constructs and cellular 3D PCL–fibrin composite constructs at days 14 and 28 depicts fibrocartilaginous tissue synthesis within the 3D PCL scaffold. Scale bar = 100 μm. Color images available online at www.liebertonline.com/ten.

FIG. 7.

Histology and immunohistochemistry of acellular 3D PCL–fibrin composite scaffolds, cellular 3D PCL constructs, and cellular 3D PCL–fibrin composite constructs at days 14 and 28. Scale bar = 1 mm. Color images available online at www.liebertonline.com/ten.