Effect of micro-fiber modulus on mesenchymal stem cell morphology and function in model micro-fiber/hydrogel composites
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1
Virginia Tech, School of Biomedical Engineering and Sciences, United States
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2
Virginia Maryland College of Veterinary Medicine, Large Animal Clinical Sciences, United States
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3
Vanderbilt, Chemical and Biomolecular Engineering, United States
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4
Virginia Tech, Chemical Engineering, United States
Introduction: Ligament ruptures afflict thousands of Americans annually, often requiring surgical replacement with autologous or allogeneic grafts to restore knee joint mechanics. However, both slow healing and resulting abnormal biomechanics can accelerate joint degeneration. Tissue engineering holds promise in overcoming limitations of existing treatments through the fabrication of scaffolding materials that support cell infiltration, proliferation, and differentiation into an organized tissue. Electrospun scaffolds with different properties (e.g., topographical, mechanical, chemical, biological) can support mesenchymal stem cell (MSC) proliferation and guide deposition of an anisotropic extracellular matrix (ECM). Since the high fiber densities of electrospun meshes impede cell infiltration, we have developed composites consisting of sparse aligned micro-fibers within a collagen hydrogel. This system permits MSC migration and proliferation while providing anisotropic mechanical and topographic cues to guide MSC morphology and function. Based on a growing body of evidence that matrix stiffness affects stem cell fate, the goal of this study was to determine how the tensile modulus of electrospun micro-fibers influences MSC morphology and function.
Methods: Micro-fiber meshes with varying modulus were prepared by electrospinning mixtures of polycaprolactone (PCL) and polyurethane (PUR) onto a rotating mandrel. Next, sparse micro-fiber/hydrogel composites were assembled by the encapsulation of thin (~5μm) fiber layer within a collagen gel (Figure 1). MSCs were cultured for up to 14 days in composites and the cell morphology and spatial distribution analyzed by confocal microscopy. Cell number and mRNA expression of ligaments markers were measured by Picogreen and PCR.
Results and Discussion: Elastic moduli of 31, 15, and 5.6 MPa (Figure 2a) were obtained for 0.6-0.7 μm diameter fiber meshes comprised of PCL, PUR, and a 25/75 wt% PCL/PUR blend, respectively. MSCs – combined with collagen and cast atop the micro-fiber layer – oriented with the underlying fiber network (Figure 2c) but remained randomly oriented in the absence of fibers (Figure 2d). After 14 days, the 5.6 MPa composites, possessed elongated and oriented cells throughout the collagen bulk. In contrast, the 31 MPa composites, exhibited randomly oriented polygonal cells in the collagen bulk. PCR at day 14 indicated that the tendon/ligament transcription factor Scleraxis and the contractile protein α-smooth muscle actin (a marker of a regenerative phenotype) were elevated for cells in the 5.6 MPa composites compared to collagen controls, while collagen 1α1 was elevated in the 31 MPa (Figure 2b). Thus, MSCs sense the mechanical properties conferred by electrospun fibers, with the softer fibers leading more promising outcomes for ligament applications: higher levels of ligament markers and better cell alignment.
Keywords:
nanocomposite,
cell phenotype,
Tissue Regeneration,
instructive microenvironment
Conference:
10th World Biomaterials Congress, Montréal, Canada, 17 May - 22 May, 2016.
Presentation Type:
Poster
Topic:
Biomaterials in mesenchymal and hematopoietic stem cell biology
Citation:
Thayer
PS,
Dahlgren
LA,
Guelcher
SA and
Goldstein
A
(2016). Effect of micro-fiber modulus on mesenchymal stem cell morphology and function in model micro-fiber/hydrogel composites.
Front. Bioeng. Biotechnol.
Conference Abstract:
10th World Biomaterials Congress.
doi: 10.3389/conf.FBIOE.2016.01.00462
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Received:
27 Mar 2016;
Published Online:
30 Mar 2016.