Evaluation of a novel oral mucosa in vitro implantation model for analysis of molecular interactions with dental abutment surfaces.

BACKGROUND: Abutment surfaces are being designed to promote gingival soft tissue attachment and integration. This forms a seal around prosthetics and consequently ensures long-term implant survival. New scalable and reproducible models are necessary to evaluate and quantify the performance of these surfaces. PURPOSE: To evaluate a novel implantation model by histomorphometric and immunohistochemical characterization of the interactions between human oral gingival tissue and titanium abutments with either novel anodized or conventional machined surface. MATERIALS AND METHODS: Abutments were inserted into an organotypic reconstructed human gingiva (RHG) model consisting of differentiated gingival epithelium cells on a fibroblast populated lamina propria hydrogel following a tissue punch. Epithelial attachment, down-growth along the abutment surface, and phenotype were assessed via histomorphology, scanning electron microscopy, and immunohistochemistry 10 days after implantation. RESULTS: The down-growing epithelium transitioned from a gingival margin to a sulcular and junctional epithelium. The sulcus depth and junctional epithelial length were similar to previously reported pre-clinical and clinical lengths. A collagen IV/laminin 5 basement membrane formed between the epithelium and the underlying connective tissue. The RHG expanded in thickness approximately 2-fold at the abutment surface. The model allowed the evaluation of protein expression of adhering soft tissue cells for both tested abutments. CONCLUSIONS: The RHG model is the first in vitro 3D model to enable the assessment of not only human epithelial tissue attachment to dental abutments but also the expression of protein markers involved in soft tissue attachment and integration. The two abutments showed no noticeable difference in epithelial attachment.

Biomaterial surface energy-driven ligand assembly strongly regulates stem cell mechanosensitivity and fate on very soft substrates.

Although mechanisms of cell-material interaction and cellular mechanotransduction are increasingly understood, the mechanical insensitivity of mesenchymal cells to certain soft amorphous biomaterial substrates has remained largely unexplained. We reveal that surface energy-driven supramolecular ligand assembly can regulate mesenchymal stem cell (MSC) sensing of substrate mechanical compliance and subsequent cell fate. Human MSCs were cultured on collagen-coated hydrophobic polydimethylsiloxane (PDMS) and hydrophilic polyethylene-oxide-PDMS (PEO-PDMS) of a range of stiffnesses. Although cell contractility was similarly diminished on soft substrates of both types, cell spreading and osteogenic differentiation occurred only on soft PDMS and not hydrophilic PEO-PDMS (elastic modulus <1 kPa). Substrate surface energy yields distinct ligand topologies with accordingly distinct profiles of recruited transmembrane cell receptors and related focal adhesion signaling. These differences did not differentially regulate Rho-associated kinase activity, but nonetheless regulated both cell spreading and downstream differentiation.

Surface-Driven Collagen Self-Assembly Affects Early Osteogenic Stem Cell Signaling.

This study reports how extracellular matrix (ECM) ligand self-assembly on biomaterial surfaces and the resulting nanoscale architecture can drive stem cell behavior. To isolate the biological effects of surface wettability on protein deposition, folding, and ligand activity, a polydimethylsiloxane (PDMS)-based platform was developed and characterized with the ability to tune wettability of elastomeric substrates with otherwise equivalent topology, ligand loading, and mechanical properties. Using this platform, markedly different assembly of covalently bound type I collagen monomers was observed depending on wettability, with hydrophobic substrates yielding a relatively rough layer of collagen aggregates compared to a smooth collagen layer on more hydrophilic substrates. Cellular and molecular investigations with human bone marrow stromal cells revealed higher osteogenic differentiation and upregulation of focal adhesion-related components on the resulting smooth collagen layer coated substrates. The initial collagen assembly driven by the PDMS surface directly affected α1ß1 integrin/discoidin domain receptor 1 signaling, activation of the extracellular signal-regulated kinase/mitogen activated protein kinase pathway, and ultimately markers of osteogenic stem cell differentiation. We demonstrate for the first time that surface-driven ligand assembly on material surfaces, even on materials with otherwise identical starting topographies and mechanical properties, can dominate the biomaterial surface-driven cell response.

Influence of physical properties of biomaterials on cellular behavior.

PURPOSE: In this study, we evaluated the effect of hydrogel structural properties on proliferation and biosynthesis activity of encapsulated chondrocytes. METHODS: Hydrogels with varying structural and mechanical properties were prepared by photopolymerizing PEGDA precursors having MWs of 3.4 kDa, 6 kDa, 10 kDa, and 20 kDa and were characterized for their swelling ratio, network structure, morphology, and mechanical properties. The effect of hydrogel structural properties on the cellular activity of encapsulated chondrocytes was studied over four weeks. RESULTS: Varying the molecular weight of PEGDA precursors exhibited a significant effect on the structural and mechanical properties of the hydrogels. Large mesh size was found to support cell proliferation. However, extracellular matrix (ECM) accumulation varied with the precursor molecular weight. Both PEGDA 6 kDa and 10 kDa hydrogels supported GAG accumulation, while PEGDA 10 kDa and 20 KDa hydrogels supported collagen accumulation. Chondrocytes cultured in PEGDA 10 kDa hydrogels expressed a relative increase in collagen type II and aggrecan expression while maintaining low collagen type I expression. CONCLUSIONS: Increasing mesh size of the hydrogels resulted in an increase in cellular proliferation exhibiting the strong correlation between mesh size and cell growth, while mesh size had a differential effect on ECM accumulation and expression of cartilage specific markers.