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Nature-inspired topographies on hydroxyapatite surfaces regulate stem cells behaviour.

ABSTRACT: Surface topography is one of the key factors in regulating interactions between materials and cells. While topographies presented to cells in vivo are non-symmetrical and in complex shapes, current fabrication techniques are limited to replicate these complex geometries. In this study, we developed a microcasting technique and successfully produced imprinted hydroxyapatite (HAp) surfaces with nature-inspired (honeycomb, pillars, and isolated islands) topographies. The in vitro biological performance of the developed non-symmetrical topographies was evaluated using adipose-derived stem cells (ADSCs). We demonstrated that ADSCs cultured on all HAp surfaces, except honeycomb patterns, presented well-defined stress fibers and expressed focal adhesion protein (paxillin) molecules. Isolated islands topographies significantly promoted osteogenic differentiation of ADSCs with increased alkaline phosphatase activity and upregulation of key osteogenic markers, compared to the other topographies and the control unmodified (flat) HAp surface. In contrast, honeycomb topographies hampered the ability of the ADSCs to proliferate and differentiate to the osteogenic lineage. This work presents a facile technique to imprint nature-derived topographies on the surface of bioceramics which opens up opportunities for the development of bioresponsive interfaces in tissue engineering and regenerative medicine.

SUBMITTER: Ramaswamy Y

PROVIDER: S-EPMC7569262 | biostudies-literature | 2021 Apr

REPOSITORIES: biostudies-literature

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Nature-inspired topographies on hydroxyapatite surfaces regulate stem cells behaviour.

Ramaswamy Yogambha Y Roohani Iman I No Young Jung YJ Madafiglio Genevieve G Chang Frank F Zhang Furong F Lu Zufu Z Zreiqat Hala H

Bioactive materials 20201016 4

Surface topography is one of the key factors in regulating interactions between materials and cells. While topographies presented to cells <i>in vivo</i> are non-symmetrical and in complex shapes, current fabrication techniques are limited to replicate these complex geometries. In this study, we developed a microcasting technique and successfully produced imprinted hydroxyapatite (HAp) surfaces with nature-inspired (honeycomb, pillars, and isolated islands) topographies. The <i>in vitro</i> biol ...[more]

PMID: 33102949

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Project description:Human epidermal stem cells initiate terminal differentiation when spreading is restricted on ECM-coated micropatterned islands, soft hydrogels or hydrogel-nanoparticle composites with high nanoparticle spacing. The effect of substrate topography, however, is incompletely understood. To explore this, primary human keratinocytes enriched for stem cells were seeded on a topographical library with over 2000 different topographies in the micrometre range. Twenty-four hours later the proportion of cells expressing the differentiation marker transglutaminase-1 was determined by high content imaging. As predicted, topographies that prevented spreading promoted differentiation. However, we also identified topographies that supported differentiation of highly spread cells. Topographies supporting differentiation of spread cells were more irregular than those supporting differentiation of round cells. Low topography coverage promoted differentiation of spread cells, whereas high coverage promoted differentiation of round cells. Based on these observations we fabricated a topography in 6-well plate format that supported differentiation of spread cells, enabling us to examine cell responses at higher resolution. We found that differentiated spread cells did not assemble significant numbers of hemidesmosomes, focal adhesions, adherens junctions, desmosomes or tight junctions. They did, however, organise the actin cytoskeleton in response to the topographies. Rho kinase inhibition and blebbistatin treatment blocked the differentiation of spread cells, whereas SRF inhibition did not. These observations suggest a potential role for actin polymerization and actomyosin contraction in the topography-induced differentiation of spread cells. STATEMENT OF SIGNIFICANCE: The epidermis is the outer covering of the skin. It is formed by layers of cells called keratinocytes. The basal cell layer contains stem cells, which divide to replace cells in the outermost layers that are lost through a process known as differentiation. In this manuscript we have developed surfaces that promote the differentiation of epidermal stem cells in order to understand the signals that control differentiation. The experimental tools we have developed have the potential to help us to devise new treatments that control diseases such as psoriasis and eczema in which epidermal stem cell proliferation and differentiation are disturbed.

Project description:Hydroxyapatite (HA), the principal component of bone mineral, shows osteoconductive properties when employed for coating metal implants as well as scaffold materials in synthetic bone grafts. With the goal of providing this material with osteoinductive capabilities to promote faster bone regeneration, we show an easy approach to functionalize HA implant surfaces and enrich them with osteoinductive properties by the use of HA-binding modular peptides. The modular peptides are designed as a combination of two domains, an HA-binding peptide motif and an osteogenic peptide motif derived from the osteogenic growth peptide (OGP) or bone morphometric protein 7 (BMP-7). To identify the best HA-binding peptide, several nature-inspired peptides derived from natural bone extracellular matrix proteins (bone sialoprotein, osteonectin, osteocalcin, and salivarin statherin) were compared for HA-binding activity, revealing concentration-dependent and incubation-time-dependent behaviours. We discovered that a Poly-E heptamer (E7) is the best HA-binding peptide, and thus combined it with a second osteogenic peptidic domain to create an osteoinductive modular peptide. After binding/release characterization, we found that the addition of the second osteogenic peptide domain did not change the binding profile of the modular peptides and caused only a slight change in their release kinetics. Mesenchymal stem cells (MSCs) were cultured on the HA substrates functionalized with modular peptides, and cell adhesion, proliferation, and differentiation in a basal medium (i.e., without any osteogenic supplements) were investigated. Gene expression data clearly showed that MSCs were committed to differentiate into osteoblasts in the presence of the modular peptides. HA discs functionalized with the E7 BMP-7 modular peptide showed the best capability in inducing the osteogenic differentiation of MSCs among all modular peptides studied. The modular peptides can easily be used to functionalize the HA implants through its constituent HA-binding motif, leaving the osteogenic peptide motif protruding from the surface for inducing osteogenesis. Our work opens up a new approach to the formulation of new bioactive HA coatings and implants for bone and dental repair.

Dataset Information

Nature-inspired topographies on hydroxyapatite surfaces regulate stem cells behaviour.

Publications

Nature-inspired topographies on hydroxyapatite surfaces regulate stem cells behaviour.

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