RESUMEN
Strategies to combine aspirated marrow cells with scaffolds to treat connective tissue defects are gaining increasing clinical attention and use. In situations such as large defects where initial survival and proliferation of transplanted connective tissue progenitors (CTPs) are limiting, therapeutic outcomes might be improved by using the scaffold to deliver growth factors that promote the early stages of cell function in the graft. Signaling by the epidermal growth factor receptor (EGFR) plays a role in cell survival and has been implicated in bone development and homeostasis. Providing epidermal growth factor (EGF) in a scaffold-tethered format may sustain local delivery and shift EGFR signaling to pro-survival modes compared to soluble ligand. We therefore examined the effect of tethered EGF on osteogenic colony formation from human bone marrow aspirates in the context of three different adhesion environments using a total of 39 donors. We found that tethered EGF, but not soluble EGF, increased the numbers of colonies formed regardless of adhesion background, and that tethered EGF did not impair early stages of osteogenic differentiation.
Asunto(s)
Ensayo de Unidades Formadoras de Colonias , Células del Tejido Conectivo/citología , Células del Tejido Conectivo/efectos de los fármacos , Factor de Crecimiento Epidérmico/farmacología , Células Madre/citología , Células Madre/efectos de los fármacos , Adsorción/efectos de los fármacos , Adhesión Celular/efectos de los fármacos , Recuento de Células , Diferenciación Celular/efectos de los fármacos , Tamaño de la Célula/efectos de los fármacos , Fibronectinas/farmacología , Humanos , Osteogénesis/efectos de los fármacos , Péptidos/farmacología , Polímeros , Suero , Solubilidad/efectos de los fármacosRESUMEN
Bone graft performance can be enhanced by addition of connective tissue progenitors (CTPs) from fresh bone marrow in a manner that concentrates the CTP cell population within the graft. Here, we used small peptide adhesion ligands presented against an otherwise adhesion-resistant synthetic polymer background in order to illuminate the molecular basis for the attachment and colony formation by osteogenic CTPs from fresh human marrow, and contrast the behavior of fresh marrow to many commonly used osteogenic cell sources. The linear GRGDSPY ligand was as effective as tissue culture polystyrene in fostering attachment of culture-expanded porcine CTPs. Although this GRGDSPY peptide was more effective than control peptides in fostering alkaline phosphatase (AP)-positive colony formation from primary human marrow in 5 of the 7 patients tested, GRGDSPY was as effective as the control glass substrate in only one patient of 7. Thus, the peptide appears capable of enabling osteoblastic development from only a subpopulation of CTPs in marrow. The bone sialoprotein-derived peptide FHRRIKA was ineffective in fostering attachment of primary culture-expanded pig CTPs, although it was as effective as GRGDSPY in fostering AP-positive colonies from fresh human marrow. This study provides insights into integrin-mediated behaviors of CTPs and highlights differences between freshly isolated marrow and culture-expanded cells.
Asunto(s)
Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/fisiología , Oligopéptidos/farmacología , Osteoblastos/citología , Osteoblastos/fisiología , Osteogénesis/fisiología , Ingeniería de Tejidos/métodos , Células de la Médula Ósea/citología , Células de la Médula Ósea/fisiología , Moléculas de Adhesión Celular/química , Moléculas de Adhesión Celular/farmacología , Técnicas de Cultivo de Célula/métodos , Diferenciación Celular , Proliferación Celular , Células Cultivadas , Materiales Biocompatibles Revestidos/química , Materiales Biocompatibles Revestidos/farmacología , Humanos , Ensayo de Materiales , Oligopéptidos/químicaRESUMEN
MSC can act as a pluripotent source of reparative cells during injury and therefore have great potential in regenerative medicine and tissue engineering. However, the response of MSC to many growth factors and cytokines is unknown. Many envisioned applications of MSC, such as treating large defects in bone, involve in vivo implantation of MSC attached to a scaffold, a process that creates an acute inflammatory environment that may be hostile to MSC survival. Here, we investigated cellular responses of MSC on a biomaterial surface covalently modified with epidermal growth factor (EGF). We found that surface-tethered EGF promotes both cell spreading and survival more strongly than saturating concentrations of soluble EGF. By sustaining mitogen-activated protein kinase kinase-extracellular-regulated kinase signaling, tethered EGF increases the contact of MSC with an otherwise moderately adhesive synthetic polymer and confers resistance to cell death induced by the proinflammatory cytokine, Fas ligand. We concluded that tethered EGF may offer a protective advantage to MSC in vivo during acute inflammatory reactions to tissue engineering scaffolds. The tethered EGF-modified polymers described here could be used together with structural materials to construct MSC scaffolds for the treatment of hard-tissue lesions, such as large bony defects. Disclosure of potential conflicts of interest is found at the end of this article.