ABSTRACT
The aim of this research is experimental investigation of the topography and evaluation of some parameters of artificial microterritories promoting osteogenic differentiation of stromal stem cells. A technique of short-term culturing of prenatal human lung stromal cells with fibroblastoid morphology on calcium phosphate substrates with known topography was used. Judging from secretory activity of the cell culture (osteocalcin, alkaline phosphatase), stromal stem cells directly interacting with calcium phosphate discs have advantage in manifestation of osteoblast-like functional activity in comparison with cells cultured on plastic. Rough surfaces of calcium phosphate discs stimulate the formation of spatial human fibroblastoid cell culture. The cells with positive reaction to acid phosphatase are located on spheroliths forming the relief of calcium phosphate coatings. The cells with positive reaction to alkaline phosphatase (marker of osteoblasts) populate hollows (niches) of the artificial surface. The niche for induction of osteogenic differentiation of human multipotent mesenchymal stem cells is apparently a structural and functional formation. It can be characterized by an index calculated as the ratio of the total area occupied by alkaline phosphatase-positive cells to the area of artificial surface occupied by one stained cell.
Subject(s)
Cell Culture Techniques/methods , Mesenchymal Stem Cells/metabolism , Osteogenesis/physiology , Stem Cell Niche , Stromal Cells/metabolism , Alkaline Phosphatase/metabolism , Calcium Phosphates , Cell Differentiation , Cells, Cultured , Humans , Lung/cytology , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/physiology , Multipotent Stem Cells , Osteoblasts/cytology , Osteoblasts/metabolism , Osteocalcin/metabolism , Pilot ProjectsABSTRACT
Thin calcium phosphate coatings were deposited on NiTi substrates (plates) by rf-magnetron sputtering. The release of nickel upon immersion in water or in saline solution (0.9% NaCl in water) was measured by atomic absorption spectroscopy (AAS) for 42 days. The coating was analyzed before and after immersion by X-ray powder diffraction (XRD), scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDX). After an initial burst during the first 7 days that was observed for all samples, the rate of nickel release decreased 0.4-0.5 ng cm(-2) d(-1) for a 0.5 mum-thick calcium phosphate coating (deposited at 290 W). This was much less than the release from uncoated NiTi (3.4-4.4 ng cm(-2) d(-1)). Notably, the nickel release rate was not significantly different in pure water and in aqueous saline solution.
Subject(s)
Calcium Phosphates/chemistry , Coated Materials, Biocompatible/pharmacokinetics , Electroplating/methods , Nickel/pharmacokinetics , Titanium/pharmacokinetics , Calcium Phosphates/pharmacology , Coated Materials, Biocompatible/chemistry , Electroplating/instrumentation , Materials Testing , Microscopy, Electron, Scanning , Nickel/chemistry , Particle Size , Sodium Chloride/metabolism , Sodium Chloride/pharmacology , Spectrometry, X-Ray Emission , Surface Properties/drug effects , Titanium/chemistry , Water/chemistry , Water/metabolism , Water/pharmacology , X-Ray DiffractionABSTRACT
Thin calcium-phosphate coatings with thickness less than 2.7 m were prepared by radio-frequency magnetron sputtering technique on the surfaces of pure titanium, titanium alloy Ti6A14V and stainless ASTM 316. Results of scanning electron microscopy showed that all coatings were dense and poreless and did not have any visible defects or microcracks. Rutherford backscattering (RBS) revealed a prepared coating consisting only of calcium 33.6 (1.6 at%, phosphorous 16.5 (1.5 at%, and oxygen 48.6 (1.2 at%. The concentration of each above-mentioned element through the coating was almost constant. The physicomechanical properties of the prepared coatings were investigated using a nanoindentation technique. The values of nano-hardness and Young's modulus calculated on the basis of the obtained data were 10 GPa and 113 GPa, respectively. These values were higher than that of non-coated substrates, except titanium alloy due to the sputtering mechanism. It was found that the coating with a thickness less than 1.6 ?m possessed more adhesion strength than coatings with greater value of thickness. However, we suggest that all coatings have great cohesive resistance that does not depend on the coating thickness.
Subject(s)
Calcium Phosphates/chemistry , Coated Materials, Biocompatible/chemistry , Biomedical Engineering/methods , Microscopy, Electron, Scanning , Surface PropertiesABSTRACT
Correlation analysis demonstrated the role of inorganic parameters of the surfaces of calcium phosphate materials in the regulation of osteogenic differentiation of mesenchymal precursors. The progenitor stromal cells were isolated from syngeneic bone marrow immobilized in vitro on calcium phosphate surfaces with different structure, phasic, and elemental composition. After 45 days of subcutaneous ectopic osteogenesis in BALB/c mice, the tissues grown on these matrixes were characterized histologically. It was found that adhesion of bone marrow cells is the initial stage determining their future proliferation (conduction) over the artificial surface and the area of formed tissue plate. The success of histogenesis depends on surface roughness. The optimal roughness class was 4-5 (Russian State Standards), which enables differentiation of progenitor stromal cells under the specific microenvironmental conditions into the connective and adipose tissue cells. Differentiation of the progenitor cells into the stromal cells producing the hemopoiesis-inducing microenvironment also takes place in the foci of active hemopoiesis. Induction of osteogenic potential of the stromal precursors (osteoinduction) is determined by the ratio between calcium and phosphate atoms in surface coatings. In our experimental system, osteogenic differentiation of stromal mechanocytes was blocked only at Ca/P<0.5.
