Molecular Indicators of Biomaterials Osteoinductivity - Cell Migration, BMP Production and Signalling Turns a Key

In this work we dissect the osteoinductive properties of selected bioactive materials obtained in a 3D form, and based on PLGA matrix and 2 types of gel-derived bioactive glasses (SBGs) of SiO2-CaO system, each with and without P2O5. The study is designed to avoid any osteogenic stimuli beyond the putative osteogenic bioactive glass compound of the studied materials. Previously we found that, when used as growth surfaces (i.e. material sheets), some of these materials were capable to support osteogenesis of bone marrow stromal cells (BMSC) without the need for any additional osteogenic cell treatment. In this work we explore further the role of BMP production and signaling upon BMSC culture on selected, SBG/PLGA 3D scaffolds as well as BMSC migration toward the condition media obtained from such cell-loaded materials. Our results show that BMP signaling of osteoprogenitor cells as well as their migration rate may present important indicators of materials osteoinductivity.

Multi-layered in vitro 3D-bone model via combination of osteogenic cell sheets with electrospun membrane interlayer

In this study, it was aimed to present an approach for the development of multi-layered tissue engineering constructs by using cell sheet engineering. Briefly, MC3T3-E1 mouse pre-osteoblast cells were cultured in temperature-responsive plates (Nunc Upcell®) in the presence of osteogenic medium and the resulting cell sheets were laminated with electrospun poly(L-lactic acid) (PLLA) membranes to obtain viable three-dimensional, thick constructs. The constructs prepared without PLLA membranes were used as control. The cell viability and death in the resulting structures were investigated by microscopic and colorimetric methods. The in vitro performance of the structures was discussed comparatively. Alkaline phosphatase (ALP) activity, collagen and sulfated glycosaminoglycan (sGAG) content values were calculated. The presented approach shows potential for engineering applications of complex tissues with at least two or more microenvironments such as osteochondral, corneal or vascular tissues.

Early Biological Response of a Ultra-Hydrophilic Implant Surface Activated by Salts and Dry Technology: In Vitro Study on MC3T3-E1 Osteogenic Cell Line

New implant surfaces created by sandblasting and acid etching enhance osseointegration processes. Surface energy seems to be an aspect of paramount importance in the first phase of healing, supporting protein adsorption and cell adhesion, proliferation, differentiation and bone mineralization. New methods were introduced to preserve over time and improve surface energy such as wet storage or bioactivation through salts coating created with dry technology. Purpose of this study was to evaluate the osteogenic response of pre-osteoblast lineage cells to dry bioactivated surface. MC3T3-E1 osteogenic cell line were cultured on ABT (SLA surface) and NANO (SLA surface with dry bioactive technology)., cell adhesion assay, proliferation assay and cell morphology were performed. Despite both surface treatments were able to support regular morphology; cell adhesion and proliferation were statistically improved ( p.value<0,05) on ABT disks. Regardless of the encouraging effects on surface energy of dry bioactivation technology with salts, this results suggest that further investigations are needed to evaluate the exact role of salts after solubilitation during the early stages of healing and the possible interactions that may hinder cells adhesion, proliferation, differentiation.

Synergistic Effect of Growth Factor Releasing Polymeric Nanoparticles and Ultrasound Stimulation on Osteogenic Differentiation

Mesenchymal stem cells (MSCs) have been extensively used in the tissue regeneration therapy. Ex vivo therapy with well-differentiated osteogenic cells is known as an efficient treatment for musculoskeletal diseases, including rheumatoid diseases. However, along with its high cost, the current therapy has limitations in terms of restoring bone regeneration procedures. An efficient process for the cell differentiation to obtain a large number of functionalized osteogenic cells is necessary. Therefore, it is strongly recommended to develop strategies to produce sufficient numbers of well-differentiated osteogenic cells from the MSCs. In general, differentiation media with growth factors have been used to facilitate cell differentiation. In the present study, the poly (lactic-co-glycolic acid) (PLGA) nanoparticles incorporating the growth factors were included in the media, resulting in releasing growth factors (dexamethasone and β-glycerophosphate) in the media in the controlled manner. Stable growth and early differentiation of osteogenic cells were achieved by the PLGA-based growth factor releasing system. Moreover, low intensity pulsed ultrasound was applied to this system to induce cell differentiation process. The results revealed that, as a biomarker at early stage of osteogenic cell differentiation, Lamin A/C nuclear protein was efficiently expressed in the cells growing in the presence of PLGA-based growth factor reservoirs and ultrasound. In conclusion, our results showed that the ultrasound stimulation combined with polymeric nanoparticles releasing growth factors could potentially induce osteogenic cell differentiation.

The molecular conformation of silk fibroin regulates osteogenic cell behavior by modulating the stability of the adsorbed protein-material interface

Silk fibroin (SF) can be used to construct various stiff material interfaces to support bone formation. An essential preparatory step is to partially transform SF molecules from random coils to β-sheets to render the material water insoluble. However, the influence of the SF conformation on osteogenic cell behavior at the material interface remains unknown. Herein, three stiff SF substrates were prepared by varying the β-sheet content (high, medium, and low). The substrates had a comparable chemical composition, surface topography, and wettability. When adsorbed fibronectin was used as a model cellular adhesive protein, the stability of the adsorbed protein-material interface, in terms of the surface stability of the SF substrates and the accompanying fibronectin detachment resistance, increased with the increasing β-sheet content of the SF substrates. Furthermore, (i) larger areas of cytoskeleton-associated focal adhesions, (ii) higher orders of cytoskeletal organization and (iii) more elongated cell spreading were observed for bone marrow-derived mesenchymal stromal cells (BMSCs) cultured on SF substrates with high vs. low β-sheet contents, along with enhanced nuclear translocation and activation of YAP/TAZ and RUNX2. Consequently, osteogenic differentiation of BMSCs was stimulated on high β-sheet substrates. These results indicated that the β-sheet content influences osteogenic differentiation of BMSCs on SF materials in vitro by modulating the stability of the adsorbed protein-material interface, which proceeds via protein-focal adhesion-cytoskeleton links and subsequent intracellular mechanotransduction. Our findings emphasize the role of the stability of the adsorbed protein-material interface in cellular mechanotransduction and the perception of stiff SF substrates with different β-sheet contents, which should not be overlooked when engineering stiff biomaterials.

Stimulating effect of whey protein hydrolysate on bone growth in MC3T3-E1 cells and a rat model

This study was conducted to investigate the effect of whey protein hydrolysate (WPH) on osteogenic cell differentiation and its growth-promoting effects in rats. Alkaline phosphatase (ALP) activity and calcium deposition...

Numerical Analysis of the Influence of Porosity and Pore Geometry on Functionality of Scaffolds Designated for Orthopedic Regenerative Medicine

Background: Scaffolds are vital for orthopedic regenerative medicine. Therefore, comprehensive studies evaluating their functionality with consideration of variable parameters are needed. The research aim was to evaluate pore geometry and scaffold porosity influence on first, cell culture efficiency in a perfusion bioreactor and second, osteogenic cell diffusion after its implantation. Methods: For the studies, five pore geometries were selected (triangular prism with a rounded and a flat profile, cube, octagonal prism, sphere) and seven porosities (up to 80%), on the basis of which 70 models were created for finite element analyses. First, scaffolds were placed inside a flow channel to estimate growth medium velocity and wall shear stress. Secondly, scaffolds were placed in a bone to evaluate osteogenic cell diffusion. Results: In terms of fluid minimal velocity (0.005 m/s) and maximal wall shear stress (100 mPa), only cubic and octagonal pores with 30% porosity and spherical pores with 20% porosity fulfilled the requirements. Spherical pores had the highest osteogenic cell diffusion efficiency for porosities up to 30%. For higher porosities, the octagonal prism’s pores gave the best results up to 80%, where no differences were noted. Conclusions: The data obtained allows for the appropriate selection of pore geometry and scaffold porosity for orthopedic regenerative medicine.