Osteogenic differentiation of rat bone mesenchymal stem cells cultured on poly (hydroxybutyrate-co-hydroxyvalerate), poly (ε-caprolactone) scaffolds.

Bioresorbable biomaterials can fill bone defects and act as temporary scaffold to recruit MSCs to stimulate their differentiation. Among the different bioresorbable polymers studied, this work focuses on poly(hydroxybutyrate-co-hydroxyvalerate) (PHBV) and poly(ε-caprolactone) (PCL). Were prepared blends of PHBV and PCL to obtain PHBV based biomaterials with good tenacity, important for bone tissue repair, associated with biocompatible properties of PCL. This study assesses the viability of Vero cells on scaffolds of PHBV, PCL, and their blends and the osteogenic differentiation of mesenchymal stem cells (MSCs). Materials were characterized in surface morphology, DSC and Impact Strength (IS). Vero cells and MSCs were assessed by MTT assay, cytochemical and SEM analysis. MSC osteogenic differentiation was evaluated through alizarin red staining and ALP activity. We found some roughness onto surface materials. DSC showed that the blends presented two distinct melting peaks, characteristic of immiscible blends. IS test confirmed that PHBV-PCL blends is an alternative for increase the tenacity of PHBV. MTT assay showed cells with high metabolic activities on extract toxicity test, but with low activity in the direct contact test. SEM analysis showed spreading cells with irregular and flattened morphology on different substrates. Cytochemical study revealed that MSCs maintained their morphology, although in smaller number for MSCs. The development of nodules of mineralized organic matrix in MSC cultures was identified by alizarin red staining and osteogenic differentiation was confirmed by the quantification of ALP activity. Thus, our scaffolds did not interfere on viability of Vero cells or the osteogenic differentiation of MSCs.

Cytotoxicity Assessment of a Poly(methyl methacrylate) Synthesized for the Direct Fabrication of Bone Tissues

ABSTRACT A cytotoxicity study is performed on a poly(methyl methacrylate) polymer (PMMA) to be used for the fabrication of bone tissue by Rapid Prototyping (RP). The solution polymerization is conducted in a pilot plant reactor using more appropriated reagents in consideration of the medical application. Moreover, the polymer is efficiently handled to avoid the side effect of the monomer, reducing the concentration of this specie to 287,731 µg MMA/kg PMMA. The cytotoxicity of the polymer is determined through growth monitoring, adherence and morphology of L-929 cells. Additionally, MTT and LIVE/DEAD tests are performed. The results showed continuous and progressive growth of the cells on the surface of the specimens. Moreover, the material did not influence on the viability of mesenchymal cells and inverted fluorescence microscopy images showed a polyanionic dye calcein well retained in the cells in contact with the PMMA as well as the negative control after 72 hours. Thus, the polymer was efficiently synthesized and handled for the expected demands.

Fibrin network architectures in pure platelet-rich plasma as characterized by fiber radius and correlated with clotting time.

Fibrin networks are obtained through activation of platelet-rich plasma (PRP) for use in tissue regeneration. The importance of fibrin networks relies on mediation of release of growth factors, proliferation of tissue cells and rheological properties of the fibrin gels. Activation of PRP usually involves the decomposition of fibrinogen by agonists, in a wide range of concentrations. Therefore fibrin networks with a large structural diversity are formed, making comparative evaluations difficult. In order to standardize the fibrin networks, we used the statistical techniques central composite rotatable design and response-surface analysis, to correlate the radius of the fibers with the ratios between the agonists (autologous serum/calcium chloride) and agonist/PRP. From an individual and interactive analysis of the variables, architectures characterized by thick, medium and thin fibers were delineated on the response-surface. Furthermore, the architectures were correlated with coagulation time. This approach is valuable for standardizing the PRP preparation for clinical applications.