Printing three-dimensional tissue analogues with decellularized extracellular matrix bioink

The ability to print and pattern all the components that make up a tissue (cells and matrix materials) in three dimensions to generate structures similar to tissues is an exciting prospect of bioprinting. However, the majority of the matrix materials used so far for bioprinting cannot represent the complexity of natural extracellular matrix (ECM) and thus are unable to reconstitute the intrinsic cellular morphologies and functions. Here, we develop a method for the bioprinting of cell-laden constructs with novel decellularized extracellular matrix (dECM) bioink capable of providing an optimized microenvironment conducive to the growth of three-dimensional structured tissue. We show the versatility and flexibility of the developed bioprinting process using tissue-specific dECM bioinks, including adipose, cartilage and heart tissues, capable of providing crucial cues for cells engraftment, survival and long-term function. We achieve high cell viability and functionality of the printed dECM structures using our bioprinting method.open11349353sciescopu

Silica nanoparticles increase human adipose tissue-derived stem cell proliferation through ERK1/2 activation

Background: Silicon dioxide composites have been found to enhance the mechanical properties of scaffolds and to support growth of human adipose tissue-derived stem cells (hADSCs) both in vitro and in vivo. Silica (silicon dioxide alone) exists as differently sized particles when suspended in culture medium, but it is not clear whether particle size influences the beneficial effect of silicon dioxide on hADSCs. In this study, we examined the effect of different sized particles on growth and mitogen-activated protein kinase signaling in hADSCs. Methods: Silica gel was prepared by a chemical reaction using hydrochloric acid and sodium silicate, washed, sterilized, and suspended in serum-free culture medium for 48 hours, and then sequentially filtered through a 0.22 mu m filter (filtrate containing nanoparticles smaller than 220 nm; silica NPs). hADSCs were incubated with silica NPs or 3 mu m silica microparticles (MPs), examined by transmission electron microscopy, and assayed for cell proliferation, apoptosis, and mitogen-activated protein kinase signaling. Results: Eighty-nine percent of the silica NPs were around 50-120 nm in size. When hADSCs were treated with the study particles, silica NPs were observed in endocytosed vacuoles in the cytosol of hADSCs, but silica MPs showed no cell entry. Silica NPs increased the proliferation of hADSCs, but silica MPs had no significant effect in this regard. Instead, silica MPs induced slight apoptosis. Silica NPs increased phosphorylation of extracellular signal-related kinase (ERK) 1/2, while silica MPs increased phosphorylation of p38. Silica NPs had no effect on phosphorylation of Janus kinase or p38. Pretreatment with PD98059, a MEK inhibitor, prevented the ERK1/2 phosphorylation and proliferation induced by silica NPs. Conclusion: Scaffolds containing silicon dioxide for tissue engineering may enhance cell growth through ERK1/2 activation only when NPs around 50-120 nm in size are included, and single component silica-derived NPs could be useful for bioscaffolds in stem cell therapy.open112523sciescopu

A Very High-Order Accurate Staggered Finite Volume Scheme for the Stationary Incompressible Navier–Stokes and Euler Equations on Unstructured Meshes

International audienceWe propose a sixth-order staggered finite volume scheme based on polynomial reconstructions to achieve high accurate numerical solutions for the incompressible Navier-Stokes and Euler equations. The scheme is equipped with a fixed-point algorithm with solution relaxation to speed-up the convergence and reduce the computation time. Numerical tests are provided to assess the effectiveness of the method to achieve up to sixth-order con-2 Ricardo Costa et al. vergence rates. Simulations for the benchmark lid-driven cavity problem are also provided to highlight the benefit of the proposed high-order scheme

Bacillus anthracis Peptidoglycan Stimulates an Inflammatory Response in Monocytes through the p38 Mitogen-Activated Protein Kinase Pathway

We hypothesized that the peptidoglycan component of B. anthracis may play a critical role in morbidity and mortality associated with inhalation anthrax. To explore this issue, we purified the peptidoglycan component of the bacterial cell wall and studied the response of human peripheral blood cells. The purified B. anthracis peptidoglycan was free of non-covalently bound protein but contained a complex set of amino acids probably arising from the stem peptide. The peptidoglycan contained a polysaccharide that was removed by mild acid treatment, and the biological activity remained with the peptidoglycan and not the polysaccharide. The biological activity of the peptidoglycan was sensitive to lysozyme but not other hydrolytic enzymes, showing that the activity resides in the peptidoglycan component and not bacterial DNA, RNA or protein. B. anthracis peptidoglycan stimulated monocytes to produce primarily TNFα; neutrophils and lymphocytes did not respond. Peptidoglycan stimulated monocyte p38 mitogen-activated protein kinase and p38 activity was required for TNFα production by the cells. We conclude that peptidoglycan in B. anthracis is biologically active, that it stimulates a proinflammatory response in monocytes, and uses the p38 kinase signal transduction pathway to do so. Given the high bacterial burden in pulmonary anthrax, these findings suggest that the inflammatory events associated with peptidoglycan may play an important role in anthrax pathogenesis

Amyloid Plaques Beyond Aβ: A Survey of the Diverse Modulators of Amyloid Aggregation

Aggregation of the amyloid-β (Aβ) peptide is strongly correlated with Alzheimer’s disease (AD). Recent research has improved our understanding of the kinetics of amyloid fibril assembly and revealed new details regarding different stages in plaque formation. Presently, interest is turning toward studying this process in a holistic context, focusing on cellular components which interact with the Aβ peptide at various junctures during aggregation, from monomer to cross-β amyloid fibrils. However, even in isolation, a multitude of factors including protein purity, pH, salt content, and agitation affect Aβ fibril formation and deposition, often producing complicated and conflicting results. The failure of numerous inhibitors in clinical trials for AD suggests that a detailed examination of the complex interactions that occur during plaque formation, including binding of carbohydrates, lipids, nucleic acids, and metal ions, is important for understanding the diversity of manifestations of the disease. Unraveling how a variety of key macromolecular modulators interact with the Aβ peptide and change its aggregation properties may provide opportunities for developing therapies. Since no protein acts in isolation, the interplay of these diverse molecules may differentiate disease onset, progression, and severity, and thus are worth careful consideration

Prion protein-specific antibodies that detect multiple TSE agents with high sensitivity

This paper describes the generation, characterisation and potential applications of a panel of novel anti-prion protein monoclonal antibodies (mAbs). The mAbs were generated by immunising PRNP null mice, using a variety of regimes, with a truncated form of recombinant ovine prion protein spanning residues 94–233. Epitopes of specific antibodies were mapped using solid-phase Pepscan analysis and clustered to four distinct regions within the PrP molecule. We have demonstrated the utility of these antibodies by use of Western blotting and immunohistochemistry in tissues from a range of different species affected by transmissible spongiform encephalopathy (TSE). In comparative tests against extensively-used and widely-published, commercially available antibodies, similar or improved results can be obtained using these new mAbs, specifically in terms of sensitivity of detection. Since many of these antibodies recognise native PrPC, they could also be applied to a broad range of immunoassays such as flow cytometry, DELFIA analysis or immunoprecipitation. We are using these reagents to increase our understanding of TSE pathogenesis and for use in potential diagnostic screening assays

Improved functionalization of oleic acid-coated iron oxide nanoparticles for biomedical applications

Superparamagnetic iron oxide nanoparticles can providemultiple benefits for biomedical applications in aqueous environments such asmagnetic separation or magnetic resonance imaging. To increase the colloidal stability and allow subsequent reactions, the introduction of hydrophilic functional groups onto the particles’ surface is essential. During this process, the original coating is exchanged by preferably covalently bonded ligands such as trialkoxysilanes. The duration of the silane exchange reaction, which commonly takes more than 24 h, is an important drawback for this approach. In this paper, we present a novel method, which introduces ultrasonication as an energy source to dramatically accelerate this process, resulting in high-quality waterdispersible nanoparticles around 10 nmin size. To prove the generic character, different functional groups were introduced on the surface including polyethylene glycol chains, carboxylic acid, amine, and thiol groups. Their colloidal stability in various aqueous buffer solutions as well as human plasma and serum was investigated to allow implementation in biomedical and sensing applications.status: publishe

Development of three-dimensional alginate encapsulated chondrocyte hybrid scaffold using microstereolithography

Hydrogels are useful materials because of their chemical similarity to extracellular matrix and their ability to rapidly diffuse hydrophilic nutrients and metabolites. Using rapid prototyping methods, we fabricated free form three-dimensional (3D) scaffolds with chondrocytes encapsulated in an alginate hydrogel. The 3D hybrid scaffold was developed as combination of two components, a trimethylene carbonate (TMC)/trimethylolpropane (TMP) framework and an alginate hydrogel within an encapsulation of chondrocytes. To develop 3D hybrid scaffolds, we employed a microstereolithography system. The biodegradable, photopolymerizable liquid prepolymer was prepared by the polymerization of TMC with TMP and subsequently end capped with an acrylate group. The meshed framework of scaffolds withstood mechanical loading effectively. The line depth and linewidth could be controlled by varying laser power scan path, and scan speed. Results of cell culture indicate that the biomimetic nature of these encapsulated chondrocyte scaffolds effectively retain the phenotypic function of chondrocytes within the scaffold structure. The proposed 3D hybrid scaffolds can be used for cartilage regeneration.X1195sciescopu

Development of three-dimensional hybrid scaffold using chondrocyte-encapsulated alginate hydrogel

Hydrogels are useful materials because of their chemical similarity to the extracellular matrix and their ability to rapidly diffuse hydrophilic nutrients and metabolites. Using rapid prototyping (R-P) methods, we fabricated freeform three-dimensional (3-D) scaffolds with chondrocytes encapsulated in an alginate hydrogel. The 3-D hybrid scaffold was developed as a combination of two components, a trimethylene carbonate (TMC)/ trimethylolpropane (TMP) framework and an alginate hydrogel containing encapsulated chondrocytes. To develop 3-D hybrid scaffolds, we employed a microstereolithography (MSTL) system. A biocompatible, biodegradable, and photopolymerizable liquid prepolymer was prepared by the polymerization of TMC/ TMP, and was subsequently end-capped with an acrylate group. The results of analyzing a cell culture indicate that because of the biomimetic nature of the encapsulated chondrocytes, scaffolds effectively retain the phenotypic function of the chondrocytes within their structure. The proposed 3-D hybrid scaffolds can be used for cartilage regeneration.X116sciescopu

Development of a bone reconstruction technique using a solid free-form fabrication (SFF)-based drug releasing scaffold and adipose-derived stem cells

For tissue regeneration, three essential components of scaffolds, signals (biomolecules), and cells are required. Moreover, because bony defects are three-dimensional in many clinical circumstances, an exact 3D scaffold is important. Therefore, we proposed an effective reconstruction tool for cranial defects using human adipose-derived stem cells (hADSCs) and a 3D functional scaffold fabricated by solid free-form fabrication (SFF) technology that secretes biomolecules. We fabricated poly(propylene fumarate)-based 3D scaffolds with embedded microsphere-deliverable bone morphogenetic protein-2 (BMP-2) by microstereolithography. BMP-2-loaded SFF scaffolds with/without hADSCs (SFF/BMP/hADSCs scaffolds and SFF/BMP scaffolds, respectively) and BMP-2-unloaded SFF scaffolds (SFF scaffolds) were then implanted in rat crania, and in vivo bone formation was observed. Analyses of bone formation areas using micro-computed tomography (micro-CT) showed the superiority of SFF/BMP/hADSCs scaffolds. Hematoxylin and eosin stain, Masson's trichrome stain, and collagen type-I stain supported the results of the micro-CT scan. And human leukocyte antigen-ABC showed that seeded, differentiated hADSCs were well grown and changed to the bone tissue at the inside of the scaffold. Results showed that our combination of a functional 3D scaffold and hADSCs may be a useful tool for improving the reconstruction quality of severe bony defects in which thick bone is required. (c) 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2013.X111312sciescopu