Fabrication and Mechanical Characterisation of Titanium Lattices with Graded Porosity

Electron Beam Melting (EBM) is an Additive Manufacturing technique which can be used to fabricate complex structures from alloys such as Ti6Al4V, for example for orthopaedic applications. Here we describe the use of EBM for the fabrication of a novel Ti6Al4V structure of a regular diamond lattice incorporating graded porosity, achieved via changes in the strut cross section thickness. Scanning Electron Microscopy and micro computed tomography analysis confirmed that generally EBM reproduced the CAD design of the lattice well, although at smaller strut sizes the fabricated lattice produced thicker struts than the model. Mechanical characterisation of the lattice in uniaxial compression showed that its behaviour under compression along the direction of gradation can be predicted to good accuracy with a simple rule of mixtures approach, knowing the properties and the behaviour of its constituent layers

Galectin-1 serum levels reflect tumor burden and adverse clinical features in classical Hodgkin lymphoma

Galectin-1 (Gal1) is a member of a highly conserved family of carbohydrate-binding proteins that modulates innate and adaptive immune responses and fosters tumor-immune escape. Hodgkin lymphoma (HL) Reed-Sternberg (RS) cells overexpress and secrete Gal1, which selectively kills Th1,Th17 and cytotoxic T cells and promotes the immunosuppressive Th2/Treg-predominant HL microenvironment. We developed a sandwich ELISA and assessed serum Gal1 levels in 315 newly diagnosed, previously untreated HL patients enrolled on 3 risk-adapted clinical trials. Serum Gal1 levels were significantly higher in HL patients than in normal controls (p < .0001). Gal1 serum levels also increased with Ann Arbor stage (p < .0001), areas of nodal involvement (p = .0001) and the International Prognostic Score (IPS) (2-7, p = .006). We conclude that Gal1 serum levels are significantly associated with tumor burden and additional adverse clinical characteristics in newly diagnosed HL Patients.Fil: Ouyang, Jing. Dana-Farber Cancer Institute. Department of Medical Oncology; Estados Unidos de América;Fil: Plütschow, Annette. German Hodgkin Study Group; Alemania;Fil: Von Strandmann, Elke Pogge. University Hospital of Cologne. Laboratory for Immunotherapy; Alemania;Fil: Reiners, Katrin S.. University Hospital of Cologne. Laboratory for Immunotherapy; Alemania;Fil: Ponader, Sabine. German Hodgkin Study Group; Alemania;Fil: Rabinovich, Gabriel Adrian. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental (i); Argentina;Fil: Neuberg, Donna. Dana-Farber Cancer Institute. Department of Biostatistics; Estados Unidos de América;Fil: Engert, Andreas. German Hodgkin Study Group; Alemania;Fil: Shipp, Margaret A.. Dana-Farber Cancer Institute. Department of Medical Oncology; Estados Unidos de América

In-vitro- und In-vivo-Untersuchungen zur Biofunktionalisierung von Knochenersatzmaterialien

Substitutes for large-scale bone defects remain a problem since autologous donor tissues are limited and allografts and xenografts involve the risk of trans-mission of pathogens. Mainly economic biocompatible solutions for the substitu-tion of the defective tissue are of increasing interest. A rising number of new material compositions and fabrication methods provide possible alternatives to conventional materials. It was the aim of this study to evaluate cellulose-based and titanium scaf-folds for bone replacement with novel surface coatings. Cellulose-based scaf-folds coated with pure sodium silicate gel, sodium silicate gels accumulated with different concentrations of the bisphosphonate pamidronate and strontium and directly coated with strontianite, pure Ti-6Al-4V surfaces with different surface structures and porosities produced by the Selective Electron Beam Melting (SEBM) process and titanium surfaces coated with oligoelectrolytes were evaluated for their suitability as matrices for the attachment, proliferation and differentiation of human fetal osteoblasts (hFOB 1.19 cells). Furthermore, the osseointegration of these materials was evaluated in vivo. By means of cell pro-liferation, vitality, osteoblastic gene expression and micrographs cellular proc-esses were observed. To evaluate the in vivo performance of the materials his-tological, histomorphometric and microradiographic analyses were evaluated. All surfaces turned out to be suitable matrices for the adhesion and elemen-tary osteoblastic processes. The bioactive silica-based surface coating influ-enced positively the cellular behaviour. An inhibitory or promoting effect could be shown for the applied drugs, which depends on the applied concentration. The coating with strontianite turned out to increase cell vitality. Smooth titanium surfaces turned out to be best suited for cell adhesion and following cellular processes. The oligoelectrolyte coating survived the steam sterilisation and had no effect on cell proliferation. All cellulose matrices showed bioactivity in the in vivo experiments. It could be shown that the silica-based coating represents a slow-release system which dissolves continuously over a period of 90 days. In vivo only few osteoblasts could penetrate the pore system of the cellulose knits. In contrast, the porous titanium scaffolds were filled up with newly formed bone tissue during the study time. Altogether, the materials tested in this study are very suitable scaffolds for bone regenerative applications.Der Ersatz großer Knochendefekte ist ein bisher noch ungelöstes Problem. Komplikationen bei der Gewinnung und die begrenzte Verfügbarkeit autologen Knochens sowie die Gefahr der Übertragung von Krankheitserregern beim Ein-satz allogener und xenogener Produkte fördern die Entwicklung immer neuer synthetischer Knochenersatzmaterialien. Kostengünstige biokompatible Materi-alien sind dabei von steigendem Interesse. Immer neue Variationen hinsichtlich Materialzusammensetzung und neueste Herstellungsverfahren bieten eine Fülle von möglichen Alternativen zu den bisher bekannten Werkstoffen. Ziel der vor-liegenden Arbeit war es, ausgewählte Knochenersatzmaterialien mit völlig neu-artigen Beschichtungssystemen auf ihre Toxizität auf Knochenzellen sowie auf ihre Eignung als Gerüste für die Adhäsion, Proliferation und Differenzierung von Osteoblasten (hFOB 1.19 Zellen) zu untersuchen sowie die biologische Körper-verträglichkeit in vivo zu studieren. Auf dreidimensionalen Cellulosegewirken sollten neuartige Beschichtungsmethoden auf Basis von bioaktivem Glas mit modernen Wirkstoffen wie Pamidronat und Strontium angewendet werden, um die Biokompatibilität des Materials zu verbessern und als Slow-Release-Systeme eine zeitlich gesteuerte Abgabe moderner Wachstumsfaktoren in den Defekt zu ermöglichen und auf diese Weise den Heilungsprozess noch zu opti-mieren. Bei dem zweiten untersuchten Werkstoff handelte es sich um kommer-ziell erhältliches TiAl6V4-Pulver, welches in einem neuartigen Fertigungsverfah-ren, dem Selektiven Elektronenstrahlschmelzen (SEBM), zu dreidimensionalen Strukturen gefertigt wurde. Dieses Verfahren erlaubt in weiten Bereichen frei einstellbare Oberflächentopographien und Porositäten. Zusätzlich sollte das In-vitro-Verhalten eines kürzlich zum ersten Mal erfolgreich durchgeführten Be-schichtungsverfahrens, der Abscheidung monomolekularer Oligoelektrolyt-schichten, auf den Titanoberflächen untersucht werden. Das Zellverhalten auf den unterschiedlichen Oberflächen wurde mittels Mes-sung der Zelladhäsion, -proliferation, -vitalität und -differenzierung sowie in mik-roskopischen Aufnahmen beurteilt. Die Osseointegration wurde mithilfe histolo-gischer und immunhistochemischer Methoden bzw. REM-Aufnahmen beurteilt. Sämtliche Oberflächen stellten sich als geeignete Gerüste für die Anheftung und darauf folgende zelluläre Vorgänge der Osteoblasten heraus. Durch die Bioglasbeschichtungen auf den Cellulosegewirken konnte positiv Einfluss auf das Zellverhalten genommen werden. Die eingebrachten Wirkstoffe Pamidronat und Strontium hatten in Abhängigkeit von der applizierten Konzentration hem-mende oder förderliche Effekte auf das Zellverhalten. Eine Direktfunktionalisie-rung der Cellulose mit Strontiumcarbonat stellte sich als förderlich für die Zellvi-talität heraus. Glatte Titanoberflächen hatten den günstigsten Effekt auf das Zellverhalten. Die Oligoelektrolytbeschichtungen stellten sich als nicht zytoto-xisch heraus. In vivo konnte der Cellulose selbst sowie den verwendeten Be-schichtungssystemen eine Bioaktivität nachgewiesen werden. Die Bioglasbe-schichtung stellte sich als Slow-Release-System heraus, welches über einen Zeitraum von über 90 Tagen kontinuierlich in den Defektbereich abgegeben wird. Das Porensystem der Cellulosegewirke konnte von Knochengewebe nur in sehr geringem Maße durchwachsen werden. Die porigen Titankonstrukte hingegen wurden weitgehend von Knochengewebe gefüllt und stellen damit geeignete Gerüste für die Knochenrekonstruktion dar

Dasatinib inhibits CXCR4 signaling in chronic lymphocytic leukaemia cells and impairs migration towards CXCL12

Chemokines and their ligands play a critical role in enabling chronic lymphocytic leukaemia (CLL) cells access to protective microenvironmental niches within tissues, ultimately resulting in chemoresistance and relapse: disruption of these signaling pathways has become a novel therapeutic approach in CLL. The tyrosine kinase inhibitor dasatinib inhibits migration of several cell lines from solid-organ tumours, but effects on CLL cells have not been reported. We studied the effect of clinically achievable concentrations of dasatinib on signaling induced by the chemokine CXCL12 through its' receptor CXCR4, which is highly expressed on CLL cells. Dasatinib pre-treatment inhibited Akt and ERK phosphorylation in CLL cells upon stimulation with CXCL12. Dasatinib also significantly diminished the rapid increase in actin polymerisation observed in CLL cells following CXCL12 stimulation. Moreover, the drug significantly inhibited chemotaxis in a transwell assay, and reduced the percentage of cells able to migrate beneath a CXCL12-expressing murine stromal cell line. Dasatinib also abrogated the anti-apoptotic effect of prolonged CXCL12 stimulation on cultured CLL cells. These data suggest that dasatinib, akin to other small molecule kinase inhibitors targeting the B-cell receptor signaling pathway, may redistribute CLL cells from protective tissue niches to the peripheral blood, and support the investigation of dasatinib in combination strategies

Protein surface mimetics: understanding how ruthenium tris(bipyridines) interact with proteins.

Protein surface mimetics achieve high affinity binding by exploiting a scaffold to project binding groups over a large area of solvent exposed protein surface to make multiple co-operative non-covalent interactions. Such recognition is a pre-requisite for competitive/ orthosteric inhibition of protein-protein interactions (PPIs). This paper describes biophysical and structural studies on ruthenium(II) tris(bipyridine) surface mimetics that recognize cytochrome (cyt) c and inhibit the cyt c/ cyt c peroxidase (CCP) PPI. Binding is electrostatically driven, with enhanced affinity achieved through enthalpic contributions thought to arise from the ability of the surface mimetics to make a greater number of non-covalent interactions with surface exposed basic residues on cyt c in comparison to CCP. High field natural abundance 1H-15N HSQC NMR experiments are consistent with surface mimetics binding to cyt c in similar manner to CCP. This provides a framework for understanding recognition of proteins by supramolecular receptors and informing the design of ligands superior to the protein partners upon which they are inspired

Precise redox-sensitive cleavage sites for improved bioactivity of siRNA lipopolyplexes

Lipo-oligomers have been proven as potent siRNA carriers based on stable electrostatic and hydrophobic complex formation and endosomal membrane destabilization. Although high stability of siRNA polyplexes is desirable in the extracellular space and cellular uptake, intracellular disassembly is important for the cytosolic release of siRNA and RNA-induced silencing complex formation. To improve the release, bioreducible sequence-defined lipo-oligomers were synthesized by solid-phase assisted synthesis using the disulfide building block Fmoc-succinoyl-cystamine for precise positioning of a disulfide unit between a lipophilic diacyl (bis-myristyl, bis-stearyl or bis-cholestanyl) domain and an ionizable oligocationic siRNA binding unit. Reducible siRNA polyplexes show higher gene silencing efficacy and lower cytotoxicity than their stable analogs, consistent with glutathione-triggered siRNA release and reduced lytic activity

The PI3-kinase delta inhibitor idelalisib (GS-1101) targets integrin-mediated adhesion of chronic lymphocytic leukemia (CLL) cell to endothelial and marrow stromal cells

CLL cell trafficking between blood and tissue compartments is an integral part of the disease process. Idelalisib, a phosphoinositide 3-kinase delta (PI3K\u3b4) inhibitor causes rapid lymph node shrinkage, along with an increase in lymphocytosis, prior to inducing objective responses in CLL patients. This characteristic activity presumably is due to CLL cell redistribution from tissues into the blood, but the underlying mechanisms are not fully understood. We therefore analyzed idelalisib effects on CLL cell adhesion to endothelial and bone marrow stromal cells (EC, BMSC). We found that idelalisib inhibited CLL cell adhesion to EC and BMSC under static and shear flow conditions. TNF\u3b1-induced VCAM-1 (CD106) expression in supporting layers increased CLL cell adhesion and accentuated the inhibitory effect of idelalisib. Co-culture with EC and BMSC also protected CLL from undergoing apoptosis, and this EC- and BMSC-mediated protection was antagonized by idelalisib. Furthermore, we demonstrate that CLL cell adhesion to EC and VLA-4 (CD49d) resulted in the phosphorylation of Akt, which was sensitive to inhibition by idelalisib. These findings demonstrate that idelalisib interferes with integrin-mediated CLL cell adhesion to EC and BMSC, providing a novel mechanism to explain idelalisib-induced redistribution of CLL cells from tissues into the blood

Multivalent glycoconjugates as vaccines and potential drug candidates

Pathogens adhere to the host cells during the first steps of infection through multivalent interactions which involve protein–glycan recognition. Multivalent interactions are also involved at different stages of immune response. Insights into these multivalent interactions generate a way to use suitable carbohydrate ligands that are attached to a basic scaffold consisting of e.g., dendrimer, polymer, nanoparticle, etc., with a suitable linker. Thus a multivalent architecture can be obtained with controllable spatial and topology parameters which can interfere with pathogen adhesion. Multivalent glycoconjugates bearing natural or unnatural carbohydrate antigen epitopes have also been used as carbohydrate based vaccines to stimulate an innate and adaptive immune response. Designing and synthesizing an efficient multivalent architecture with optimal ligand density and a suitable linker is a challenging task. This review presents a concise report on the endeavors to potentially use multi- and polyvalent glycoconjugates as vaccines as well as anti-infectious and anti-inflammatory drug candidates