Publications

The present study investigated early interactions between three alloplastic materials (calcium phosphate (CaP), titanium alloy (Ti), and polyetheretherketone (PEEK) with human whole blood using an established in vitro slide chamber model. After 60 min of contact with blood, coagulation (thrombin-antithrombin complexes, TAT) was initiated on all test materials (Ti > PEEK > CaP), with a significant increase only for Ti. All materials showed increased contact activation, with the KK-AT complex significantly increasing for CaP (p < 0.001), Ti (p < 0.01), and PEEK (p < 0.01) while only CaP demonstrated a notable rise in KK-C1INH production (p < 0.01). The complement system had significant activation across all materials, with CaP (p < 0.0001, p < 0.0001) generating the most pronounced levels of C3a and sC5b-9, followed by Ti (p < 0.001, p < 0.001) and lastly, PEEK (p < 0.001, p < 0.01). This activation correlated with leukocyte stimulation, particularly myeloperoxidase release. Consequently, the complement system may assume a more significant role in the early stages post implantation in response to CaP materials than previously recognized. Activation of the complement system and the inevitable activation of leukocytes might provide a more favorable environment for tissue remodeling and repair than has been traditionally acknowledged. While these findings are limited to the early blood response, complement and leukocyte activation suggest improved healing outcomes, which may impact long-term clinical outcomes.

JTD Keywords: activation, biomaterial, calcium phosphate, coagulation, complement, cranioplasty, human whole blood, platelets, receptors, surface, titanium, Biomaterials, Calcium phosphate, Coagulation, Complement, Human whole blood, Peroxidase enzymes

Galleria mellonella is an alternative animal model of infection. The use of this species presents a wide range of advantages, as its maintenance and rearing are both easy and inexpensive. Moreover, its use is considered to be more ethically acceptable than other models, it is conveniently sized for manipulation, and its immune system has multiple similarities with mammalian immune systems. Hemocytes are immune cells that help encapsulate and eliminate pathogens and foreign particles. All of these reasons make this insect a promising animal model. However, cultivating G. mellonella hemocytes in vitro is not straightforward and it has many difficult challenges. Here, we present a methodologically optimized protocol to establish and maintain a G. mellonella hemocyte primary culture. These improvements open the door to easily and quickly study the toxicity of nanoparticles and the interactions of particles and materials in an in vitro environment.

JTD Keywords: cell culture, galleria mellonella, infection, nanoparticle, Bacteria, Cell culture, Galleria mellonella, Hemolin, Infection, Insect hemocytes, Larvae, Lepidoptera, Nanoparticle, Phagocytosis, Prophenoloxidase, Suspension, Systems

Electrochemical sensors for real-time detection of several bioanalytes have been prepared by additive manufacturing, shaping non-conductive poly(lactic acid) (PLA) filaments, and applying a physical treatment to create excited species. The latter process, which consists of the application of power discharge of 100 W during 2 min in a chamber at a low pressure of O-2, converts electrochemically inert PLA into an electrochemically responsive material. The electric discharge caused the oxidation of the PLA surface as evidenced by the increment in the quantity of oxygenated species detected by FTIR spectroscopy and X-ray photoelectron spectroscopy (XPS). Indeed, changes in the surface chemical composition became more pronounced with increasing O-2 pressure. After demonstrating the performance of the chemically modified material as individual dopamine and glucose sensors, multiplexed detection has been achieved by measuring simultaneously the two voltammetric signals. This has been performed by collecting the signals in two different regions, a naked chemically modified PLA for dopamine detection and a chemically modified PLA region functionalized with Glucose Oxidase. These outcomes led to define a new paradigm for manufacturing electrodes for electrochemical sensors based on 3D printing without using conducting materials at any stage of the process.

JTD Keywords: Additive manu f a c turing, Carbon, Conductivity, Degradation, Dopamine, Dopamine detection, Glucose detection, Glucose sensors, Immobilization, Multiplexed detect i o n, Oxidase, Plasma treatment

Despite tremendous progress in the understanding of COVID-19, mechanistic insight into immunological, disease-driving factors remains limited. We generated maVie16, a mouse-adapted SARS-CoV-2, by serial passaging of a human isolate. In silico modeling revealed how only three Spike mutations of maVie16 enhanced interaction with murine ACE2. maVie16 induced profound pathology in BALB/c and C57BL/6 mice, and the resulting mouse COVID-19 (mCOVID-19) replicated critical aspects of human disease, including early lymphopenia, pulmonary immune cell infiltration, pneumonia, and specific adaptive immunity. Inhibition of the proinflammatory cyto-kines IFN? and TNF substantially reduced immunopathology. Importantly, genetic ACE2-deficiency completely prevented mCOVID-19 development. Finally, inhalation therapy with recombinant ACE2 fully protected mice from mCOVID-19, revealing a novel and efficient treatment. Thus, we here present maVie16 as a new tool to model COVID-19 for the discovery of new therapies and show that disease severity is determined by cytokine-driven immunopathology and critically dependent on ACE2 in vivo. © Gawish et al.

JTD Keywords: covid-19 mouse model, covid-19 therapy, cytokine storm, immunology, inflammation, mavie16, mouse, mouse-adapted sars-cov-2, program, recombinant soluble ace2, tmprss2, Adaptive immunity, Angiotensin converting enzyme 2, Angiotensin-converting enzyme 2, Animal, Animal cell, Animal experiment, Animal model, Animal tissue, Animals, Apoptosis, Article, Bagg albino mouse, Breathing rate, Bronchoalveolar lavage fluid, C57bl mouse, Cell composition, Cell infiltration, Controlled study, Coronavirus disease 2019, Coronavirus spike glycoprotein, Covid-19, Cytokeratin 18, Cytokine production, Dipeptidyl carboxypeptidase, Disease model, Disease models, animal, Disease severity, Drosophila-melanogaster, Enzyme linked immunosorbent assay, Expression vector, Flow cytometry, Gamma interferon, Gene editing, Gene expression, Gene mutation, Genetic engineering, Genetics, Glycosylation, High mobility group b1 protein, Histology, Histopathology, Immune response, Immunocompetent cell, Immunology, Immunopathology, Interferon-gamma, Interleukin 2, Metabolism, Mice, inbred balb c, Mice, inbred c57bl, Mouse-adapted sars-cov-2, Myeloperoxidase, Neuropilin 1, Nonhuman, Nucleocapsid protein, Pathogenicity, Peptidyl-dipeptidase a, Pyroptosis, Recombinant soluble ace2, Renin angiotensin aldosterone system, Rna extraction, Rna isolation, Sars-cov-2, Severe acute respiratory syndrome coronavirus 2, Spike glycoprotein, coronavirus, T lymphocyte activation, Trabecular meshwork, Tumor necrosis factor, Virology, Virus load, Virus replication, Virus transmission, Virus virulence

A conducting nanocomposite hydrogel is developed for the detection of L-lactate. The hydrogel is based on a mixture of alginate (Alg) and poly(3,4-ethylenedioxythiophene) (PEDOT), which is loaded with gold nanoparticles (GNP). In this novel hydrogel, Alg provides 3D structural support and flexibility, PEDOT confers conductivity and sensing capacity, and GNP provides signal amplification with respect to simple voltammetric and chronoamperometric response. The synergistic combination of the properties provided by each component results in a new flexible nanocomposite with outstanding capacity to detect hydrogen peroxide, which has been used to detect the oxidation of L-lactate. The hydrogel detects hydrogen peroxide with linear response and limits of detection of 0.91 ?M and 0.02 ?M by cyclic voltammetry and chronoamperometry, respectively. The hydrogel is functionalized with lactate oxidase, which catalyzes the oxidation of L-lactate to pyruvate, forming hydrogen peroxide. For L-lactate detection, the functionalized biosensor works in two linear regimes, one for concentrations lower than 5 mM with a limit of detection of 0.4 mM, and the other for concentrations up to 100 mM with a limit of detection of 3.5 mM. Because of its linear range interval, the developed biosensor could be suitable for a wide number of biological fluids. © 2021

JTD Keywords: biosensor, dehydrogenase, enzymatic oxidation, films, hardness, indentation, lactate oxidase, Biosensor, Elastic-modulus, Enzymatic oxidation, Lactate, Lactate oxidase, Reacciones enzimáticas

Diamine oxydase and peroxidase have been co-immobilized onto layered double hydroxide (LDH) thin films for the development of real-time histamine biosensors. The chosen LDH materials are Mg2AlCO3, Mg4FeCl and Ca2AlCl. Prepared bi-enzymatic hybrid nanomaterials are capable of detecting histamine through the electrochemical oxidation of H(2)O(2)and are used as the sensitive membrane for potentiometric microelectrode. Histamine biosensors developed in this work have fast response of less than 20 s, are sensitive and selective, with a large dynamic range of 10(-8)-10(-3) M and a limit of detection of less than 10(-8) M. The detection limit of the developed bi-enzymatic biosensors is relatively higher than those corresponding with gas and liquid chromatography, which are still considered as the reference methods. Finally, the reproducibility, the specificity and the storage stability of the biosensors were studied.

JTD Keywords: Biogenic-amines, Biosensor, Diamine oxidase, Film, Fish, Histamine, Hybrid nanomaterial, Immobilization, Layer double hydroxide, Potentiometric biosensor, Specificity