Lysozyme Release Research Articles

Enhanced Antibacterial Activity of CuS-BSA/Lysozyme under Near Infrared Light Irradiation.

The synthesis of multifunctional photothermal nanoagents for antibiotic loading and release remains a challenging task in nanomedicine. Herein, we investigated a simple, low-cost strategy for the preparation of CuS-BSA nanoparticles (NPs) loaded with a natural enzyme, lysozyme, as an antibacterial drug model under physiological conditions. The successful development of CuS-BSA NPs was confirmed by various characterization tools such as transmission electron microscopy (TEM), X-ray diffraction (XRD), Raman spectroscopy, and X-ray photoelectron spectroscopy (XPS). Lysozyme loading onto CuS-BSA NPs was evaluated by UV/vis absorption spectroscopy, Fourier-transform infrared spectroscopy (FTIR), zeta potential, and dynamic light scattering measurements. The CuS-BSA/lysozyme nanocomposite was investigated as an effective means for bacterial elimination of B. subtilis (Gram-positive) and E. coli (Gram-negative), owing to the combined photothermal heating performance of CuS-BSA and lysozyme release under 980 nm (0.7 W cm−2) illumination, which enhances the antibiotic action of the enzyme. Besides the photothermal properties, CuS-BSA/lysozyme nanocomposite possesses photodynamic activity induced by NIR illumination, which further improves its bacterial killing efficiency. The biocompatibility of CuS-BSA and CuS-BSA/Lysozyme was elicited in vitro on HeLa and U-87 MG cancer cell lines, and immortalized human hepatocyte (IHH) cell line. Considering these advantages, CuS-BSA NPs can be used as a suitable drug carrier and hold promise to overcome the limitations of traditional antibiotic therapy.

Synergistic Lysozyme‐Photodynamic Therapy Against Resistant Bacteria based on an Intelligent Upconversion Nanoplatform

AbstractThe rapid emergence of drug‐resistant bacteria has raised a great social concern together with the impetus for exploring advanced antibacterial ways. NIR‐triggered antimicrobial photodynamic therapy (PDT) by lanthanide‐doped upconversion nanoparticles (UCNP) as energy donor exhibits the advantages of high tissue penetration, broad antibacterial spectrum and less acquired resistance, but is still limited by its low efficacy. Now we designed a bio‐inorganic nanohybrid and combined lysozyme (LYZ) with UCNP‐PDT system to enhance the efficiency against resistant bacteria. Benefiting from the rapid adhesion to bacteria, intelligently bacteria‐responsive LYZ release and synergistic LYZ‐PDT effect, the nanoplatform achieves an exceptionally strong bactericidal capacity and conspicuous bacteriostasis on methicillin‐resistant S. aureus. These findings pave the way for designing efficiently antibacterial nanomaterials and provide a new strategy for combating deep‐tissue bacterial infection.

Synergistic Lysozyme-Photodynamic Therapy Against Resistant Bacteria based on an Intelligent Upconversion Nanoplatform.

The rapid emergence of drug-resistant bacteria has raised a great social concern together with the impetus for exploring advanced antibacterial ways. NIR-triggered antimicrobial photodynamic therapy (PDT) by lanthanide-doped upconversion nanoparticles (UCNP) as energy donor exhibits the advantages of high tissue penetration, broad antibacterial spectrum and less acquired resistance, but is still limited by its low efficacy. Now we designed a bio-inorganic nanohybrid and combined lysozyme (LYZ) with UCNP-PDT system to enhance the efficiency against resistant bacteria. Benefiting from the rapid adhesion to bacteria, intelligently bacteria-responsive LYZ release and synergistic LYZ-PDT effect, the nanoplatform achieves an exceptionally strong bactericidal capacity and conspicuous bacteriostasis on methicillin-resistant S. aureus. These findings pave the way for designing efficiently antibacterial nanomaterials and provide a new strategy for combating deep-tissue bacterial infection.

Efficacy of biodegradable, antimicrobial packaging on safety and quality parameters maintenance of a pear juice and rice milk-based smoothie product

In this study, the effect of a Polylactic acid (PLA) antimicrobial biodegradable packaging activated with lysozyme by cold plasma on a pear juice and rice milk-based smoothie was investigated. The antimicrobial effect of the active innovative packaging was evaluated In-vitro and on the smoothie inoculated with Listeria monocytogenens and Lactobacillus plantarum. After a preliminary evaluation of the lysozyme release kinetics in different conditions, its influence on some smoothie quality parameters (water activity, pH, colour and microbial growth) was evaluated. In-vitro trials showed an antimicrobial activity of the activated film against different microorganisms. Inoculated smoothies packed in activated and not materials were stored at 10 and 4 °C and analysed overtime. Results showed the capability of the activated package to inhibit Listeria monocytogenes and to maintain a better and a more stable colour compared to control ones. Activated pouches showed the best antimicrobial effect on samples stored at 10 °C compared to 4 °C, difference due to the faster lysozyme release kinetic from the packaging material at the highest storage temperature.Obtained results highlight the potentiality of the biodegradable packaging activated with lysozyme to be applied successfully in food industry, to improve safety and extend shelf-life of juice-based product.

Functional and Morphological Changes Induced in Mytilus Hemocytes by Selected Nanoparticles.

Nanoparticles (NPs) show various properties depending on their composition, size, and surface coating, which shape their interactions with biological systems. In particular, NPs have been shown to interact with immune cells, that represent a sensitive surveillance system of external and internal stimuli. In this light, in vitro models represent useful tools for investigating nano-bio-interactions in immune cells of different organisms, including invertebrates. In this work, the effects of selected types of NPs with different core composition, size and functionalization (custom-made PVP-AuNP and commercial nanopolystyrenes PS-NH2 and PS-COOH) were investigated in the hemocytes of the marine bivalve Mytilus galloprovincialis. The role of exposure medium was evaluated using either artificial seawater (ASW) or hemolymph serum (HS). Hemocyte morphology was investigated by scanning electron microscopy (SEM) and different functional parameters (lysosomal membrane stability, phagocytosis, and lysozyme release) were evaluated. The results show distinct morphological and functional changes induced in mussel hemocytes depending on the NP type and exposure medium. Mussel hemocytes may represent a powerful alternative in vitro model for a rapid pre-screening strategy for NPs, whose utilization will contribute to the understanding of the possible impact of environmental exposure to NPs in marine invertebrates.

Structural characterization and developability assessment of sustained release hydrogels for rapid implementation during preclinical studies

Sustained-release formulations are important tools to convert efficacious molecules into therapeutic products. Hydrogels enable the rapid assessment of sustained-release strategies, which are important during preclinical development where drug quantities are limited and fast turnaround times are the norm. Most research in hydrogel-based drug delivery has focused around synthesizing new materials and polymers, with limited focus on structural characterization, technology developability and implementation. Two commercially available thermosensitive hydrogel systems, comprised of block copolymers of poly(lactic-co-glycolic acid)-b-poly(ethylene glycol)-b-poly(lactic-co-glycolic acid) (PLGA) and poly(lactide-co-caprolactone)-b-poly(ethyleneglycol)-b-poly(lactide-co-caprolactone) (PLCL), were evaluated during this study. The two block copolymers described in the study were successfully formulated to form hydrogels which delayed the release of lysozyme (> 20 days) in vitro. Characterization of formulation attributes of the hydrogels like Tsol-gel temperature, complex viscosity and injection force showed that these systems are amenable to rapid implementation in preclinical studies. Understanding the structure of the gel network is critical to determine the factors controlling the release of therapeutics out of these gels. The structures were characterized via the gel mesh sizes, which were estimated using two orthogonal techniques: small angle X-ray scattering (SAXS) and rheology. The mesh sizes of these hydrogels were larger than the hydrodynamic radius (size) of lysozyme (drug), indicating that release through these gels is expected to be diffusive at all time scales rather than sub-diffusive. In vitro drug release experiments confirm that diffusion is the dominating mechanism for lysozyme release; with no contribution from degradation, erosion, relaxation, swelling of the polymer network or drug-polymer interactions. PLGA hydrogel was found to have a much higher complex viscosity than PLCL hydrogel, which correlates with the slower diffusivity and release of lysozyme seen from the PLGA hydrogel as compared to PLCL hydrogel. This is due to the increased frictional drag experienced by the lysozyme molecule in the PLGA hydrogel network, as described by the hydrodynamic theory.

Prevention of Pulmonary Injury in Isolated Perfused Rat Lungs by Activated Human Neutrophils Preincubated With Anti-Mo1 Monoclonal Antibody

Neutrophil activation results in neutrophil adherence and may subsequently cause lung injury through the generation of oxidants, release of granule proteases, and generation of a variety of mediator substances. We hypothesized that inhibition of neutrophil adherence and subsequent lung sequestration would attenuate the lung injury caused by activated neutrophils. Using isolated perfused rat lungs, we determined if anti-Mo1 monoclonal antibody (binds to the alpha subunit of a neutrophil glycoprotein [gp 155.94] that facilitates adherence) would attenuate lung neutrophil sequestration and lung injury caused by human neutrophils stimulated by phorbol myristate acetate (PMA). PMA-stimulated neutrophils but not PMA or neutrophils alone caused lung injury as assessed by accumulation of 125I-bovine serum albumin into lung parenchyma and alveolar lavage fluid. Incubation of neutrophils with anti-Mo1 antibody prior to stimulation with PMA attenuated lung injury and neutrophil sequestration. Furthermore, a histological survey revealed that anti-Mo1 antibody inhibited neutrophils present in the lung from spreading following exposure to PMA. Anti-Mo1 antibody did not inhibit PMA-stimulated neutrophil release of granule constituents or toxic O2 metabolites as evidenced by lysozyme and lactoferrin release or the reduction of ferricytochrome c in the lung perfusate. The inhibition of lung injury caused by the anti-Mo1 antibody was not likely due to a nonspecific effect of the antibody, since another murine monoclonal antibody of the same class (anti-Mo5) did not inhibit lung neutrophil sequestration or lung injury. Thus, in this experimental model, interference with the close approximation of the neutrophil to its target site inhibited the ability of the activated human neutrophil to cause injury.

Polystyrene-block-poly(ethylene oxide) Thin Films Fabricated from a Solvent Mixture for the Co-Assembly of Polymers and Proteins.

The co-assembly of peptides and proteins in poly(styrene-block-ethylene oxide) (PS-b-PEO) thin films has proven to be a promising method to fabricate polymer–biomolecule functional materials. Contrary to the covalent immobilization of biomolecules on surfaces, co-assembly presents the opportunity to arrange cargo within thin films, which can be released upon exposure to an aqueous environment. The use of a mixed solvent system ensures the solubilization of hydrophobic polymer as well as the solubilization and protection of the biomolecule cargo. However, to produce largely defect-free films of PS-b-PEO from a solvent mixture containing water is challenging due to the narrow range of solvent miscibility and polymer/protein solubility. This work explores the limits of using a benzene/methanol/water solvent mixture for the production of thin PS-b-PEO films and provides a template for the fabrication optimization of block copolymer thin films in different complex solvent systems. The film quality is analyzed using optical microscopy and atomic force microscopy and correlated to the solvent composition. By adjusting the solvent composition to 80/18.8/1.2 vol % benzene/methanol/water, it was possible to reliably fabricate thin films with less than 1% macroscopic defect surface coverage. Using the optimized solvent composition, we also demonstrate the fabrication of ordered PS-b-PEO films containing lysozyme. Furthermore, we show the release of lysozyme into aqueous media, which highlights the potential use of such films for drug delivery applications.

Production of Lysozyme-PLGA-Loaded Microparticles for Controlled Release Using Hot-Melt Extrusion.

Biopharmaceuticals are usually administered intravenously with frequent dosing regimens which may decrease patient compliance. Controlled-release formulations allow to reduce the frequency of injections while providing a constant dosing of the biopharmaceutical over extended periods. These formulations are typically produced by emulsions, requiring high amounts of organic solvents and have limited productivity. Hot-melt extrusion (HME) is an alternative technology to produce controlled drug delivery systems. It is a continuous solvent-free process, leading to a small ecological footprint and higher productivity. However, it may induce thermolabile compounds' degradation. In this work, the impact of the formulation and extrusion temperature on lysozyme's bioactivity and release profile of poly(lactic-co-glycolic acid) (PLGA)-based extended release formulations were evaluated using a design-of-experiments (DoE) approach. The lysozyme-loaded PLGA microparticles were produced by HME followed by milling. It was observed that the in vitro release (IVR) profile was mainly affected by the drug load; higher drug load led to higher burst and total lysozyme release after 14days. HME temperature seemed to decrease lysozyme's activity although this correlation was not statistically significant (p value = 0.0490). Adding polyethylene glycol 400 (PEG 400) as a plasticizer to the formulation had no significant impact on the lysozyme release profile. The burst release was effectively mitigated with the inclusion of a washing step. Washing the microparticles with water reduced the burst release by 80% whereas washing them with a poly(vinyl alcohol) (PVA) aqueous solution eliminated it. In conclusion, HME is demonstrated to be suitable in producing controlled-release microparticles of small biopharmaceuticals. Graphical abstract.

Intelligent magnetic imprinted nanoparticles for efficient lysozyme separation from egg whites and sustained antibacterial activity

Intelligent magnetic materials are capable of responding to environmental stimuli in an on/off manner. The photo- and thermal-responsive nanogel layer was surface-grafted onto several magnetic nanoparticles to prepare the magnetic imprinted nanoparticles (Fe3O4 MIPs). The Fe3O4 MIPs show dramatic temperature and light, dual-dependent rebinding and release characteristics for lysozyme. Above the volume phase transition temperature of the PNIPAAm, the imprinted protein (lysozyme) can be selectively separated from egg whites and the activity remained unchanged. By combining low temperature and UV irradiation, a more finely tuned release of the binding protein can be achieved. Time-dependent antibacterial activity was observed, and it was found that Fe3O4 MIPs-loaded lysozyme almost completely suppressed the growth of E. coli with the continuous lysozyme release. More importantly, the Fe3O4 MIPs could easily be separated by an external magnetic field and can be recycled. The photo- and thermal-responsive Fe3O4 MIPs pave a new way for the development of an advanced, sustainable material for use in bioseparation, drug delivery, and antibacterial materials.

Development of gel-based pads loaded with lysozyme and green tea extract: Characterization of pads and test of their antilisterial potential on cold-smoked salmon

The objective of this work is to develop antilisterial gel-based pads (g-pad) for active packaging of cold-smoked salmon. Lysozyme (LYS) and/or green tea extract (GTE) (1% w/w) were incorporated into g-pads of gelatin (GEL), and its mechanically improved composite g-pads obtained by mixing GEL with rice starch (GEL/RS) or candelilla wax (GEL/CW). GEL g-pad with LYS and GEL/RS g-pad with LYS or LYS + GTE, and GEL/CW g-pad with LYS + GTE caused ≥ ~1.8 and ~1.7 log reduction in broth media against Listeria innocua within 11 and 15 days, respectively. All g-pads with LYS or LYS + GTE inhibited L. innocua growth, and gave cold-smoked salmon with 1.5–1.9 log lower Listeria load than controls after 15 days at +4 °C. GEL g-pad with LYS was the most potent on salmon surface since it achieved faster LYS release (1.3–1.8-fold) than other g-pads. Sustained release of GEL/RS g-pads for LYS, and GEL/CW g-pads for GTE was promising for extended storage. Composite g-pads were more compatible with GTE than GEL g-pads since they prevented discoloration of polyphenols. Water absorption capacities showed that GEL/RS g-pads (75%) could prevent drip-loss better than GEL (59%) and GEL/CW (57%) g-pads. Active g-pads provide alternative to active edible films to reduce listeriosis from processed high-risk food.

Microgels prepared from corn starch with an improved capacity for uptake and release of lysozyme

The combination of 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO)-oxidization and annealing was applied to modify structural properties of corn starch in order to obtain starch polymer microgels. Microgels were characterized by X-ray diffraction, ATR-FTIR and thermogravimetry analysis (TGA). The effects of microgels on the uptake and release of lysozyme were also analyzed. The formation of oxidized microgels through the crosslinking between starch molecules and sodium trimetaphosphate was confirmed by X-ray diffraction. The ATR-FTIR analysis suggested that TEMPO-oxidation results in the formation of carboxyl groups. TGA data indicated that oxidized-annealed microgels had less thermal stability than oxidized microgels. Furthermore, the oxidized-annealed microgels had an improved capacity for uptake and release of lysozyme. At this stage of the exploration, microgels prepared from corn starches by annealing and oxidation is simple and efficient.

P34 Early-onset neutropenia in rituximab treated rheumatoid arthritis patients: a case series

Abstract Background Rituximab is a chimeric monoclonal anti-CD20 antibody, used in the management of autoimmunity. It is known to have a favourable safety profile, however, a rare complication includes early-onset neutropenia (EON), defined as neutropenia having occurred within four weeks of rituximab treatment. Only eight case reports have been published so far, the majority of which relate to its use in SLE and none in rheumatoid arthritis (RA). We report here the first dedicated case series of post-rituximab EON in RA. Methods We identified 4 RA patients with EON. All were anti-citrullinated protein antibody (ACPA) and rheumatoid factor (RF) positive. Demographics were female:male ratio 3:1, with mean age 58 (range 33-69). Our first patient (65F) had a past medical history (PMH) of asymptomatic primary biliary cirrhosis (ANA+ve, AMA+ve, M2+ve), the second (65F) had Sjogren’s syndrome (ANA+ve, dsDNA+ve, IgG+ve) and the third (69F) had diverticulosis, Raynaud’s phenomenon and subacute cutaneous lupus with no systemic manifestations (ANA+ve, Ro and La +ve). The fourth patient (33M) had no PMH. Three had previously reported intermittent neutropenia on prior disease modifying anti-rheumatic drug (DMARD) therapy, but none were on DMARDs/doses known to cause this at the time of Rituximab. Standard Rituximab regimens were administered (1g with 100mg IV methylprednisolone, two weeks apart) with two patients (33M, 69F) on concurrent methotrexate (15mg) and hydroxychloroquine (200mg) respectively. Results EON occurred at 4, 11 and 14 days (n = 2) (mean 10 days) post-infusion with mean nadir 0.57x109/L (range 0.05 - 1.17x109/L). It was the first treatment cycle for two patients (65F and 33M), whilst the others (65F and 69F) had received 2-3 previous cycles uneventfully. Patient 2, 65F and ANA+ve, subsequently developed neutropenic sepsis and was treated with standard protocol and G-CSF (granulocyte-colony stimulating factor). Cultures were negative and other investigations did not identify any infective source. The other three patients’ (33M, 69F) neutropenia resolved spontaneously without required intervention. Conclusion EON is a rare but serious complication of rituximab but to date no clear mechanism has been identified. It has been suggested that EON may be due to a bystander effect of lysozyme and granzyme release, secondary to B cell killing and that individuals with a high-affinity FcγRIIIa 158 V allele are more susceptible to this mechanism and therefore prone to a greater depth of neutropenia. Because of its relative acute onset, we may actually be under-recognising uncomplicated episodes of EON due to variability in routine blood monitoring post-treatment. We therefore want to highlight the need for vigilance in this patient cohort, particularly in those who are already susceptible to drug-induced neutropenia or are ANA+ve, in addition to vigilance irrespective of cycle number. Future monitoring of early neutrophil counts in Rituximab-treated RA patients may provide additional insight into this rare complication. Disclosures C. Lin None. A.R. Lorenzi None. F.A.H. Cooles None.

Effects of recombinant human lactoferrin on calcium signaling and functional responses of human neutrophils

Lactoferrin is a non-heme iron-binding glycoprotein with multiple health-beneficial functions including antimicrobial, antioxidant, anticarcinogenic, and immunomodulatory effects. There is emerging evidence that neutrophils may serve as targets of lactoferrin in vivo, and here we show how recombinant human lactoferrin (rhLf) can contribute to this regulation. Indeed, our results demonstrate that rhLf binds efficiently to human neutrophils and induces a variety of early cellular responses such as mobilization of intracellular Ca2+, remodeling of actin cytoskeleton, and degranulation (release of lysozyme and myeloperoxidase). In addition, rhLf facilitates lectin-induced H2O2 production and stabilization of lectin-induced cellular aggregates. The role of calcium signaling seems to be essential for rhLf-induced activation of neutrophils, as Ca2+-chelators inhibit degranulation response while lectin-induced H2O2 production correlates significantly with cytoplasmic Ca2+ elevation. Taken together, our findings justify that rhLf can activate neutrophil functions in a calcium-dependent manner and hence, can potentiate innate immune responses.

Controlled release of lysozyme based core/shells structured alginate beads with CaCO3 microparticles using Pickering emulsion template and in situ gelation

To employ dual advantages of emulsion and gel, a facile approach was investigated to fabricate core/shells structured hydrogel beads based on sodium alginate (SA) via Pickering emulsion template and in situ gelation. The encapsulation and controlled release behavior were further studied using lysozyme (Ly) as the model protein. The optical micrographs and SEM images indicated the SA beads could well disperse with the size about 150 μm. CaCO3 microparticles were strong adhesive onto SA gel. It showed that 96.51 ± 0.62% Ly was loaded into the hydrogel beads. The released behavior of Ly could be regulated by external pH condition, and displayed highest release rate at pH 5.0. Whereas the lowest release rate was recorded at pH 7.0. The released behavior well followed the Hixcon-Crowell model which indicated that the release mechanism of Ly followed the corrosion diffusion law. The worth-while endeavor provide an artful and facile approach using Pickering emulsion template and in situ gelation to fabricate core/shells structured SA beads with high load capacity and controlled regulation of the entrapped functional component.

Antimicrobial activity of lysozyme-nisin co-encapsulated in liposomes coated with polysaccharides

Natural antimicrobials are an innovative alternative for food safety and encapsulation can improve their controlled release and stability. In the present work, lysozyme and its combination with nisin were encapsulated into phosphatidylcholine (PC) liposomes coated with pectin or polygalacturonic acid. The mean particle sizes of liposomes encapsulating lysozyme were 92, 116, and 97 nm for PC, PC-pectin and PC-polygalacutronic acid, respectively. Co-encapsulation of lysozyme and nisin resulted in particle sizes of 86, 77, and 80 nm, respectively. The formulations showed high encapsulation efficiency (77–87%), zeta potential around −30 mV, and spherical structures were observed by transmission electron microscopy. Liposomes encapsulating lysozyme inhibited Listeria monocytogens, but not Salmonella Enteritidis, while liposomes co-encapsulating the mixture nisin/lysozyme inhibited both bacteria. The PC-pectin liposomes were more efficient in inhibiting Listeria spp. when compared to the other liposomes tested. The antimicrobial activity was assessed at 37 °C using milk as model, showing that PC-pectin liposomes reduced the population of L. monocytogenes by 2 log CFU/mL in whole milk and 5 log CFU/mL in skim milk. Under refrigeration temperature, PC-pectin liposomes reduced the population of L. monocytogenes to below detection limit for up to 25 days in skim milk, indicating that liposomes containing polysaccharides can be a promising technology for the controlled release of lysozyme and nisin in foods.

Surface interactions between 2D Ti3C2/Ti2C MXenes and lysozyme

This work presents a comprehensive study on the surface-property changes of 2D flakes of MXenes (Ti3C2 and Ti2C) upon interacting with the model cationic protein – lysozyme, using time-resolved dynamic light scattering and zeta potential. The study was also aimed at investigating how MXenes stoichiometry, structure, surface chemistry and exposure concentration effects its interactions with lysozyme. We found that the adsorption of lysozyme on 2D Ti2C and Ti3C2 flakes occurred effectively, leading to changes in MXenes zeta potential in a concentration-dependent manner. The lysozyme adsorption efficiency varied, depending on the surface chemistry and stoichiometry of the investigated MXenes. The results showed that the Ti3C2 MXene exhibited a higher adsorption of lysozyme in comparison to Ti2C MXene. The zeta potential assays revealed that the adsorption mechanism for MXenes was mainly electrostatic. We also demonstrated the lysozyme release from the MXene surface during pH changes to basic environment. Our findings highlight the importance of evaluating MXenes interactions with biomacromolecules and the mechanism for potential protein adsorption prior to designing e.g. drug delivery systems. We have shown that protein adsorption dramatically changes the surface-physicochemical properties of the MXenes, which may have a significant impact on the effectiveness and safety of future delivery, separation and purification systems dedicated to particular proteins.

Responses of Mytilus galloprovincialis to challenge with the emerging marine pathogen Vibrio coralliilyticus

Vibrio coralliilyticus has emerged as a coral pathogen of concern throughout the Indo-Pacific reef. The interest towards understanding its ecology and pathogenic potential has increased since V. coralliilyticus was shown to be strongly virulent also for other species; in particular, it represents a serious threat for bivalve aquaculture, being one of the most important emerging pathogen responsible for oyster larval mortalities worldwide. V. coralliilyticus has a tightly regulated temperature-dependent virulence and it has been related to mass mortalities events of benthic invertebrates also in the temperate northwestern Mediterranean Sea. However, no data are available on the effects of V. coralliilyticus in the mussel Mytilus galloprovincialis, the most abundant aquacultured species in this area. In this work, responses of M. galloprovincialis to challenge with V. coralliilyticus (ATCC BAA-450) were investigated. In vitro, short term responses of mussel hemocytes were evaluated in terms of lysosomal membrane stability, bactericidal activity, lysozyme release, ROS and NO production, and ultrastructural changes, evaluated by TEM. In vivo, hemolymph parameters were measured in mussels challenged with V. coralliilyticus at 24h p.i. Moreover, the effects of V. coralliilyticus on mussel early embryo development (at 48 hpf) were evaluated. The results show that both in vitro and in vivo, mussels were unable to activate immune response towards V. coralliilyticus, and that challenge mainly induced lysosomal stress in the hemocytes. Moreover, V. coralliilyticus showed a strong and concentration-dependent embryotoxicity. Overall, the results indicate that, although M. galloprovincialis is considered a resistant species to vibrio infections, the emerging pathogen V. coralliilyticus can represent a potential threat to mussel aquaculture.

Studies on immunological and degranulation properties of a galectin-1 purified from goat (Capra hircus) heart

Galectins are mammalian lectins characterized by affinity for β-galactosides and the presence of a conserved carbohydrate recognition domain (CRD). Galectins play crucial role in the causation and progression of deadly human diseases like cancer, neurodegenerative disorders and cardiovascular disorders. Available literature reports relevant roles of galectins in innate as well as adaptive immune responses, along with the modulation of acute inflammatory response. In the current study, we purified the goat heart galectin-1 (GHG-1) and carried out its extensive immunological studies. Immunodiffusion studies revealed that anti-GHG-1 antibodies recognize the GHG-1 more readily as compared to the other galectins, suggesting its preferred utilization in various recognition studies. Antigenic cross-reactivity between galectins isolated from different tissues and species suggest their evolutionary preserved fundamental biological roles. A gradual increase in the lysozyme release was evident when the neutrophils were treated with various neutrophil activating agents. The findings of the present study confirm the increase in lysozyme production under the presence of various neutrophil activators, and thus add new information on GHG-1 induced degranulation.

Fabrication of Sericin/Agrose Gel Loaded Lysozyme and Its Potential in Wound Dressing Application.

Sericin is a biomaterial resource for its significant biodegradability, biocompatibility, hydrophilicity, and reactivity. Designing a material with superabsorbent, antiseptic, and non-cytotoxic wound dressing properties is advantageous to reduce wound infection and promote wound healing. Herein, we propose an environment-friendly strategy to obtain an interpenetrating polymer network gel through blending sericin and agarose and freeze-drying. The physicochemical characterizations of the sericin/agarose gel including morphology, porosity, swelling behavior, crystallinity, secondary structure, and thermal property were well characterized. Subsequently, the lysozyme loaded sericin/agarose composite gel was successfully prepared by the solution impregnation method. To evaluate the potential of the lysozyme loaded sericin/agarose gel in wound dressing application, we analyzed the lysozyme loading and release, antimicrobial activity, and cytocompatibility of the resulting gel. The results showed the lysozyme loaded composite gel had high porosity, excellent water absorption property, and good antimicrobial activities against Escherichia coli and Staphylococcus aureus. Also, the lysozyme loaded gel showed excellent cytocompatibility on NIH3T3 and HEK293 cells. So, the lysozyme loaded sericin/agarose gel is a potential alternative biomaterial for wound dressing.