Adriamycin loading and release characteristics of albumin-heparin conjugate microspheres

Biodegradable ion-exchange microspheres, prepared from a prefabricated conjugate of albumin and heparin were investigated as carriers for adriamycin. The ion-exchange microspheres could be loaded with adriamycin giving payloads up to 33% w/w, depending on the heparin content of the conjugate. In vitro adriamycin release depended on the ionic strength of the release medium. In ion containing media, for instance saline, 90% of the drug was released within 45 min, whereas in non-ionic media, such as distilled water, only 30% was released. Drug release profiles could be modelled by combining ion-exchange kinetics and diffusion controlled drug release models

Preparation and characterization of albumin-heparin microspheres

Albumin-heparin microspheres were prepared by a two-step process which involved the preparation of a soluble albumin-heparin conjugate, followed by formation of microspheres from this conjugate or by a double cross-linking technique involving both coupling of soluble albumin and heparin and microsphere stabilization in one step. The first technique was superior since it allowed better control over the composition and the homogeneity of the microspheres. Microspheres could be prepared with a diameter of 5¿35¿m. The size could be controlled by adjusting the emulsification conditions. The degree of swelling of the microspheres was sensitive to external stimuli, and increased with increasing pH and decreasing ionic strength of the medium

Switching the mode of drug release from a reaction-coupled low-molecular-weight gelator system by altering its reaction pathway

Low-molecular-weight hydrogels are attractive scaffolds for drug delivery applications because of their modular and facile preparation starting from inexpensive molecular components. The molecular design of the hydrogelator results in a commitment to a particular release strategy, where either noncovalent or covalent bonding of the drug molecule dictates its rate and mechanism. Herein, we demonstrate an alternative approach using a reaction-coupled gelator to tune drug release in a facile and user-defined manner by altering the reaction pathway of the low-molecular-weight gelator (LMWG) and drug components through an acylhydrazone-bond-forming reaction. We show that an off-the-shelf drug with a reactive handle, doxorubicin, can be covalently bound to the gelator through its ketone moiety when the addition of the aldehyde component is delayed from 0 to 24 h, or noncovalently bound with its addition at 0 h. We also examine the use of an L-histidine methyl ester catalyst to prepare the drugloaded hydrogels under physiological conditions. Fitting of the drug release profiles with the Korsmeyer-Peppas model corroborates a switch in the mode of release consistent with the reaction pathway taken: increased covalent ligation drives a transition from a Fickian to a semi-Fickian mode in the second stage of release with a decreased rate. Sustained release of doxorubicin from the reaction-coupled hydrogel is further confirmed in an MTT toxicity assay with MCF-7 breast cancer cells. We demonstrate the modularity and ease of the reaction-coupled approach to prepare drug-loaded self-assembled hydrogels in situ with tunable mechanics and drug release profiles that may find eventual applications in macroscale drug delivery.Supramolecular & Biomaterials Chemistr

Coverage of endangered species in environmental risk assessments at EFSA

The EFSA performs environmental risk assessment (ERA) for single potential stressors such as plantprotection products, genetically modified organisms and feed additives, and for invasive alien speciesthat are harmful to plant health. This ERA focusses primarily on the use or spread of such potentialstressors in an agricultural context, but also considers the impact on the wider environment. It isimportant to realise that the above potential stressors in most cases contribute a minor proportion ofthe total integrated pressure that ecosystems experience. The World Wildlife Fund listed the relativeattribution of threats contributing to the declines in animal populations as follows: 37% fromexploitation (fishing, hunting, etc.), 31% habitat degradation and change, 13% from habitat loss, 7%from climate change, and only 5% from invasive species, 4% from pollution and 2% from disease. Inthis scientific opinion, the Scientific Committee gathered scientific knowledge on the extent of coverageof endangered species in current ERA schemes that fall under the remit of EFSA. The legal basis andthe relevant ecological and biological features used to classify a species as endangered areinvestigated. The characteristics that determine vulnerability of endangered species are reviewed.Whether endangered species are more at risk from exposure to potential stressors than other non-target species is discussed, but specific protection goals for endangered species are not given. Due toa lack of effect and exposure data for the vast majority of endangered species, the reliability of usingdata from other species is a key issue for their ERA. This issue and other uncertainties are discussedwhen reviewing the coverage of endangered species in current ERA schemes. Potential tools, such aspopulation and landscape modelling and trait-based approaches, for extending the coverage ofendangered species in current ERA schemes, are explored and reported

Assessing the health status of managed honeybee colonies (HEALTHY-B): a toolbox to facilitate harmonised data collection

Tools are provided to assess the health status of managed honeybee colonies by facilitating further harmonisation of data collection and reporting, design of field surveys across the European Union (EU) and analysis of data on bee health. The toolbox is based on characteristics of a healthy managed honeybee colony: an adequate size, demographic structure and behaviour; an adequate production of bee products (both in relation to the annual life cycle of the colony and the geographical location); and provision of pollination services. The attributes ‘queen presence and performance’, ‘demography of the colony’, ‘in-hive products’ and ‘disease, infection and infestation’ could be directly measured in field conditions across the EU, whereas ‘behaviour and physiology’ is mainly assessed through experimental studies. Analysing the resource providing unit, in particular land cover/use, of a honeybee colony is very important when assessing its health status, but tools are currently lacking that could be used at apiary level in field surveys across the EU. Data on ‘beekeeping management practices’ and ‘environmental drivers’ can be collected via questionnaires and available databases, respectively. The capacity to provide pollination services is regarded as an indication of a healthy colony, but it is assessed only in relation to the provision of honey because technical limitations hamper the assessment of pollination as regulating service (e.g. to pollinate wild plants) in field surveys across the EU. Integrating multiple attributes of honeybee health, for instance, via a Health Status Index, is required to support a holistic assessment. Examples are provided on how the toolbox could be used by different stakeholders. Continued interaction between the Member State organisations, the EU Reference Laboratory and EFSA is required to further validate methods and facilitate the efficient use of precise and accurate bee health data that are collected by many initiatives throughout the EU

A mouse model for oral squamous cell carcinoma

Despite recent advances, the prognosis of oral squamous cell carcinoma is still poor. Therapeutic options such as radiotherapy, chemotherapy, surgery and the novel treatment option gene therapy are being investigated in animal models. Diverse models have been studied to induce oral squamous cell carcinomas. The carcinogenic 4-nitroquinoline-1-oxide (4NQO) model has proven to be successful although until now it is unknown at what time point the established tumor is a representative squamous cell carcinoma and has a suitable volume for scientific treatment. For this end we applied 4NQO 3 times a week during 16 weeks and measured the volume of tumor tissue each week until the end of the experiment at 40 weeks. Concurrent histopathology at different time points up to the end of the experiment revealed that all mice bearing oral tumors were diagnosed with squamous cell carcinoma. Immunohistochemistry with markers cyclin D1 and E-cadherin revealed that the generated mouse oral tumors showed strong similarities with the described immunopathology in human oral tumors. The 4NQO model is a suitable alternative for preclinical gene therapy experiments with primary oral tumors. Future survey of therapeutic options in the carcinogenic 4NQO model should be conducted around 40 weeks after the start of the treatment

Transcriptomic Profiling of Virus-Host Cell Interactions following Chicken Anaemia Virus (CAV) Infection in an In Vivo Model.

Chicken Anaemia Virus (CAV) is an economically important virus that targets lymphoid and erythroblastoid progenitor cells leading to immunosuppression. This study aimed to investigate the interplay between viral infection and the host's immune response to better understand the pathways that lead to CAV-induced immunosuppression. To mimic vertical transmission of CAV in the absence of maternally-derived antibody, day-old chicks were infected and their responses measured at various time-points post-infection by qRT-PCR and gene expression microarrays. The kinetics of mRNA expression levels of signature cytokines of innate and adaptive immune responses were determined by qRT-PCR. The global gene expression profiles of mock-infected (control) and CAV-infected chickens at 14 dpi were also compared using a chicken immune-related 5K microarray. Although in the thymus there was evidence of induction of an innate immune response following CAV infection, this was limited in magnitude. There was little evidence of a Th1 adaptive immune response in any lymphoid tissue, as would normally be expected in response to viral infection. Most cytokines associated with Th1, Th2 or Treg subsets were down-regulated, except IL-2, IL-13, IL-10 and IFNγ, which were all up-regulated in thymus and bone marrow. From the microarray studies, genes that exhibited significant (greater than 1.5-fold, false discovery rate <0.05) changes in expression in thymus and bone marrow on CAV infection were mainly associated with T-cell receptor signalling, immune response, transcriptional regulation, intracellular signalling and regulation of apoptosis. Expression levels of a number of adaptor proteins, such as src-like adaptor protein (SLA), a negative regulator of T-cell receptor signalling and the transcription factor Special AT-rich Binding Protein 1 (SATB1), were significantly down-regulated by CAV infection, suggesting potential roles for these genes as regulators of viral infection or cell defence. These results extend our understanding of CAV-induced immunosuppression and suggest a global immune dysregulation following CAV infection

An untargeted multi-technique metabolomics approach to studying intracellular metabolites of HepG2 cells exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin

<p>Abstract</p> <p>Background</p> <p><it>In vitro </it>cell systems together with omics methods represent promising alternatives to conventional animal models for toxicity testing. Transcriptomic and proteomic approaches have been widely applied <it>in vitro </it>but relatively few studies have used metabolomics. Therefore, the goal of the present study was to develop an untargeted methodology for performing reproducible metabolomics on <it>in vitro </it>systems. The human liver cell line HepG2, and the well-known hepatotoxic and non-genotoxic carcinogen 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), were used as the <it>in vitro </it>model system and model toxicant, respectively.</p> <p>Results</p> <p>The study focused on the analysis of intracellular metabolites using NMR, LC-MS and GC-MS, with emphasis on the reproducibility and repeatability of the data. State of the art pre-processing and alignment tools and multivariate statistics were used to detect significantly altered levels of metabolites after exposing HepG2 cells to TCDD. Several metabolites identified using databases, literature and LC-nanomate-Orbitrap analysis were affected by the treatment. The observed changes in metabolite levels are discussed in relation to the reported effects of TCDD.</p> <p>Conclusions</p> <p>Untargeted profiling of the polar and apolar metabolites of <it>in vitro </it>cultured HepG2 cells is a valid approach to studying the effects of TCDD on the cell metabolome. The approach described in this research demonstrates that highly reproducible experiments and correct normalization of the datasets are essential for obtaining reliable results. The effects of TCDD on HepG2 cells reported herein are in agreement with previous studies and serve to validate the procedures used in the present work.</p