Establishment of a Transgenic Mouse Model Specifically Expressing Human Serum Amyloid A in Adipose Tissue

Obesity and obesity co-morbidities are associated with a low grade inflammation and elevated serum levels of acute phase proteins, including serum amyloid A (SAA). In the non-acute phase in humans, adipocytes are major producers of SAA but the function of adipocyte-derived SAA is unknown. To clarify the role of adipocyte-derived SAA, a transgenic mouse model expressing human SAA1 (hSAA) in adipocytes was established. hSAA expression was analysed using real-time PCR analysis. Male animals were challenged with a high fat (HF) diet. Plasma samples were subjected to fast protein liquid chromatography (FPLC) separation. hSAA, cholesterol and triglyceride content were measured in plasma and in FPLC fractions. Real-time PCR analysis confirmed an adipose tissue-specific hSAA gene expression. Moreover, the hSAA gene expression was not influenced by HF diet. However, hSAA plasma levels in HF fed animals (37.7±4.0 µg/mL, n = 7) were increased compared to those in normal chow fed animals (4.8±0.5 µg/mL, n = 10; p<0.001), and plasma levels in the two groups were in the same ranges as in obese and lean human subjects, respectively. In FPLC separated plasma samples, the concentration of hSAA peaked in high-density lipoprotein (HDL) containing fractions. In addition, cholesterol distribution over the different lipoprotein subfractions as assessed by FPLC analysis was similar within the two experimental groups. The established transgenic mouse model demonstrates that adipose tissue produced hSAA enters the circulation, resulting in elevated plasma levels of hSAA. This new model will enable further studies of metabolic effects of adipose tissue-derived SAA

Is hyperoxaemia helping or hurting patients during extracorporeal membrane oxygenation? Review of a complex problem

Extracorporeal membrane oxygenation (ECMO) facilitates organ support in patients with refractory cardiorespiratory failure whilst disease-modifying treatments can be administered. Improvements to the ECMO process have resulted in its increased utilisation. However, iatrogenic injuries remain, with bleeding and thrombosis the most significant concerns. Many factors contribute to the formation of thrombi, with the hyperoxaemia experienced during ECMO a potential contributor. Outside of ECMO, emerging evidence associates hyperoxaemia with increased mortality. Currently, no universal definition of hyperoxaemia exists, a gap in clinical standards that may impact patient outcomes. Hyperoxaemia has the potential to induce platelet activation, aggregation and, subsequently, thrombosis through markedly increasing the production of reactive oxygen species. There are minimal data in the current literature that explore the relationship between ECMO-induced hyperoxaemia and the production of reactive oxygen species – a putative link towards pathology. Furthermore, there is limited research directly linking hyperoxaemia and platelet activation. These are areas that warrant investigation as definitive data regarding the nascence of these pathological processes may delineate and define the relative risk of supranormal oxygen tension. These data could then assist in defining optimal oxygenation practice, reducing the risks associated with extracorporeal support

Controlled oxidation of iron nanoparticles in chemical vapour synthesis

In the present study, iron oxide nanoparticles (primary particle size of 80-90 nm) with controlled oxidation state were prepared via an atmospheric pressure chemical vapour synthesis (APCVS) method. Iron pentacarbonyl [Fe(CO)5], a precursor material, was thermally decomposed to iron in the APCVS reactor. Subsequently, the iron was oxidized with controlled amount of oxygen in the reactor to produce nearly pure magnetite or haematite particles depending on the oxygen concentration. Size, morphology and crystal structure of the synthesized nanoparticles were studied with scanning mobility particle sizer (SMPS), transmission electron microscopy (TEM) and X-ray diffraction (XRD). In addition, thermodynamic equilibrium calculations and computational fluid dynamics model were used to predict the oxidation state of the iron oxides and the reaction conditions during mixing. Aggregates of crystalline particles were formed, determined as magnetite at the oxygen volumetric fraction of 0.1 % and haematite at volumetric fraction of 0.5 %, according to the XRD. The geometric mean electrical mobility diameter of the aggregates increased from 110 to 155 nm when the volumetric fraction of oxygen increased from 0.1 to 0.5 %, determined using the SMPS. The aggregates were highly sintered based on TEM analyses. As a conclusion, APCVS method can be used to produce nearly pure crystalline magnetite or haematite nanoparticles with controlled oxidation in a continuous one-stage gas-phase proces

Dose Dependent Side Effect of Superparamagnetic Iron Oxide Nanoparticle Labeling on Cell Motility in Two Fetal Stem Cell Populations

Multipotent stem cells (SCs) could substitute damaged cells and also rescue degeneration through the secretion of trophic factors able to activate the endogenous SC compartment. Therefore, fetal SCs, characterized by high proliferation rate and devoid of ethical concern, appear promising candidate, particularly for the treatment of neurodegenerative diseases. Super Paramagnetic Iron Oxide nanoparticles (SPIOn), routinely used for pre-clinical cell imaging and already approved for clinical practice, allow tracking of transplanted SCs and characterization of their fate within the host tissue, when combined with Magnetic Resonance Imaging (MRI). In this work we investigated how SPIOn could influence cell migration after internalization in two fetal SC populations: human amniotic fluid and chorial villi SCs were labeled with SPIOn and their motility was evaluated. We found that SPIOn loading significantly reduced SC movements without increasing production of Reactive Oxygen Species (ROS). Moreover, motility impairment was directly proportional to the amount of loaded SPIOn while a chemoattractant-induced recovery was obtained by increasing serum levels. Interestingly, the migration rate of SPIOn labeled cells was also significantly influenced by a degenerative surrounding. In conclusion, this work highlights how SPIOn labeling affects SC motility in vitro in a dose-dependent manner, shedding the light on an important parameter for the creation of clinical protocols. Establishment of an optimal SPIOn dose that enables both a good visualization of grafted cells by MRI and the physiological migration rate is a main step in order to maximize the effects of SC therapy in both animal models of neurodegeneration and clinical studies

Silver and Gold Nanoparticles Exposure to In Vitro Cultured Retina – Studies on Nanoparticle Internalization, Apoptosis, Oxidative Stress, Glial- and Microglial Activity

The complex network of neuronal cells in the retina makes it a potential target of neuronal toxicity - a risk factor for visual loss. With growing use of nanoparticles (NPs) in commercial and medical applications, including ophthalmology, there is a need for reliable models for early prediction of NP toxicity in the eye and retina. Metal NPs, such as gold and silver, gain much of attention in the ophthalmology community due to their potential to cross the barriers of the eye. Here, NP uptake and signs of toxicity were investigated after exposure to 20 and 80 nm Ag- and AuNPs, using an in vitro tissue culture model of the mouse retina. The model offers long-term preservation of retinal cell types, numbers and morphology and is a controlled system for delivery of NPs, using serum-free defined culture medium. AgNO3-treatment was used as control for toxicity caused by silver ions. These end-points were studied; gross morphological organization, glial activity, microglial activity, level of apoptosis and oxidative stress, which are all well described as signs of insult to neural tissue. TEM analysis demonstrated cellular- and nuclear uptake of all NP types in all neuronal layers of the retina. Htx-eosin staining showed morphological disruption of the normal complex layered retinal structure, vacuole formation and pyknotic cells after exposure to all Ag- and AuNPs. Significantly higher numbers of apoptotic cells as well as an increased number of oxidative stressed cells demonstrated NP-related neuronal toxicity. NPs also caused increased glial staining and microglial cell activation, typical hallmarks of neural tissue insult. This study demonstrates that low concentrations of 20 and 80 nm sized Ag- and AuNPs have adverse effects on the retina, using an organotypic retina culture model. Our results motivate careful assessment of candidate NP, metallic or-non-metallic, to be used in neural systems for therapeutic approaches