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Neeraj Prabhakar,1,2 Jixi Zhang,3 Diti Desai,1 Eudald Casals,1 Tina Gulin-Sarfraz,1 Tuomas Näreoja,2,4 Jukka Westermarck,5,6 Jessica M Rosenholm1 1Pharmaceutical Sciences Laboratory, Faculty of Science and Engineering, Åbo Akademi University, 2Laboratory of Biophysics, Faculty of Medicine, University of Turku, Turku, Finland; 3College of Bioengineering, Chongqing University, Chongqing, People’s Republic of China; 4Department of Neuroscience, Karolinska Institute, Stockholm, Sweden; 5Centre for Biotechnology, University of Turku and Åbo Akademi, 6Department of Pathology, University of Turku, Turku, Finland Abstract: Small interfering RNA (siRNA) is a highly potent drug in gene-based therapy with the challenge being to deliver it in a sustained manner. The combination of mesoporous silica nanoparticles (MSNs) and polycations in the confined pore space allows for incorporation and controlled release of therapeutic siRNA payloads. We hereby constructed MSNs with expanded mesopores and pore-surface-hyperbranched poly(ethyleneimine) (PEI) tethered with redox-cleavable linkers that could carry a high payload of siRNA (120 mg·g-1). The developed nanocarriers were efficiently taken up by cancer cells and were subsequently able to escape to the cytoplasm from the endosomes, most likely owing to the integrated PEI. Triggered by the intracellular redox conditions, the siRNA was sustainably released inside the cells over a period of several days. Functionality of siRNAs was demonstrated by using cell-killing siRNA as cargo. Despite not being the aim of the developed system, in vitro experiments using cell-killing siRNAs showed that the efficacy of siRNA transfection was comparable to the commercial in vitro transfection agent Lipofectamine. Consequently, the developed MSN-based delivery system offers a potential approach to hybrid nanocarriers for more efficient and long-term siRNA delivery and, in a longer perspective, in vivo gene silencing for RNA interference (RNAi) therapy. Keywords: mesoporous silica nanoparticles, RNAi therapy, siRNA delivery, stimuli-responsive drug release, hybrid nanocarriers

M Rosa Hernández1, Patricia Urbán2, Elisenda Casals3, Joan Estelrich3, Ginés Escolar1, Ana M Galán11Department of Hemotherapy and Hemostasis, Hospital Clinic, CDB, IDIBAPS, UB, Barcelona, Spain; 2Nanobioengineering Group, Institute for Bioengineering of Catalonia, Barcelona, Spain; 3Department of Physical Chemistry, Faculty of Pharmacy, University of Barcelona, SpainBackground: The contribution of fibrinogen (FBN) to hemostasis acting on platelet aggregation and clot formation is well established. It has been suggested that FBN-coated liposomes could be useful in restoring hemostasis. In the present study, we evaluated the modifications induced by multilamellar raw liposomes (MLV) or fibrinogen-coated liposomes (MLV-FBN) on hemostatic parameters.Materials and methods: Different experimental settings using whole blood or thrombocytopenic blood were used. Thromboelastometry, aggregation studies, platelet function analyzer (PFA-100®) tests and studies under flow conditions were applied to detect the effect of MLV-FBN on hemostatic parameters.Results: The presence of MLV-FBN in whole blood modified its viscoelastic properties, prolonging clot formation time (CFT) (226.5 ± 26.1 mm versus 124.1 ± 9.4 mm; P < 0.01) but reducing clot firmness (45.4 ± 1.8 mm versus 35.5 ± 2.3 mm; P < 0.05). Under thrombocytopenic conditions, FIBTEM analysis revealed that MLV-FBN shortened clotting time (CT) compared to MLV (153.3 ± 2.8 s versus 128.0 ± 4.6 s; P < 0.05). Addition of either liposome decreased fibrin formation on the subendothelium (MLV 8.1% ± 4.7% and MLV-FBN 0.8% ± 0.5% versus control 36.4% ± 6.7%; P < 0.01), whereas only MLV-FBN significantly reduced fibrin deposition in thrombocytopenic blood (14.4% ± 6.3% versus control 34.5% ± 5.2%; P < 0.05). MLV-FBN inhibited aggregation induced by arachidonic acid (52.1% ± 8.1% versus 88.0% ± 2.1% in control; P < 0.01) and ristocetin (40.3% ± 8.8% versus 94.3% ± 1.1%; P < 0.005), but it did not modify closure times in PFA-100® studies. In perfusion experiments using whole blood, MLV and MLV-FBN decreased the covered surface (13.25% ± 2.4% and 9.85% ± 2.41%, respectively, versus control 22.0% ± 2.0%; P < 0.01) and the percentage of large aggregates (8.4% ± 2.3% and 3.3% ± 1.01%, respectively, versus control 14.6% ± 1.8%; P < 0.01).Conclusion: Our results reveal that, in addition to the main contribution of fibrinogen to hemostasis, MLV-FBN inhibits platelet-mediated hemostasis and coagulation mechanisms.Keywords: thrombocytopenia, hemostasis, fibrinogen, liposomes, procoagulant activity, fibrin

Abstract Background In nanotoxicology, the exact role of particle shape, in relation to the composition, on the capacity to induce toxicity is largely unknown. We investigated the toxic and immunotoxic effects of silver wires (length: 1.5 - 25 μm; diameter 100 - 160 nm), spherical silver nanoparticles (30 nm) and silver microparticles ( Methods Wires and nanoparticles were synthesized by wet-chemistry methods and extensively characterized. Cell viability and cytotoxicity were assessed and potential immunotoxic effects were investigated. To compare the effects on an activated and a resting immune system, cells were stimulated with rhTNF-α or left untreated. Changes in intracellular free calcium levels were determined using calcium imaging. Finally, ion release from the particles was assessed by ICP-MS and the effects of released ions on cell viability and cytotoxicity were tested. Results No effects were observed for the spherical particles, whereas the silver wires significantly reduced cell viability and increased LDH release from A549 cells. Cytokine promoter induction and NF-κB activation decreased in a concentration dependent manner similar to the decrease seen in cell viability. In addition, a strong increase of intracellular calcium levels within minutes after addition of wires was observed. This toxicity was not due to free silver ions, since the samples with the highest ion release did not induce toxicity and ion release control experiments with cells treated with pre-incubated medium did not show any effects either. Conclusions These data showed that silver wires strongly affect the alveolar epithelial cells, whereas spherical silver particles had no effect. This supports the hypothesis that shape is one of the important factors that determine particle toxicity.

Abstract Background With the increasing use of nanomaterials, the need for methods and assays to examine their immunosafety is becoming urgent, in particular for nanomaterials that are deliberately administered to human subjects (as in the case of nanomedicines). To obtain reliable results, standardised in vitro immunotoxicological tests should be used to determine the effects of engineered nanoparticles on human immune responses. However, before assays can be standardised, it is important that suitable methods are established and validated. Results In a collaborative work between European laboratories, existing immunological and toxicological in vitro assays were tested and compared for their suitability to test effects of nanoparticles on immune responses. The prototypical nanoparticles used were metal (oxide) particles, either custom-generated by wet synthesis or commercially available as powders. Several problems and challenges were encountered during assay validation, ranging from particle agglomeration in biological media and optical interference with assay systems, to chemical immunotoxicity of solvents and contamination with endotoxin. Conclusion The problems that were encountered in the immunological assay systems used in this study, such as chemical or endotoxin contamination and optical interference caused by the dense material, significantly affected the data obtained. These problems have to be solved to enable the development of reliable assays for the assessment of nano-immunosafety.

Soil permeability tests require a time series of water level measurements to determine system losses, including both infiltration and evaporation. Laboratory measurements of flow are standardised by international regulations such as ASTM International, ISO or UNE, but field measurements are not as well described and in some cases may require definition and specification of test conditions. This is the case for geosynthetic clay liner (GCL) products, where permeability is assessed by a laboratory measurement using a flexible wall permeameter as defined in standard test method D 5887-04. This method is not able to evaluate the performance of such products in the field and therefore cannot guarantee their ability to be used for the repair of landfill liner overlays. For this reason, we have defined a field test in a confined steel ring and developed a real-time ultrasonic IoT device to evaluate water losses over a period of time. The test method was applied in Mallorca (Spain) and as a result the quality of a landfill cover repair solution was evaluated, the corresponding civil works were carried out and the basis for future field measurements of soil permeability tests on different materials and conditions was established., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Author(s).)

Advanced oxidation processes using TiO 2 -based nanomaterials are sustainable technologies that hold great promise for the degradation of many types of pollutants including pharmaceutical residues. A wide variety of heterostructures coupling TiO 2 with visible-light active nanomaterials have been explored to shift its photocatalytic properties to harness sun irradiation but a systematic comparison between them is lacking in the current literature. Furthermore, the high number of proposed nanostructures with different size, morphology, and surface area, and the often complex synthesis processes hamper the transition of these materials into commercial and effective solutions for environmental remediation. Herein, we have designed a facile and cost-effective method to synthesize two heterostructured photocatalysts representative of two main families of novel structures proposed, hybrids of TiO 2 with metal (Au) and semiconductor (CeO 2 ) nanomaterials. The photocatalysts have been extensively characterized to ensure a good comparability in terms of co-catalyst doping characteristics, morphology and surface area. The photocatalytic degradation of ciprofloxacin and sulfamethoxazole as target pollutants, two antibiotics of high concern polluting water sources, has been evaluated and CeO 2 /TiO 2 exhibited the highest activity, achieving complete antibiotic degradation at very low photocatalyst concentrations. Our study provides new insights into the development of inexpensive heterostructured photocatalysts and suggests that the non-stoichiometry and characteristic d and f electronic orbital configuration of CeO 2 have a significantly improved role in the enhancement of the photocatalytic reaction., Competing Interests: The authors declare no competing financial interest., (This journal is © The Royal Society of Chemistry.)

In response to the threat of increasing antimicrobial resistance, we must increase the amount of available high-quality genomic data gathered on antibiotic-resistant bacteria. To this end, we developed an integrated pipeline for high-throughput long-read sequencing, assembly, annotation and analysis of bacterial isolates and used it to generate a large genomic data set of carbapenemase-producing Enterobacterales (CPE) isolates collected in Spain. The set of 461 isolates were sequenced with a combination of both Illumina and Oxford Nanopore Technologies (ONT) DNA sequencing technologies in order to provide genomic context for chromosomal loci and, most importantly, structural resolution of plasmids, important determinants for transmission of antimicrobial resistance. We developed an informatics pipeline called Assembly and Annotation of Carbapenem-Resistant Enterobacteriaceae (AACRE) for the full assembly and annotation of the bacterial genomes and their complement of plasmids. To explore the resulting genomic data set, we developed a new database called inCREDBle that not only stores the genomic data, but provides unique ways to filter and compare data, enabling comparative genomic analyses at the level of chromosomes, plasmids and individual genes. We identified a new sequence type, ST5000, and discovered a genomic locus unique to ST15 that may be linked to its increased spread in the population. In addition to our major objective of generating a large regional data set, we took the opportunity to compare the effects of sample quality and sequencing methods, including R9 versus R10 nanopore chemistry, on genome assembly and annotation quality. We conclude that converting short-read and hybrid microbial sequencing and assembly workflows to the latest nanopore chemistry will further reduce processing time and cost, truly enabling the routine monitoring of resistance transmission patterns at the resolution of complete chromosomes and plasmids.

Cerium oxide nanoparticles (CeO 2 NPs) have exceptional catalytic properties, rendering them highly effective in removing excessive reactive oxygen species (ROS) from biological environments, which is crucial in safeguarding these environments against radiation-induced damage. Additionally, the Ce atom's high Z number makes it an ideal candidate for utilisation as an X-ray imaging contrast agent. We herein show how the injection of albumin-stabilised 5 nm CeO 2 NPs into mice revealed substantial enhancement in X-ray contrast, reaching up to a tenfold increase at significantly lower concentrations than commercial or other proposed contrast agents. Remarkably, these NPs exhibited prolonged residence time within the target organs. Thus, upon injection into the tail vein, they exhibited efficient uptake by the liver and spleen, with 85% of the injected dose (%ID) recovered after 7 days. In the case of intratumoral administration, 99% ID of CeO 2 NPs remained within the tumour throughout the 7-day observation period, allowing for observation of disease dynamics. Mass spectrometry (ICP-MS) elemental analysis confirmed X-ray CT imaging observations.

Nanoparticle (NP) pharmacokinetics significantly differ from traditional small molecule principles. From this emerges the need to create new tools and concepts to harness their full potential and avoid unnecessary risks. Nanoparticle pharmacokinetics strongly depend on size, shape, surface functionalisation, and aggregation state, influencing their biodistribution, accumulation, transformations, and excretion profile, and hence their efficacy and safety. Today, while NP biodistribution and nanoceria biodistribution have been studied often at short times, their long-term accumulation and excretion have rarely been studied. In this work, 3 nm nanoceria at 5.7 mg/kg of body weight was intravenously administrated in a single dose to healthy mice. Biodistribution was measured in the liver, spleen, kidney, lung, brain, lymph nodes, ovary, bone marrow, urine, and faeces at different time points (1, 9, 30, and 100 days). Biodistribution and urinary and faecal excretion were also studied in rats placed in metabolic cages at shorter times. The similarity of results of different NPs in different models is shown as the heterogeneous nanoceria distribution in organs. After the expectable accumulation in the liver and spleen, the concentration of cerium decays exponentially, accounting for about a 50% excretion of cerium from the body in 100 days. Cerium ions, coming from NP dissolution, are most likely excreted via the urinary tract, and ceria nanoparticles accumulated in the liver are most likely excreted via the hepatobiliary route. In addition, nanoceria looks safe and does not damage the target organs. No weight loss or apathy was observed during the course of the experiments.

Circulating levels of 2-hydroxybutyrate (2HB) are highly related to glycemic status in different metabolomic studies. According to recent evidence, 2HB is an early biomarker of the future development of dysglycemia and type 2 diabetes mellitus and may be causally related to the progression of normal subjects to impaired fasting glucose or insulin resistance. In the present study, we developed and validated a simple, specific and sensitive gas chromatography-mass spectrometry (GC-MS) method specifically intended to quantify serum levels of 2HB. Liquid-liquid extraction with ethyl acetate was followed by 2 min of microwave-assisted derivatization. The method presented acceptable accuracy, precision and recovery, and the limit of quantification was 5 µM. Levels of 2HB were found to be stable in serum after three freeze-thaw cycles, and at ambient temperature and at a temperature of 4 °C for up to 24 h. Extracts derivatized under microwave irradiation were stable for up to 96 h. No differences were found in 2HB concentrations measured in serum or plasma EDTA samples. In summary, the method is useful for a rapid, precise and accurate quantification of 2HB in serum samples assessed for the evaluation of dysglycemia and diabetes mellitus .

The immune system contributes to maintaining the body's functional integrity through its two main functions: recognizing and destroying foreign external agents (invading microorganisms) and identifying and eliminating senescent cells and damaged or abnormal endogenous entities (such as cellular debris or misfolded/degraded proteins). Accordingly, the immune system can detect molecular and cellular structures with a spatial resolution of a few nm, which allows for detecting molecular patterns expressed in a great variety of pathogens, including viral and bacterial proteins and bacterial nucleic acid sequences. Such patterns are also expressed in abnormal cells. In this context, it is expected that nanostructured materials in the size range of proteins, protein aggregates, and viruses with different molecular coatings can engage in a sophisticated interaction with the immune system. Nanoparticles can be recognized or passed undetected by the immune system. Once detected, they can be tolerated or induce defensive (inflammatory) or anti-inflammatory responses. This paper describes the different modes of interaction between nanoparticles, especially inorganic nanoparticles, and the immune system, especially the innate immune system. This perspective should help to propose a set of selection rules for nanosafety-by-design and medical nanoparticle design.

The wide prevalence of oxidative stress-induced diseases has led to a growing demand for antioxidant therapeutics worldwide. Nanozyme antioxidants are drawing enormous attention as practical alternatives for conventional antioxidants. The considerable body of research over the last decade and the promising results achieved signify the potential of nanozyme antioxidants to secure a place in the expanding market of antioxidant therapeutics. Nonetheless, there is no report on clinical trials for their further evaluation. Through analyzing in-depth selected papers which have conducted in vivo studies on nanozyme antioxidants, this review aims to pinpoint and discuss possible reasons impeding development of research toward clinical studies and to offer some practical solutions for future studies to bridge the gap between preclinical and clinical stages.

Glioblastoma (GB) is the most frequent malignant tumor originating from the central nervous system. Despite breakthroughs in treatment modalities for other cancer types, GB remains largely irremediable due to the high degree of intratumoral heterogeneity, infiltrative growth, and intrinsic resistance towards multiple treatments. A sub-population of GB cells, glioblastoma stem cells (GSCs), act as a reservoir of cancer-initiating cells and consequently, constitute a significant challenge for successful therapy. In this study, we discovered that PEI surface-functionalized mesoporous silica nanoparticles (PEI-MSNs), without any anti-cancer drug, very potently kill multiple GSC lines cultured in stem cell conditions. Very importantly, PEI-MSNs did not affect the survival of established GB cells, nor other types of cancer cells cultured in serum-containing medium, even at 25 times higher doses. PEI-MSNs did not induce any signs of apoptosis or autophagy. Instead, as a potential explanation for their lethality under stem cell culture conditions, we demonstrate that the internalized PEI-MSNs accumulated inside lysosomes, subsequently causing a rupture of the lysosomal membranes. We also demonstrate blood-brain-barrier (BBB) permeability of the PEI-MSNs in vitro and in vivo. Taking together the recent indications for the vulnerability of GSCs for lysosomal targeting and the lethality of the PEI-MSNs on GSCs cultured under stem cell culture conditions, the results enforce in vivo testing of the therapeutic impact of PEI-functionalized nanoparticles in faithful preclinical GB models.

The metabolic ratios lactate/pyruvate and β-hydroxybutyrate/acetoacetate are considered valuable tools to evaluate the in vivo redox cellular state by estimating the free NAD+/NADH in cytoplasm and mitochondria, respectively. The aim of the current study was to validate a gas-chromatography mass spectrometry method for simultaneous determination of the four metabolites in plasma and liver tissue. The procedure included an o-phenylenediamine microwave-assisted derivatization, followed by liquid-liquid extraction with ethyl acetate and silylation with bis(trimethylsilyl)trifluoroacetamide:trimethylchlorosilane 99:1. The calibration curves presented acceptable linearity, with a limit of quantification of 0.001 mM for pyruvate, β-hydroxybutyrate and acetoacetate and of 0.01 mM for lactate. The intra-day and inter-day accuracy and precision were within the European Medicines Agency's Guideline specifications. No significant differences were observed in the slope coefficient of three-point standard metabolite-spiked curves in plasma or liver and water, and acceptable recoveries were obtained in the metabolite-spiked samples. Applicability of the method was tested in precision-cut liver rat slices and also in HepG2 cells incubated under different experimental conditions challenging the redox state. In conclusion, the validated method presented good sensitivity, specificity and reproducibility in the quantification of lactate/pyruvate and β-hydroxybutyrate/acetate metabolites and may be useful in the evaluation of in vivo redox states.

Oxidative stress induced by the overproduction of free radicals or reactive oxygen species (ROS) has been considered as a key pathogenic mechanism contributing to the initiation and progression of injury in liver diseases. Consequently, during the last few years antioxidant substances, such as superoxide dismutase (SOD), resveratrol, colchicine, eugenol, and vitamins E and C have received increasing interest as potential therapeutic agents in chronic liver diseases. These substances have demonstrated their efficacy in equilibrating hepatic ROS metabolism and thereby improving liver functionality. However, many of these agents have not successfully passed the scrutiny of clinical trials for the prevention and treatment of various diseases, mainly due to their unspecificity and consequent uncontrolled side effects, since a minimal level of ROS is needed for normal functioning. Recently, cerium oxide nanoparticles (CeO 2 NPs) have emerged as a new powerful antioxidant agent with therapeutic properties in experimental liver disease. CeO 2 NPs have been reported to act as a ROS and reactive nitrogen species (RNS) scavenger and to have multi-enzyme mimetic activity, including SOD activity (deprotionation of superoxide anion into oxygen and hydrogen peroxide), catalase activity (conversion of hydrogen peroxide into oxygen and water), and peroxidase activity (reducing hydrogen peroxide into hydroxyl radicals). Consequently, the beneficial effects of CeO 2 NPs treatment have been reported in many different medical fields other than hepatology, including neurology, ophthalmology, cardiology, and oncology. Unlike other antioxidants, CeO 2 NPs are only active at pathogenic levels of ROS, being inert and innocuous in healthy cells. In the current article, we review the potential of CeO 2 NPs in several experimental models of liver disease and their safety as a therapeutic agent in humans as well.

The COVID-19 pandemic has posed and is continuously posing enormous societal and health challenges worldwide. The research community has mobilized to develop novel projects to find a cure or a vaccine, as well as to contribute to mass testing, which has been a critical measure to contain the infection in several countries. Through this article, we share our experiences and learnings as a group of volunteers at the Centre for Genomic Regulation (CRG) in Barcelona, Spain. As members of the ORFEU project, an initiative by the Government of Catalonia to achieve mass testing of people at risk and contain the epidemic in Spain, we share our motivations, challenges and the key lessons learnt, which we feel will help better prepare the global society to address similar situations in the future., Competing Interests: No competing interests were disclosed., (Copyright: © 2020 Ghose R et al.)

Cerium oxide nanoparticles (CeO 2 NPs) possess powerful antioxidant properties, thus emerging as a potential therapeutic tool in non-alcoholic fatty liver disease (NAFLD) progression, which is characterized by a high presence of reactive oxygen species (ROS). The aim of this study was to elucidate whether CeO 2 NPs can prevent or attenuate oxidant injury in the hepatic human cell line HepG2 and to investigate the mechanisms involved in this phenomenon. The effect of CeO 2 NPs on cell viability and ROS scavenging was determined, the differential expression of pro-inflammatory and oxidative stress-related genes was analyzed, and a proteomic analysis was performed to assess the impact of CeO 2 NPs on cell phosphorylation in human hepatic cells under oxidative stress conditions. CeO 2 NPs did not modify HepG2 cell viability in basal conditions but reduced H 2 O 2 - and lipopolysaccharide (LPS)-induced cell death and prevented H 2 O 2 -induced overexpression of MPO, PTGS1 and iNOS. Phosphoproteomic analysis showed that CeO 2 NPs reverted the H 2 O 2 -mediated increase in the phosphorylation of peptides related to cellular proliferation, stress response, and gene transcription regulation, and interfered with H 2 O 2 effects on mTOR, MAPK/ERK, CK2A1 and PKACA signaling pathways. In conclusion, CeO 2 NPs protect HepG2 cells from cell-induced oxidative damage, reducing ROS generation and inflammatory gene expression as well as regulation of kinase-driven cell survival pathways.

Background and Aims: Cerium oxide nanoparticles are effective scavengers of reactive oxygen species and have been proposed as a treatment for oxidative stress-related diseases. Consequently, we aimed to investigate the effect of these nanoparticles on hepatic regeneration after liver injury by partial hepatectomy and acetaminophen overdose., Methods: All the in vitro experiments were performed in HepG2 cells. For the acetaminophen and partial hepatectomy experimental models, male Wistar rats were divided into three groups: (1) nanoparticles group, which received 0.1 mg/kg cerium nanoparticles i.v. twice a week for 2 weeks before 1 g/kg acetaminophen treatment, (2) N-acetyl-cysteine group, which received 300 mg/kg of N-acetyl-cysteine i.p. 1 h after APAP treatment and (3) partial hepatectomy group, which received the same nanoparticles treatment before partial hepatectomy. Each group was matched with vehicle-controlled rats., Results: In the partial hepatectomy model, rats treated with cerium oxide nanoparticles showed a significant increase in liver regeneration, compared with control rats. In the acetaminophen experimental model, nanoparticles and N-acetyl-cysteine treatments decreased early liver damage in hepatic tissue. However, only the effect of cerium oxide nanoparticles was associated with a significant increment in hepatocellular proliferation. This treatment also reduced stress markers and increased cell cycle progression in hepatocytes and the activation of the transcription factor NF-κB in vitro and in vivo., Conclusions: Our results demonstrate that the nanomaterial cerium oxide, besides their known antioxidant capacities, can enhance hepatocellular proliferation in experimental models of liver regeneration and drug-induced hepatotoxicity.

Non-alcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease worldwide, ranging from steatosis to non-alcoholic steatohepatitis (NASH). Recently, cerium oxide nanoparticles (CeO 2 NPs) have emerged as a new antioxidant agent with hepatoprotective properties in experimental liver disease. The aim of the current investigation was to elucidate whether CeO 2 NPs display beneficial effects in an experimental model of NAFLD.Therefore, fifteen Wistar rats were subjected to a methionine and choline deficient diet (MCDD) for 6 weeks and intravenously treated with CeO 2 NP or vehicle during the weeks three and four of the diet. The effect of CeO 2 NPs on serum biochemistry, hepatic steatosis, inflammation, fatty acid content and expression of reactive oxygen species (ROS) and lipid metabolism related genes was assessed. MCDD fed rats showed increased inflammation, enhanced hepatic lipid accumulation of both saturated and unsaturated fatty acids (FAs) and overexpression of genes related to fatty liver and ROS metabolism. Treatment with CeO 2 NPs was able to reduce the size and content of hepatocyte lipid droplets, the hepatic concentration of triglyceride- and cholesterol ester-derived FAs and the expression of several genes involved in cytokine, adipokine and chemokine signaling pathways. These findings suggest that CeO 2 NPs could be of beneficial value in NAFLD.

Nonalcoholic fatty liver disease (NAFLD) is characterized by hepatic accumulation of lipids. Antisteatotic effects of cerium oxide nanoparticles (CeO 2 NPs) have recently been shown in animal models of liver disease. However, it is unclear whether the activity of CeO 2 NPs is related solely to the decrease in oxidative stress or, in addition, they directly decrease liver fatty acid accumulation. To address this question, in this work, we used an in vitro model of hepatocellular steatosis, exposing HepG2 cells to oleic and palmitic acid. Cell uptake of CeO 2 NPs and their effect on oxidative stress and viability of hepatic cells cultured with H 2 O 2 were also evaluated. Results show that CeO 2 NPs were uptaken by HepG2 cells and reduced oxidative stress and improved cell viability. Treatment with oleic and palmitic acid increased lipogenesis and the content of different fatty acids. CeO 2 NPs reduced palmitic and stearic acid and most fatty acids consisting of more than 18 carbon atoms. These effects were associated with significant changes in elongase and desaturase activity. In conclusion, CeO 2 NPs directly protected HepG2 cells from cell injury in oxidative stress conditions and reduced fatty acid content in steatotic conditions by inducing specific changes in fatty acid metabolism, thus showing potential in the treatment of NAFLD.

Background and Aims: The occurrence of endothelial alterations in the liver and in the splanchnic vasculature of cirrhotic patients and experimental models of liver diseases has been demonstrated. However, the pathological role of the portal vein endothelium in this clinical context is scarcely studied and, therefore, deserves attention. In this context, we aimed to investigate whether pathological endothelial activation occurs in the portal vein of cirrhotic rats., Methods: Cirrhosis was induced in wistar rats by CCl4 inhalation. We generated immortalized endothelial cells from the portal vein of control (CT-iPVEC) and cirrhotic rats (CH-iPVEC) by retroviral transduction of the SV40 T antigen. We assessed differential gene expression and intracellular reactive oxygen species (ROS) levels in iPVECs and in portal veins of control and cirrhotic rats. Finally, we assessed the therapeutic effectiveness of cerium oxide nanoparticles (CeO2NP) on reversing PVEC activation and macrophage polarization., Results: CH-iPVECs overexpressed collagen-I, endothelin-1, TIMP-1, TIMP-2, IL-6 and PlGF genes. These results were consistent with the differential expression showed by whole portal veins from cirrhotic rats. In addition, CH-iPVECs showed a significant increase in intracellular ROS and the capacity of potentiating M1 polarization in macrophages. The treatment of CH-iPVECs with CeO2NPs blocked intracellular ROS formation and IL-6 and TIMP-2 gene overexpression. In agreement with the in vitro results, the chronic treatment of cirrhotic rats with CeO2NPs also resulted in the blockade of both ROS formation and IL-6 gene overexpression in whole portal veins., Conclusions: Endothelial cells from portal vein of cirrhotic rats depicted an abnormal phenotype characterized by a differential gene expression and the induction of M1 polarization in macrophages. We identified the excess of intracellular reactive oxygen species (ROS) as a major contributor to this altered phenotype. In addition, we demonstrated the utility of the nanomaterial cerium oxide as an effective antioxidant capable of reverse some of these pathological features associated with the portal vein in the cirrhosis condition., Competing Interests: The authors have declared that no competing interests exist.

Motivation: DNA methylation is essential for normal embryogenesis and development in mammals and can be captured at single base pair resolution by whole genome bisulfite sequencing (WGBS). Current available analysis tools are becoming rapidly outdated as they lack sensible functionality and efficiency to handle large amounts of data now commonly created., Results: We developed gemBS, a fast high-throughput bioinformatics pipeline specifically designed for large scale BS-Seq analysis that combines a high performance BS-mapper (GEM3) and a variant caller specifically for BS-Seq data (BScall). gemBS provides genotype information and methylation estimates for all genomic cytosines in different contexts (CpG and non-CpG) and a set of quality reports for comprehensive and reproducible analysis. gemBS is highly modular and can be easily automated, while producing robust and accurate results., Availability and Implementation: gemBS is released under the GNU GPLv3+ license. Source code and documentation are freely available from www.statgen.cat/gemBS., Supplementary Information: Supplementary data are available at Bioinformatics online., (© The Author(s) 2018. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Gene therapy using siRNA molecules is nowadays considered as a promising approach. For successful therapy, development of a stable and reliable vector for siRNA is crucial. Non-viral and non-organic vectors like mesoporous silica nanoparticles (MSN) are associated with lack of most viral vector drawbacks, such as toxicity, immunogenicity, but also generally a low nucleic acid carrying capacity. To overcome this hurdle, we here modified the pore walls of MSNs with surface-hyperbranching polymerized poly(ethyleneimine) (hbPEI), which provides an abundance of amino-groups for loading of a larger amount of siRNA molecules via electrostatic adsorption. After loading, the particles were covered with a second layer of pre-polymerized PEI to provide better protection of siRNA inside the pores, more effective cellular uptake and endosomal escape. To test the transfection efficiency of PEI covered siRNA/MSNs, MDA-MB 231 breast cancer cells stably expressing GFP were used. We demonstrate that PEI-coated siRNA/MSN complexes provide more effective delivery of siRNAs compared to unmodified MSNs. Thus, it can be concluded that appropriately surface-modified MSNs can be considered as prospective vectors for therapeutic siRNA delivery., (© 2018 Shenyang Pharmaceutical University. Published by Elsevier B.V.)

[This corrects the article on p. 970 in vol. 8, PMID: 28855907.].

Mesoporous silica nanoparticles (MSNs) are extensively explored as drug delivery systems, but in depth understanding of design-toxicity relationships is still scarce. We used zebrafish (Danio rerio) embryos to study toxicity profiles of differently surface functionalized MSNs. Embryos with the chorion membrane intact, or dechoroniated embryos, were incubated or microinjected with amino (NH 2 -MSNs), polyethyleneimine (PEI-MSNs), succinic acid (SUCC-MSNs) or polyethyleneglycol (PEG-MSNs) functionalized MSNs. Toxicity was assessed by viability and cardiovascular function. NH 2 -MSNs, SUCC-MSNs and PEG-MSNs were well tolerated, 50 µg/ml PEI-MSNs induced 100% lethality 48 hours post fertilization (hpf). Dechoroniated embryos were more sensitive and 10 µg/ml PEI-MSNs reduced viability to 5% at 96hpf. Sensitivity to PEG- and SUCC-, but not NH 2 -MSNs, was also enhanced. Typically cardiovascular toxicity was evident prior to lethality. Confocal microscopy revealed that PEI-MSNs penetrated into the embryos whereas PEG-, NH2- and SUCC-MSNs remained aggregated on the skin surface. Direct exposure of inner organs by microinjecting NH 2 -MSNs and PEI-MSNs demonstrated that the particles displayed similar toxicity indicating that functionalization affects the toxicity profile by influencing penetrance through biological barriers. The data emphasize the need for careful analyses of toxicity mechanisms in relevant models and constitute an important knowledge step towards the development of safer and sustainable nanotherapies.

We review the apparent discrepancies between studies that report anti-inflammatory effects of cerium oxide nanoparticles (CeO 2 NPs) through their reactive oxygen species-chelating properties and immunological studies highlighting their toxicity. We observe that several underappreciated parameters, such as aggregation size and degree of impurity, are critical determinants that need to be carefully addressed to better understand the NP biological effects in order to unleash their potential clinical benefits. This is because NPs can evolve toward different states, depending on the environment where they have been dispersed and how they have been dispersed. As a consequence, final characteristics of NPs can be very different from what was initially designed and produced in the laboratory. Thus, aggregation, corrosion, and interaction with extracellular matrix proteins critically modify NP features and fate. These modifications depend to a large extent on the characteristics of the biological media in which the NPs are dispersed. As a consequence, when reviewing the scientific literature, it seems that the aggregation state of NPs, which depends on the characteristics of the dispersing media, may be more significant than the composition or original size of the NPs. In this work, we focus on CeO 2 NPs, which are reported sometimes to be protective and anti-inflammatory, and sometimes toxic and pro-inflammatory.