A newly developed in vitro model of the human epithelial airway barrier to study the toxic potential of nanoparticles

The potential health effects of inhaled engineered nanoparticles are almost unknown. To avoid and replace toxicity studies with animals, a triple cell co-culture system composed of epithelial cells, macrophages and dendritic cells was established, which simulates the most important barrier functions of the epithelial airway. Using this model, the toxic potential of titanium dioxide was assessed by measuring the production of reactive oxygen species and the release of tumour necrosis factor alpha. The intracellular localisation of titanium dioxide nanoparticles was analyzed by energy filtering transmission electron microscopy. Titanium dioxide nanoparticles were detected as single particles without membranes and in membrane-bound agglomerates. Cells incubated with titanium dioxide particles showed an elevated production of reactive oxygen species but no increase of the release of tumour necrosis factor alpha. Our in vitro model of the epithelial airway barrier offers a valuable tool to study the interaction of particles with lung cells at a nanostructural level and to investigate the toxic potential of nanoparticles.

In vitro models of the human epithelial airway barrier to study the toxic potential of particulate matter

BACKGROUND: Several epidemiological studies show that inhalation of particulate matter may cause increased pulmonary morbidity and mortality. Of particular interest are the ultrafine particles that are particularly toxic. In addition more and more nanoparticles are released into the environment; however, the potential health effects of these nanoparticles are yet unknown. OBJECTIVES: To avoid particle toxicity studies with animals many cell culture models have been developed during the past years. METHODS: This review focuses on the most commonly used in vitro epithelial airway and alveolar models to study particle-cell interactions and particle toxicity and highlights advantages and disadvantages of the different models. RESULTS/CONCLUSION: There are many lung cell culture models but none of these models seems to be perfect. However, they might be a great tool to perform basic research or toxicity tests. The focus here is on 3D and co-culture models, which seem to be more realistic than monocultures.

Translocation and cellular entering mechanisms of nanoparticles in the respiratory tract

Anthropogenic nano-sized particles (NSP), ie, particles with a diameter of less than 100 nm, are generated with or without purpose as chemically and physically well-defined materials or as a consequence of combustion processes respectively. Inhalation of NSP occurs on a regular basis due to air pollution and is associated with an increase in respiratory and cardiovascular morbidity and mortality. Manufactured NSP may intentionally be inhaled as pharmaceuticals or unintentionally during production at the workplace. Hence the interactions of NSP with the respiratory tract are currently under intensive investigation. Due to special physicochemical features of NSP, its biological behaviour may differ from that of larger sized particles. Here we review two important themes of current research into the effects of NSP on the lungs: 1) The potential of NSP to cross the blood-air barrier of the lungs, thus gaining access to the circulation and extrapulmonary organs. It is currently accepted that a small fraction of inhaled NSP may translocate to the circulation. The significance of this translocation requires further research. 2) The entering mechanisms of NSP into different cell types. There is evidence that NSP are taken up by cells via well-known pathways of endocytosis but also via different mechanisms not well understood so far. Knowledge of the quantitative relationship between the different entering mechanisms and cellular responses is not yet available but is urgently needed in order to understand the effects of intentionally or unintentionally inhaled NSP on the respiratory tract.

Empirical evidence of bias in treatment effect estimates in controlled trials with different interventions and outcomes: meta-epidemiological study

OBJECTIVE: To examine whether the association of inadequate or unclear allocation concealment and lack of blinding with biased estimates of intervention effects varies with the nature of the intervention or outcome. DESIGN: Combined analysis of data from three meta-epidemiological studies based on collections of meta-analyses. DATA SOURCES: 146 meta-analyses including 1346 trials examining a wide range of interventions and outcomes. MAIN OUTCOME MEASURES: Ratios of odds ratios quantifying the degree of bias associated with inadequate or unclear allocation concealment, and lack of blinding, for trials with different types of intervention and outcome. A ratio of odds ratios <1 implies that inadequately concealed or non-blinded trials exaggerate intervention effect estimates. RESULTS: In trials with subjective outcomes effect estimates were exaggerated when there was inadequate or unclear allocation concealment (ratio of odds ratios 0.69 (95% CI 0.59 to 0.82)) or lack of blinding (0.75 (0.61 to 0.93)). In contrast, there was little evidence of bias in trials with objective outcomes: ratios of odds ratios 0.91 (0.80 to 1.03) for inadequate or unclear allocation concealment and 1.01 (0.92 to 1.10) for lack of blinding. There was little evidence for a difference between trials of drug and non-drug interventions. Except for trials with all cause mortality as the outcome, the magnitude of bias varied between meta-analyses. CONCLUSIONS: The average bias associated with defects in the conduct of randomised trials varies with the type of outcome. Systematic reviewers should routinely assess the risk of bias in the results of trials, and should report meta-analyses restricted to trials at low risk of bias either as the primary analysis or in conjunction with less restrictive analyses.

Identification of novel mutations in X-linked retinitis pigmentosa families and implications for diagnostic testing

PURPOSE: The goal of this study was to identify mutations in X-chromosomal genes associated with retinitis pigmentosa (RP) in patients from Germany, The Netherlands, Denmark, and Switzerland. METHODS: In addition to all coding exons of RP2, exons 1 through 15, 9a, ORF15, 15a and 15b of RPGR were screened for mutations. PCR products were amplified from genomic DNA extracted from blood samples and analyzed by direct sequencing. In one family with apparently dominant inheritance of RP, linkage analysis identified an interval on the X chromosome containing RPGR, and mutation screening revealed a pathogenic variant in this gene. Patients of this family were examined clinically and by X-inactivation studies. RESULTS: This study included 141 RP families with possible X-chromosomal inheritance. In total, we identified 46 families with pathogenic sequence alterations in RPGR and RP2, of which 17 mutations have not been described previously. Two of the novel mutations represent the most 3'-terminal pathogenic sequence variants in RPGR and RP2 reported to date. In exon ORF15 of RPGR, we found eight novel and 14 known mutations. All lead to a disruption of open reading frame. Of the families with suggested X-chromosomal inheritance, 35% showed mutations in ORF15. In addition, we found five novel mutations in other exons of RPGR and four in RP2. Deletions in ORF15 of RPGR were identified in three families in which female carriers showed variable manifestation of the phenotype. Furthermore, an ORF15 mutation was found in an RP patient who additionally carries a 6.4 kbp deletion downstream of the coding region of exon ORF15. We did not identify mutations in 39 sporadic male cases from Switzerland. CONCLUSIONS: RPGR mutations were confirmed to be the most frequent cause of RP in families with an X-chromosomal inheritance pattern. We propose a screening strategy to provide molecular diagnostics in these families.

Acute disseminated encephalomyelitis in adults: a reappraisal of clinical, CSF, EEG, and MRI findings

OBJECTIVES: To establish an adequate definition of acute disseminated encephalomyelitis (ADEM) in adults, based on our clinical observations of a case-series. METHODS: Over a period of three years 10 adult patients with a para- or postinfectious disseminated (diffuse or multifocal) syndrome of the CNS fulfilling predefined strict criteria for the diagnosis of ADEM were encountered and systematically followed. RESULTS: The age ranged from 21 to 62 years, two were men. MRI was normal in 5 patients and only mildly abnormal in the remaining patients. CSF was normal in 5 patients and mildly abnormal in the remainder, EEG was abnormal in 7/8 patients. All patients survived and were followed over a period of 30 months (range: 8 to 48 months). Nine patients were left with some residual defects, consisting most often of a mild cognitive impairment. CONCLUSIONS: The EEG as an investigation of brain function can be crucial in establishing the organic nature of disease. MRI is important to exclude other diffuse or multifocal encephalopathies. However, in contrast to previous reports in the literature abnormal MRI should not be considered mandatory in adult ADEM. Difficulties in the diagnosis of ADEM are discussed and the importance of clinical and paraclinical findings for establishing the diagnosis is outlined.