Using Rapid Prototyping to Develop a Cell-Based Platform with Electrical Impedance Sensor Membranes for In Vitro RPMI2650 Nasal Nanotoxicology Monitoring.

Due to advances in additive manufacturing and prototyping, affordable and rapid microfluidic sensor-integrated assays can be fabricated using additive manufacturing, xurography and electrode shadow masking to create versatile platform technologies aimed toward qualitative assessment of acute cytotoxic or cytolytic events using stand-alone biochip platforms in the context of environmental risk assessment. In the current study, we established a nasal mucosa biosensing platform using RPMI2650 mucosa cells inside a membrane-integrated impedance-sensing biochip using exclusively rapid prototyping technologies. In a final proof-of-concept, we applied this biosensing platform to create human cell models of nasal mucosa for monitoring the acute cytotoxic effect of zinc oxide reference nanoparticles. Our data generated with the biochip platform successfully monitored the acute toxicity and cytolytic activity of 6 mM zinc oxide nanoparticles, which was non-invasively monitored as a negative impedance slope on nasal epithelial models, demonstrating the feasibility of rapid prototyping technologies such as additive manufacturing and xurography for cell-based platform development.

The Cultivation Modality and Barrier Maturity Modulate the Toxicity of Industrial Zinc Oxide and Titanium Dioxide Nanoparticles on Nasal, Buccal, Bronchial, and Alveolar Mucosa Cell-Derived Barrier Models.

As common industrial by-products, airborne engineered nanomaterials are considered important environmental toxins to monitor due to their potential health risks to humans and animals. The main uptake routes of airborne nanoparticles are nasal and/or oral inhalation, which are known to enable the transfer of nanomaterials into the bloodstream resulting in the rapid distribution throughout the human body. Consequently, mucosal barriers present in the nose, buccal, and lung have been identified and intensively studied as the key tissue barrier to nanoparticle translocation. Despite decades of research, surprisingly little is known about the differences among various mucosa tissue types to tolerate nanoparticle exposures. One limitation in comparing nanotoxicological data sets can be linked to a lack of harmonization and standardization of cell-based assays, where (a) different cultivation conditions such as an air-liquid interface or submerged cultures, (b) varying barrier maturity, and (c) diverse media substitutes have been used. The current comparative nanotoxicological study, therefore, aims at analyzing the toxic effects of nanomaterials on four human mucosa barrier models including nasal (RPMI2650), buccal (TR146), alveolar (A549), and bronchial (Calu-3) mucosal cell lines to better understand the modulating effects of tissue maturity, cultivation conditions, and tissue type using standard transwell cultivations at liquid-liquid and air-liquid interfaces. Overall, cell size, confluency, tight junction localization, and cell viability as well as barrier formation using 50% and 100% confluency was monitored using trans-epithelial-electrical resistance (TEER) measurements and resazurin-based Presto Blue assays of immature (e.g., 5 days) and mature (e.g., 22 days) cultures in the presence and absence of corticosteroids such as hydrocortisone. Results of our study show that cellular viability in response to increasing nanoparticle exposure scenarios is highly compound and cell-type specific (TR146 6 ± 0.7% at 2 mM ZnO (ZnO) vs. ~90% at 2 mM TiO2 (TiO2) for 24 h; Calu3 93.9 ± 4.21% at 2 mM ZnO vs. ~100% at 2 mM TiO2). Nanoparticle-induced cytotoxic effects under air-liquid cultivation conditions declined in RPMI2650, A549, TR146, and Calu-3 cells (~0.7 to ~0.2-fold), with increasing 50 to 100% barrier maturity under the influence of ZnO (2 mM). Cell viability in early and late mucosa barriers where hardly influenced by TiO2 as well as most cell types did not fall below 77% viability when added to Individual ALI cultures. Fully maturated bronchial mucosal cell barrier models cultivated under ALI conditions showed less tolerance to acute ZnO nanoparticle exposures (~50% remaining viability at 2 mM ZnO for 24 h) than the similarly treated but more robust nasal (~74%), buccal (~73%), and alveolar (~82%) cell-based models.

Does Tap Water Quality Compromise the Production of Aedes Mosquitoes in Genetic Control Projects?

A mosquito's life cycle includes an aquatic phase. Water quality is therefore an important determinant of whether or not the female mosquitoes will lay their eggs and the resulting immature stages will survive and successfully complete their development to the adult stage. In response to variations in laboratory rearing outputs, there is a need to investigate the effect of tap water (TW) (in relation to water hardness and electrical conductivity) on mosquito development, productivity and resulting adult quality. In this study, we compared the respective responses of Aedes aegypti and Ae. albopictus to different water hardness/electrical conductivity. First-instar larvae were reared in either 100% water purified through reverse osmosis (ROW) (low water hardness/electrical conductivity), 100% TW (high water hardness/electrical conductivity) or a 80:20, 50:50, 20:80 mix of ROW and TW. The immature development time, pupation rate, adult emergence, body size, and longevity were determined. Overall, TW (with higher hardness and electrical conductivity) was associated with increased time to pupation, decreased pupal production, female body size in both species and longevity in Ae. albopictus only. However, Ae. albopictus was more sensitive to high water hardness/EC than Ae. aegypti. Moreover, in all water hardness/electrical conductivity levels tested, Ae. aegypti developed faster than Ae. albopictus. Conversely, Ae. albopictus adults survived longer than Ae. aegypti. These results imply that water with hardness of more than 140 mg/l CaCO3 or electrical conductivity more than 368 µS/cm cannot be recommended for the optimal rearing of Aedes mosquitoes and highlight the need to consider the level of water hardness/electrical conductivity when rearing Aedes mosquitoes for release purposes.

Amygdalin Modulates Prostate Cancer Cell Adhesion and Migration In Vitro.

The natural compound, amygdalin, is notably popular with prostate cancer patients as an alternative or complementary treatment option. However, knowledge about its mode of action is sparse. We investigated amygdalin's impact on prostate cancer adhesion and motile behavior. DU-145 and PC3 cancer cells were exposed to amygdalin. Adhesion to human vascular endothelium or immobilized collagen was then explored. The influence of amygdalin on chemotaxis and migration was also investigated, as well as amygdalin induced alteration to surface and total cellular α and ß integrin expression. Integrin knockdown was performed to evaluate the integrin influence on chemotaxis and adhesion. Amygdalin significantly reduced chemotactic activity, migration, and adhesion of DU-145 but not of PC3 cells. Amygdalin elevated integrin α2 in both cell lines. Integrin α6 was reduced by amygdalin only in DU-145 cells, whereas ß1 increased only in PC3 cells. Functional blocking revealed a negative association of α2 with PC3 and DU-145 chemotaxis. The ß1 increase correlated with enhanced chemotaxis, the diminished α6 expression with reduced chemotaxis. Amygdalin acted on prostate cancer cells in vitro. It induced downregulation of α6 integrin in DU-145 but not in PC3 cells, suggesting that exposing certain prostate cancer cells to amygdalin might inhibit metastatic spread promoted by this particular integrin.

Laser spectroscopy steered 13 C-labelling of plant material in a walk-in growth chamber.

RATIONALE: Carbon-13 (13 C)-labelled plant material forms the basis for experiments elucidating soil organic carbon dynamics and greenhouse gas emissions. Quantitative field-scale tracing is only possible if plants are labelled homogeneously in large quantities. By using a laser spectrometer to automatically steer the isotopic ratio in the chamber, it is possible to obtain large amounts of homogeneously labelled plant material. METHODS: Ninety-six maize plants were labelled for 25 days until tassel formation in a 15 m3 walk-in growth chamber with a continuous air δ13 C-CO2 value of 400‰. A Los Gatos Research laser absorption spectrometer controlled the ambient δ13 C-CO2 value in the chamber through steering of the mass flow controllers with 13 C-enriched and natural abundance CO2 gas. RESULTS: Laser absorption spectroscopy steering kept the δ13 C value of chamber air between 368 and 426‰. The resulting 1 kg dry matter of 13 C-labelled shoots showed an average δ13 C value of 384‰ and accuracy of 8‰ (half width of the 95% confidence interval). Only the oldest leaves showed larger heterogeneity. The growth chamber eliminated variability between plants. The δ13 C value of the stabile material did not differ significantly from that of bulk material. CONCLUSIONS: Laser spectroscopy controlled 13 C labelling of plants in a walk-in growth chamber successfully kept the isotopic ratio of the CO2 in the chamber air constant. Therefore, large quantities of material were labelled homogeneously at the inter- and intra-plant level, thus establishing a method to provide high-quality input for quantitative isotopic tracer studies.

Crystallization speed of salbutamol as a function of relative humidity and temperature.

Spray dried salbutamol sulphate and salbutamol base particles are amorphous as a result of spray drying. As there is always the risk of recrystallization of amorphous material, the aim of this work is the evaluation of the temperature and humidity dependent recrystallization of spray dried salbutamol sulphate and base. Therefore in-situ Powder X-ray Diffraction (PXRD) studies of the crystallization process at various temperature (25 and 35 °C) and humidity (60%, 70%, 80%, 90% relative humidity) conditions were performed. It was shown that the crystallization speed of salbutamol sulphate and base is a non-linear function of both temperature and relative humidity. The higher the relative humidity the higher is the crystallization speed. At 60% relative humidity salbutamol base as well as salbutamol sulphate were found to be amorphous even after 12 h, however samples changed optically. At 70% and 90% RH recrystallization of salbutamol base is completed after 3 h and 30 min and recrystallization of salbutamol sulphate after 4h and 1h, respectively. Higher temperature (35 °C) also leads to increased crystallization speeds at all tested values of relative humidity.

Uptake of (14)C-atropine and/or its transformation products from soil by wheat (Triticum aestivum var Kronjet) and their translocation to shoots.

Plant uptake of toxins and their translocation to edible plant parts are important processes in the transfer of contaminants into the food chain. Atropine, a highly toxic muscarine receptor antagonist produced by Solanacea species, is found in all plant tissues and can enter the soil and hence be available for uptake by crops. The absorption of atropine and/or its transformation products from soil by wheat (Triticum aestivum var Kronjet) and its distribution to shoots was investigated by growing wheat in soil spiked with unlabeled or (14)C-labeled atropine. Radioactivity attributable to (14)C-atropine and its transformation products was measurable in plants sampled at 15 d after sowing (DAS) and thereafter until the end of experiment. The highest accumulation of (14)C-atropine and/or its transformation products by plants was detected in leaves (between 73 and 90% of the total accumulated) with lower amounts in stems, roots, and seeds (approximately 14%, 9%, and 3%, respectively). (14)C-Atropine and/or its transformation products were detected in soil leachate at 30, 60, and 90 DAS and were strongly adsorbed to soil, with 60% of the applied dose adsorbed at 30 DAS, plateauing at 70% from 60 DAS. Unlabeled atropine was detected in shoots 30 DAS at a concentration of 3.9 ± 0.1 µg kg(-1) (mean ± SD). The observed bioconcentration factor was 2.3 ± 0.04. The results suggest a potential risk of atropine toxicity to consumers.