Cultex Laboratories GmbH

Hannover, Germany

Cultex Laboratories GmbH

Hannover, Germany
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Aufderheide M.,Cultex Laboratories GmbH | Emura M.,Cultex Laboratories GmbH
Experimental and Toxicologic Pathology | Year: 2017

3D constructs composed of differentiated immortalized primary normal human bronchial epithelial (NHBE) cells (CL-1548) were repeatedly exposed at the air-liquid interface to non-lethal concentrations of mainstream cigarette smoke (4 cigarettes a day, 5. days/week, 8 repetitions in total) and e-cigarette vapor (50 puffs a day, 5 days/week, 8 repetitions in total) to build up a permanent burden on the cells. Samples were taken after 4, 6 and 8 times of repeated smoke exposure and the cultures were investigated using histopathological methods Compared to the clean air-exposed cultures (process control) and incubator control, the aerosol-exposed cultures showed a reduction of ciliated, mucus-producing and club cells. At the end of the exposure phase, we even found metaplastic areas positive for CK13 antibody in the cultures exposed to mainstream cigarette smoke and e-liquid vapor, commonly seen in squamous cells as a marker for non-cornified squamous epithelium. The control cultures (incubator cells) showed no comparable phenotypical changes. In conclusion, our in vitro model presents a valuable tool to study the induction of phenotypical changes after exposure to hazardous airborne material. © 2017.


Aufderheide M.,Cultex Laboratories GmbH | Ito S.,Japan Tobacco Inc. | Ishikawa S.,Japan Tobacco Inc. | Emura M.,Cultex Laboratories GmbH
Experimental and Toxicologic Pathology | Year: 2017

3D constructs composed of primary normal differentiated human bronchiolar epithelial (NHBE) cells as mono- or co-culture in combination with normal human lung fibroblasts were exposed repeatedly at the air-liquid interface with non-lethal concentrations of mainstream cigarette smoke (4 cigarettes a day, 5. days/week, 13 times repetition in total) to build up a permanent burden on the cells. Samples were taken after 4, 8 and 13 times of repeated smoke exposure and the cultures were analyzed by histopathological methods In comparison with the clean air exposure (process control) and incubator control cells the cigarette smoke exposed cultures showed a reduction of cilia bearing as well as mucus producing cells. In both mono- as well as co-cultures, hyperplasia was induced showing different histological cell types (undifferentiated secretory and squamous cell types). At the end of the exposure phase, we observed the development of non-hyperplastic areas strongly positive to CK13 antibody, commonly seen in squamous cells as a marker for non-cornified squamous epithelium, thus suggesting a transition of the normal bronchial epithelial cells towards metaplastic cells. The control cultures (clean air exposed and incubator cells) showed no comparable phenotypic changes. In conclusion, our in vitro model presents a valuable tool to study the induction of metaplastic alterations after exposure to airborne material. © 2017.


Rach J.,CULTEX Laboratories GmbH | Budde J.,CULTEX Laboratories GmbH | Mohle N.,CULTEX Laboratories GmbH | Aufderheide M.,CULTEX Laboratories GmbH
Journal of Applied Toxicology | Year: 2014

In toxicology, the strategies for testing the hazardous potential of substances are changing as a result of the ongoing progress in the development of in vitro methods and the demand of the authorities to reduce animal testing. Even in the complex field of inhalation toxicology with its high requirements on the technical implementation and cell culture models, the preconditions for using such methods are fulfilled. We here introduce a sophisticated technique that enables the stable and reproducible exposure of cultivated cells to airborne substances at the air-liquid interface by means of the CULTEX® Radial Flow System (RFS) module. The feasibility and suitability of the experimental setup is demonstrated by dose-response investigations of mainstream cigarette smoke and particulate matter of four substances in different lung epithelial cell lines. A dose-dependent cytotoxcity of the test substances was verified by applying different exposure times. The high reproducibility of the results indicate the reliability of the presented method and recommend the integration of such in vitro approaches in the field of inhalation toxicology by advancing their regulatory validation. © 2013 John Wiley & Sons, Ltd.


Aufderheide M.,Cultex Laboratories GmbH | Halter B.,Halter Engineering GmbH | Mohle N.,Cultex Laboratories GmbH | Hochrainer D.,Von der Hardt 16
BioMed Research International | Year: 2013

The EU Regulation on Registration, Evaluation, Authorization and Restriction of Chemicals (REACH) demands the implementation of alternative methods for analyzing the hazardous effects of chemicals including particulate formulations. In the field of inhalation toxicology, a variety of in vitro models have been developed for such studies. To simulate the in vivo situation, an adequate exposure device is necessary for the direct exposure of cultivated lung cells at the air-liquid interface (ALI). The CULTEX RFS fulfills these requirements and has been optimized for the exposure of cells to atomized suspensions, gases, and volatile compounds as well as micro- and nanosized particles. This study provides information on the construction and functional aspects of the exposure device. By using the Computational Fluid Dynamics (CFD) analysis, the technical design was optimized to realize a stable, reproducible, and homogeneous deposition of particles. The efficiency of the exposure procedure is demonstrated by exposing A549 cells dose dependently to lactose monohydrate, copper(II) sulfate, copper(II) oxide, and micro- and nanoparticles. All copper compounds induced cytotoxic effects, most pronounced for soluble copper(II) sulfate. Micro- and nanosized copper(II) oxide also showed a dose-dependent decrease in the cell viability, whereby the nanosized particles decreased the metabolic activity of the cells more severely. © 2013 Michaela Aufderheide et al.


Aufderheide M.,CULTEX Laboratories GmbH | Scheffler S.,CULTEX Laboratories GmbH | Mohle N.,CULTEX Laboratories GmbH | Halter B.,Halter Engineering GmbH | Hochrainer D.,Vor der Hardt 16
Analytical and Bioanalytical Chemistry | Year: 2011

In the field of inhalation toxicology, progress in the development of in vitro methods and efficient exposure strategies now offers the implementation of cellular-based systems. These can be used to analyze the hazardous potency of airborne substances like gases, particles, and complex mixtures (combustion products). In addition, the regulatory authorities require the integration of such approaches to reduce or replace animal experiments. Although the animal experiment currently still has to provide the last proof of the toxicological potency and classification of a certain compound, in vitro testing is gaining more and more importance in toxicological considerations. This paper gives a brief characterization of the CULTEX® Radial Flow System exposure device, which allows the exposure of cultivated cells as well as bacteria under reproducible and stable conditions for studying cellular and genotoxic effects after the exposure at the air-liquid or air-agar interface, respectively. A commercial bronchial epithelial cell line (16HBE14o-) as well as Salmonella typhimurium tester strains were exposed to smoke of different research and commercial available cigarettes. A dose-dependent reduction of cell viability was found in the case of 16HBE14o- cells; S. typhimurium responded with a dose-dependent induction of revertants. The promising results recommend the integration of cellular studies in the field of inhalation toxicology and their regulatory acceptance by advancing appropriate validation studies. © 2011 The Author(s).


Rach J.,CULTEX Laboratories GmbH | Halter B.,Halter Engineering GmbH | Aufderheide M.,CULTEX Laboratories GmbH
Experimental and Toxicologic Pathology | Year: 2013

The development and validation of new in vitro methods in the field of toxicology have gained more importance in recent years due to stricter guidelines for animal testing, especially in the European Union. Consequently, advances in the construction of technical devices for the exposure of cell or tissue cultures to test substances are necessary. Here, to obtain reliable results, it is important to exclusively use materials that do not interfere with the cell viability. Thus, similar to the biomaterials testing of medical devices which is regulated in the Directive 93/42/EEC, the biocompatibility of the materials has to be verified prior to the construction of such devices. We present here a novel approach for biomaterials testing which allows the quantitative and qualitative assessment of cytotoxicity of material samples. Stainless steel and silicone are often used for laboratory equipment, due to their high chemical, thermal and mechanical resistance. However, our results highlight that some types of silicone may have adverse effects on cultured cells. Moreover, special methods for the surface treatment of metals may also be a critical factor for in vitro devices. Therefore, the testing of all materials coming in contact with cell cultures is highly recommended. © 2013 Elsevier GmbH.


Emura M.,M/A-COM | Aufderheide M.,M/A-COM | Mohr U.,Cultex Laboratories GmbH
Experimental and toxicologic pathology : official journal of the Gesellschaft für Toxikologische Pathologie | Year: 2015

Recent advancement in research on stem/progenitor cells of respiratory organs is breathtaking, benefiting from the rapid development of technology to create transgenic mice. There is now a great deal of knowledge capable of direct translation from mice to humans. Nevertheless, one has to be careful, since there may be unexpected pitfalls. First of all, there are differences anatomically, histologically and ultrastructurally in the airway epithelia of the two species. In parallel with these structural differences, regionally specific cell types behave and function, particularly in regenerative instances, differently between the two species, at least to some extent. From the viewpoint of important human respiratory diseases, one of the most susceptible regions of the respiratory tract is the bronchiole. In our approach to develop in vitro systems utilizing human bronchiolar epithelial cells, we are currently leaning on the data obtained from mouse studies in spite of the above-mentioned species differences. With the help of such in vitro systems we should be able to investigate the damaging effects and mechanisms of environmental pollutants in the human respiratory epithelium and consequently achieve results useful for quantitative analyses of the impact on human respiratory health. While pursuing this goal, the mouse data have suggested that it should be worthwhile to pay close attention to the stem/progenitor cells contained in the human bronchiolar epithelia and eventually make use of them. The mouse data have further shown that these stem/progenitor cells possess a very close association with the immature and variant club cells and the neuroendocrine cells, and our own unpublished preliminary data with human cells are, apparently, at least partly consistent with what the mouse data are telling us. Copyright © 2014 Elsevier GmbH. All rights reserved.


Deschl U.,Boehringer Ingelheim | Vogel J.,Boehringer Ingelheim | Aufderheide M.,Cultex Laboratories GmbH
Experimental and Toxicologic Pathology | Year: 2011

Respiratory diseases like asthma or COPD are gaining more and more importance worldwide due to an increased exposure of humans to inhalable compounds such as cigarette smoke, diesel exhaust or other forms of environmental pollution. Therefore, a high impact on national health systems is expected, meaning long-term treatment, with periodic examinations accompanied by high costs. Although a number of efficient drugs for these disease patterns, like Tiotropium (antimuscarinic), Salmetron (β-antagonist) or corticosteroids, are already available, a great deal of effort has to be put into the development of new drugs and therapy concepts. In this context, in vitro methods may be useful to establish more efficient prescreening procedures to analyze, for example, the toxicity of new compounds during the research and development process. These studies should aim to achieve a better selection of substances relevant for further development and a final reduction in the number of animal experiments. Therefore, we established an in vitro exposure device that allows the analysis of inhalable compounds for their pharmacological and toxicological effects. This CULTEX ® device is composed of an exposure entity representing the in vivo respiratory air compartment and a basal feeding compartment representing the subepithelium. Both compartments are connected by porous transwells on which cells form an epithelium-like cell layer. We have used this system for exposing human lung cells directly to liquid aerosols and present the first data with regard to aerosolized model substances. © 2010.


Scheffler S.,Cultex Laboratories GmbH | Dieken H.,Cultex Laboratories GmbH | Krischenowski O.,Cultex Laboratories GmbH | Aufderheide M.,Cultex Laboratories GmbH
International Journal of Environmental Research and Public Health | Year: 2015

The in vitro toxicological evaluation of e-liquid aerosol is an important aspect of consumer protection, but the cell model is of great significance. Due to its water solubility, e-liquid aerosol is deposited in the conducting zone of the respiratory tract. Therefore, primary normal human bronchial epithelial (NHBE) cells are more suitable for e-liquid aerosol testing than the widely used alveolar cell line A549. Due to their prolonged lifespan, immortalized cell lines derived from primary NHBE cells, exhibiting a comparable in vitro differentiation, might be an alternative for acute toxicity testing. In our study, A549 cells freshly isolated NHBE cells and the immortalized cell line CL-1548 were exposed at the air-liquid interface to e-liquid aerosol and cigarette mainstream smoke in a CULTEX® RFS compact module. The cell viability was analyzed 24 h post-exposure. In comparison with primary NHBE cells, the CL-1548 cell line showed lower sensitivity to e-liquid aerosol but significantly higher sensitivity compared to A549 cells. Therefore, the immortalized cell line CL-1548 is recommended as a tool for the routine testing of e-liquid aerosol and is preferable to A549 cells. © 2015 by the authors; licensee MDPI, Basel, Switzerland.


Nara H.,Japan Tobacco Inc. | Fukano Y.,Japan Tobacco Inc. | Nishino T.,Japan Tobacco Inc. | Aufderheide M.,Cultex Laboratories GmbH
Experimental and Toxicologic Pathology | Year: 2013

For the biological evaluation of cigarette smoke in vitro, the particulate phase (PP) and the gas vapor phase (GVP) of mainstream smoke have usually been collected individually and exposed to biological material such as cultured cells. Using this traditional method, the GVP is collected by bubbling in an aqueous solution such as phosphate-buffered saline (PBS). In such a way the water-insoluble GVP fraction is excluded from the GVP, meaning that the toxic potential of the water-insoluble GVP fraction has hardly been investigated so far. In our experiments we used a direct exposure method to expose cells at the air-liquid interface (ALI) to the water-insoluble GVP fraction for demonstrating its toxicological/biological activity.In order to isolate the water-insoluble GVP fraction from mainstream smoke, the GVP was passed through 6 impingers connected in series with PBS. After direct exposure of Chinese hamster ovary cells (CHO-K1) with the water-insoluble GVP fraction in the CULTEX® system its cytotoxicity was assayed by using the neutral red uptake assay. The water-insoluble GVP fraction was proven to be less cytotoxic than the water-soluble GVP fraction, but showed a significant effect in a dose-dependent manner. The results of this study showed that the direct exposure of cultivated cells at the air-liquid interface offers the possibility to analyze the biological and toxicological activities of all fractions of cigarette smoke including the water-insoluble GVP fraction. © 2012 Elsevier GmbH.

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