Chromosomal aberrations and deoxyribonucleic acid single-strand breaks in adipose-derived stem cells during long-term expansion in vitro.

BACKGROUND AIMS: Adipose-derived stem cells (ASCs) are a promising mesenchymal cell source for tissue engineering approaches. To obtain an adequate cell amount, in vitro expansion of the cells may be required in some cases. To monitor potential contraindications for therapeutic applications in humans, DNA strand breaks and chromosomal aberrations in ASCs during in vitro expansion were examined. METHODS: After isolation of ASC from human lipoaspirates of seven patients, in vitro expansion over 10 passages was performed. Cells from passages 1, 2, 3, 5 and 10 were used for the alkaline single-cell microgel electrophoresis (comet) assay to detect DNA single-strand breaks and alkali labile as well as incomplete excision repair sites. Chromosomal changes were examined by means of the chromosomal aberration test. RESULTS: During in vitro expansion, ASC showed no DNA single-strand breaks in the comet assay. With the chromosomal aberration test, however, a significant increase in chromosomal aberrations were detected. CONCLUSIONS: The study showed that although no DNA fragmentation could be determined, the safety of ASC cannot be ensured with respect to chromosome stability during in vitro expansion. Thus, reliable analyses for detecting ASC populations, which accumulate chromosomal aberrations or even undergo malignant transformation during extensive in vitro expansion, must be implemented as part of the safety evaluation of these cells for stem cell-based therapy.

Dynamic changes of the neurogenic potential in the rat cochlear nucleus during post-natal development.

Neuronal stem cells have been described in the post-natal cochlear nucleus recently. The aim of the study was to analyse the neurogenic potential in the cochlear nucleus from the early post-natal days until adulthood. Cochlear nuclei from Sprague-Dawley rats from post-natal day P3 up to P40 were examined. Neurosphere assays showed persistent neurosphere formation from the early post-natal days until adulthood. The numbers of generated neurospheres were fewer in older ages. Neurospheres were smaller, but displayed the same pattern of neuronal stem cell markers. The markers GFAP, MBP and ß-III Tubulin showed differentiation of dissociated cells from the neurospheres in all cells of the neuronal lineage. BrdU incorporation could be detected, in an age-dependent decrease, in whole-mount experiments of the cochlear nucleus on all examined days. BrdU co-labelled with Atoh1 and ß-III Tubulin. In addition, gene expression and cellular distribution studies of the neuronal stem cell markers displayed an age-dependent reduction in both quantity and numbers. The presented results display a possible neurogenic potential until adulthood in the cochlear nucleus by in vitro and in vivo experiments. The fact that this potential is highest at a critical period of development reveals possible functional importance for the development of the cochlear nucleus and the auditory function. The persistent neurogenic potential displayed until adulthood could be a neurogenic niche in the adult cochlear nucleus, which might be used for potential therapeutic strategies.

Nitrogen dioxide is genotoxic in urban concentrations.

In the discussion on toxic and genotoxic thresholds of air pollutants such as nitrogen dioxide (NO2), realistically low urban concentration ranges are of major interest. For NO2, the WHO defines the annual limit value as corresponding to 0.02 ppm. In the present study, the toxicity and genotoxicity of NO2 is set at a concentration under this limit value and examined in human nasal epithelium at different exposure durations in vitro. Nasal epithelial mucosa samples of 10 donors were harvested during nasal air passage surgery and cultured as an air-liquid interface. Exposure to 0.01 ppm NO2 or synthetic air as a control was performed for 0.5, 1, 2 and 3 h. Analysis included the caspase-3 ELISA, the single cell microgel electrophoresis (comet) assay and the micronucleus assay. The caspase-3 activity was not influenced by NO2 exposure, DNA strand fragmentation correlated with exposure durations to NO2 at 0.01 ppm NO2, and no cytotoxic effects such as apoptosis, necrosis or disturbances of cell proliferation were present. However, micronucleus induction as a sign of genotoxicity at an exposure duration of 3 h could be shown. Shorter exposures did not induce micronucleus formation. In summary, genotoxicity of NO2 could be demonstrated at a common urban concentration in vitro, but a threshold of NO2 genotoxicity could not be defined based on the present experiments.

Isolation and characterization of neural stem cells from the neonatal rat cochlear nucleus.

Neural stem cells have been identified in multiple parts of the postnatal mammalian brain, as well as in the inner ear. No investigation of potential neural stem cells in the cochlear nucleus has yet been performed. The aim of this study was to investigate potential neural stem cells from the cochlear nucleus by neurosphere assay and in histological sections to prove their capacity for self-renewal and for differentiation into progenitor cells and cells of the neuronal lineage. For this purpose, cells of the cochlear nucleus of postnatal day 6 rats were isolated and cultured for generation of primary neurospheres. Spheres were dissociated and cells analyzed for capacity for mitosis and differentiation. Cell division was detected by cell-counting assay and BrdU incorporation. Differentiated neural progenitor cells showed distinct labeling for Nestin and for Atoh1. Positive staining of ß-III Tubulin, glial fibrillary acid protein (GFAP) and myelin basic protein (MBP) showed differentiation into neurons, astrocytes and oligodendrocytes. Furthermore, Nestin- and BrdU-labeled cells could also be detected in histological sections. In conclusion, the isolated cells from the cochlear nucleus presented all the features of neural stem cells: cell division, presence of progenitor cells and differentiation into different cells of the neuronal lineage. The existence of neural stem cells may add to the understanding of developmental features in the cochlear nucleus.

Multipotent stromal cells for autologous cell therapy approaches in the guinea pig model.

Multipotent stromal cells have become of increasing interest due to their potential to provide therapeutic approaches for autologous tissue repair. However, these cells are not well defined in the guinea pig, which represents an important model in hearing research. Adipose-tissue-derived stem cells (ADSC) and bone-marrow-derived stem cells (BMSC) were isolated from different donor sites, and growth curves were generated to judge the proliferation potential. Adipogenic, chondrogenic and osteogenic differentiation was induced and confirmed histologically. Finally, the capability of guinea pig ADSC to differentiate into neuron-like cells was investigated. With regard to the expansion potential, total cell number and doubling time, ADSC from the neck were the most suitable cells of the tested donor sites. Both ADSC and BMSC showed nearly identical behaviour and ability to undergo multilineage differentiation. Thus, we identified ADSC from the neck as a promising cell source for autologous cell-based approaches in hearing research using the guinea pig model.

Demands on caring relatives of head and neck cancer patients.

BACKGROUND: Relatives of cancer patients experience high levels of stress that influence the quality of life of these individuals. To investigate whether there is a necessity for simultaneous supportive care of patient relatives, we performed for the first time a study asking the closest relatives of head and neck cancer patients about their needs during and after the treatment to consider how to optimize the situation for such patient groups. MATERIAL AND METHODS: Patients' relatives were assessed using an anonymous self-report questionnaire that was established in our department by expanding on a questionnaire for cancer patients' relatives from the psycho-oncologic society in Switzerland. The evaluation was multidimensional, cancer specific, and relative based. RESULTS: Relatives feel confronted themselves with cancer, although indirectly. The majority of the respondents were of the opinion that simultaneous psychological care of the patients and for the caring relatives would be helpful to cope with the situation. CONCLUSION: This study shows the significant impact of cancer on caring relatives of head and neck cancer patients. In our opinion, health services should become more aware of this potential to ensure that the needs of the involved patient relatives are met as well as those of the patients.

Genotoxic effects of zinc oxide nanoparticles in nasal mucosa cells are antagonized by titanium dioxide nanoparticles.

Titanium dioxide nanoparticles (TiO2-NPs) and zinc oxide nanoparticles (ZnO-NPs) are often used in sunscreens and other consumer products due to their photoprotective properties. However, concern exists regarding them possibly causing cyto- and genotoxic effects. The aim of this study was to assess cyto- and genotoxicity of these nanomaterials after single or combined exposure. For this purpose, a battery of cell culture test systems for human nasal mucosa (monolayer, air-liquid interface and mini organ culture) were exposed to 0.1-20µg/ml of TiO2- and ZnO-NPs alone and in combination. Cytotoxicity was measured by the MTT assay, and DNA damage and repair capacity were investigated using the comet assay. TiO2-NPs did not exhibit any cyto- or genotoxic potential within the tested concentrations. However, results of the study indicated cyto- and genotoxicity resulting from ZnO-NPs. The genotoxicity could be antagonized by TiO2-NPs. Furthermore, the DNA repair capacity after ZnO-NP-induced DNA damage was enhanced by TiO2-NPs. The adsorption of dissolved zinc ions onto TiO2-NPs is discussed as the major antagonistic mechanism. The combination of both metal oxide nanoparticles interferes with the genotoxicity of ZnO-NPs and should be discussed as a reasonable and safe alternative to the sole use of ZnO-NPs in consumer products.

Effects of salinomycin and CGP37157 on head and neck squamous cell carcinoma cell lines in vitro.

Surgery, radiation, chemotherapy or a combinations of these are all accepted modalities for the treatment of head and neck squamous cell carcinoma (HNSCC). Despite this, 40­60% of patients suffering from HNSCC develop loco­regional failure and/or distant metastases. Salinomycin has been demonstrated to be >100­fold more effective than paclitaxel at causing cancer stem cell death, therefore, it may offer an important improvement in cancer therapy. However, the toxicity of salinomycin is of concern. A possible solution may be the administration of additive drugs, which reduce the toxicity. By inhibiting the mitochondrial Na+/Ca2+ exchanger using the benzodiazepine derivate, CGP37157 (CGP), a significant reduction in salinomycin neuronal toxicity has been observed. This raises the question of whether CGP also inhibits the tumor toxicity of salinomycin. In the present study, the FaDu and HLaC79 C1 HNSCC cell lines were treated with salinomycin with or without CGP. Comparative viability assessments were performed using microscopy, a fluorescein diacetate assay, an MTT assay, a clonogenic assay and annexin V­propidium iodide staining. The expression levels of MDR­1 were monitored using reverse transcription­quantitative polymerase chain reaction. Salinomycin alone, and in combination with CGP, achieved a significant attenuation of cell viability and increased apoptosis in a dose­dependent manner. However, the tumor toxicity of salinomycin was not inhibited by CGP. The HLaC79 C1 cells were more sensitive to salinomycin, compared with the FaDu cells, with this sensitivity being due to high expression levels of MDR­1 by the HLaC79 C1 cells. In conclusion, CGP did not counteract the tumor toxicity of salinomycin in vitro and may be a promising drug in future anticancer therapy. The results of the present study encourages further investigation of the toxicological aspects of salinomycin, particularly in human cells and animal models.

A hydrogel coating for cochlear implant arrays with encapsulated adipose-derived stem cells allows brain-derived neurotrophic factor delivery.

CONCLUSION: Human adipose-derived stem cells (ASCs), encapsulated in a fibrin-collagen hydrogel for the coating of an electrode array, produce sufficient amounts of neurotrophic factors and may be suitable for enhancing the bioelectric interface of cochlear implants (CIs). OBJECTIVES: To evaluate different hydrogel compositions loaded with ASCs with regard to delivery of neuroactive substances and mechanical suitability for the coating of a CI electrode array. METHODS: ASCs were cultivated in hydrogels consisting of collagen and fibrin in varying fractions (0:1, 1:1, 1:2, and 1:0). The cell proliferation and viability, as well as the production of brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), and laminin were determined. Two hydrogel compositions were used as a coating for CI electrode arrays and tested in a scala tympani model. RESULTS: Cell proliferation was best in collagen/fibrin hydrogel compositions (1:1 and 1:2) and increasing amounts of BDNF (up to 2.59 ng/ml) and laminin (up to 320 ng/ml) were detected. GDNF production was inconsistent and markedly lower. A sufficient coating of a CI electrode carrier in terms of stability and flexibility was achieved only with mixed compositions, although hydrogels formed bulky and uneven layers on the silicone surfaces.

Nanoparticle-induced photocatalytic head and neck squamous cell carcinoma cell death is associated with autophagy.

AIM: To characterize molecular mechanisms underlying photocatalytic cell death of head and neck squamous cell carcinoma (HNSCC) by zinc oxide nanoparticles (ZnO-NPs). MATERIALS & METHODS: Human HNSCC-derived FaDu cells were incubated with ZnO-NPs followed by UVA-1 irradiation. Cytotoxicity was assessed by MTT assay and annexin-V propidium iodide test. Autophagy was detected by autophagosome accumulation, conversion of light chain 3 I to II, and lysosomal activity. The generation of reactive oxygen species was measured using the 2',7'-dichlorofluorescein-diacetate test. RESULTS: Apoptosis-independent cytotoxic effects were induced by 0.2- and 2-µg/ml ZnO-NPs and UVA-1. FaDu cells promoted autophagosome formation. Significantly elevated light chain 3 II and reactive oxygen species were seen after the combined application of both ZnO-NPs and UVA-1 as photocatalytic treatment. Autophagy probably mediates cell survival under UVA-1 or ZnO-NP exposure alone but induces self-digestive cell death after combined treatment. CONCLUSION: The effect of autophagy on HNSCC viability after nanoparticle-induced photocatalytic treatment seems to depend on the impact of the physicochemical trigger.

Nicotine derived genotoxic effects in human primary parotid gland cells as assessed in vitro by comet assay, cytokinesis-block micronucleus test and chromosome aberrations test.

Genotoxic effects of nicotine were described in different human cells including salivary gland cells. Based on the high nicotine concentration in saliva of smokers or patients using therapeutic nicotine patches, the current study was performed to evaluate the genotoxic potential of nicotine in human salivary gland cells. Therefore, primary salivary gland cells from 10 patients undergoing parotid gland surgery were exposed to nicotine concentrations between 1 µM and 1000 µM for 1 h in the absence of exogenous metabolic activation. The acinar phenotype was proven by immunofluorescent staining of alpha-amylase. Genotoxic effects were evaluated using the Comet assay, the micronucleus test and the chromosome aberration test. Cytotoxicity and apoptosis were determined by trypan blue exclusion test and Caspase-3 assay. Nicotine was able to induce genotoxic effects in all three assays. The chromosome aberration test was the most sensitive and increases in numerical and structural (chromatid-type and chromosome-type) aberrations were seen at ≥1 µM, whereas increases in micronuclei frequency were detected at 10 µM and DNA damage as measured in the Comet assay was noted at >100 µM. No cytotoxic damage or influence of apoptosis could be demonstrated. Nicotine as a possible risk factor for tumor initiation in salivary glands is still discussed controversially. Our results demonstrated the potential of nicotine to induce genotoxic effects in salivary gland cells. These results were observed at saliva nicotine levels similar to those found after oral or transdermal exposure to nicotine and suggest the necessity of careful monitoring of the use of nicotine in humans.

Functional responses of human adipose tissue-derived mesenchymal stem cells to metal oxide nanoparticles in vitro.

Nanoparticles (NPs) are frequently applied in biomedical applications. The use of human mesenchymal stem cells (hMSC) in biomedicine is pivotal, especially in oncology and tissue engineering. Titanium dioxide (TiO2) and zinc oxide (ZnO) NPs are interesting agents in experimental oncology and stem cells are discussed to be a potential vehicle for NPs to tumor sites. However, little is known about hazardous effects of NPs in hMSC. The aim of the present study was to analyze functional impairment of hMSC by ZnO- and TiO2-NPs. Cytotoxic effects of NPs were evaluated by the MTT-assay. Furthermore, multi-differentiation capacity, spheroid formation and migration were assessed. The immunophenotype was observed by flow cytometry. Cytotoxic effects were observed at 625 nM ZnO-NPs whereas no cytotoxicity was seen in hMSC by TiO2-NPs. The differentiation capacity of hMSC into osteogenic and adipose lineages was unchanged. A long-term period cultivation of hMSC for 3 weeks after NP exposure revealed a persistence of NPs in the cytoplasm. The migration capability was impaired whereas the ability to form spheroids was not affected. Flow cytometric analysis revealed distinct alteration of cell surface markers CD 90 and CD 73. Major functional properties of hMSC were unaffected by TiO2- and ZnO-NPs. However, restricted migration might critically influence wound healing capacity. Further information is needed to assess the clinical impact of these findings.

Intraoperative monitoring using cochlear microphonics in cochlear implant patients with residual hearing.

OBJECTIVES: To describe a technique for the intraoperative monitoring of residual hearing during a cochlear implant (CI) procedure and, thus, to identify the time of occurrence and surgical steps leading to hearing loss. STUDY DESIGN: Prospective audiologic analysis in a patient series. SETTING: Tertiary referral center. SUBJECTS: Patients with residual hearing subjected to cochlear implantation. INTERVENTIONS: During cochlear implantation, cochlear microphonics (CMs) in response to frequency-specific stimuli were recorded in patients with residual hearing. Thresholds were determined before and after opening of the cochlea, with a limited portion of the electrode array inserted, and after full insertion. RESULTS: Monitoring of the hearing state using CMs was quick, reliable, and capable of detecting an intracochlear trauma. In a first series of patients, thresholds were preserved in all patients after opening of the cochlea. Thresholds were preserved in 5 of 6 patients after limited insertion and half of the patients after full insertion of the electrode array. Despite threshold preservation until the end of surgery, the residual hearing was lost in patients with deep insertions 1 week postoperative. CONCLUSION: Intraoperative monitoring of CM thresholds may be valuable for identifying the exact point of time at which residual hearing is affected in CI patients. Opening of the cochlea itself seems to be unrelated to hearing loss. A significant proportion of patients may have hearing loss caused by secondary effects rather than a direct trauma.

Antitumor activity of photo-stimulated zinc oxide nanoparticles combined with paclitaxel or cisplatin in HNSCC cell lines.

Zinc oxide nanoparticles (ZnO-NPs) exhibit photocatalytic properties and are used in sunscreen cosmetics and for the degradation of environmental pollutants. Furthermore, ZnO-NPs have proven to induce tumor-selective cell death in human squamous cell carcinoma (HNSCC) in vitro. The aim of the current study was to evaluate cytotoxic effects of UVA-1-activated ZnO-NPs in combination with paclitaxel and cisplatin in HNSCC. Three HNSCC cell lines (HLaC-78, Cal-27 and PJ-41) were incubated with ZnO-NPs at concentrations of 0.02 and 0.2 µg/ml in combination with paclitaxel (0.5-5 nM) or cisplatin (0.03-3 µM) for 24 h. Afterwards, cells were irradiated with UVA-1 for 15 min. Viability was measured by MTT assay, fluorescein-diacetate (FDA) test and annexin/propidiumiodide flow cytometry. A significant decrease in viable cells could be observed in all three HNSCC cell lines treated by photocatalytic therapy with 0.2 µg/cm(2) ZnO-NPs alone. A combination with paclitaxel or cisplatin at low concentrations resulted in a further increase in cytotoxicity in vitro revealing a synergistic effect. Flow cytometry revealed a combination of apoptosis and necrosis. These results indicate that photocatalytic therapy of HNSCC with ZnO-NPs could enhance the cytotoxic action of chemotherapeutic agents synergistically, indicating a promising potential for ZnO-NPs in antitumor applications.

Nanosized titanium dioxide particles do not induce DNA damage in human peripheral blood lymphocytes.

Industrial application of titanium dioxide nanoparticles (TiO(2) -NPs) as an additive in pharmaceutical and cosmetic products is increasing. However, the knowledge about the toxicity of this material is still incomplete and data concerning health and environmental safety and results of recent studies on TiO(2) nanotoxicology are inconsistent. The in vitro geno- and cytotoxicity of TiO(2) -NPs in the anatase crystal phase was evaluated in human peripheral blood lymphocytes from 10 male donors. Initially, transmission electron microscopy (TEM) was performed to describe particle morphology and size, the degree of particle aggregation, and the intracellular distribution. Cells were exposed to nanoparticles in increasing concentrations of 20, 50, 100, and 200 µg/ml for 24 hr. Cytotoxic effects were analyzed by trypan blue exclusion test and the single-cell microgel electrophoresis (comet) assay was applied to detect DNA double-strand breakage. TiO(2) -NPs were sphere shaped with a diameter of 15-30 nm. Despite dispersive pretreatment, a strong tendency to form aggregates was observed. Particles were detected in the cytoplasm of lymphocytes, but also a transfer into the nucleus was seen. The trypan blue exclusion test did not show any decrease in lymphocyte viability, and there was no evidence of genotoxicity in the comet assay for any of the tested concentrations. In conclusion, TiO(2) -NPs reached the cytoplasm as well as the nucleus and did not induce cyto- or genotoxic effects in human peripheral blood lymphocytes. Complement investigations on different human cell systems will be performed to estimate the biocompatibility of TiO(2) -NPs. Environ. Mol. Mutagen.

Intraoperative monitoring of active middle ear implant function in patients with normal and pathologic middle ears.

OBJECTIVE: To describe a technique enabling the intraoperative assessment of the mechanical coupling of active middle ear implants in patients with normal or dysfunctional middle ears. PATIENTS: Patients with sensorineural or mixed hearing loss subjected to revision surgery of an active middle ear implant. INTERVENTION: Recording of compound action potentials (CAPs) of the auditory nerve in response to stimulation via an active middle ear implant during revision surgery. MAIN OUTCOME MEASURES: CAP thresholds as a measure of mechanical coupling and system integrity. CONCLUSION: Determining CAP thresholds may be suitable for identifying a proper mechanical coupling of active middle ear implants in patients with a dysfunctional middle ear, even if the ossicular chain is disrupted.

The effect of wound fluid on adipose-derived stem cells in vitro: a study in human cell materials.

After surgery, wound healing begins with a well-orchestrated integration of several cytokines, cells, and extracellular matrix. Some studies show an involvement of stem cells in wound healing. However, little is known about the mechanism that leads to the migration of stem cells. Wound fluid (WF) with its cytokines may play an important role. We investigated in the present study the in vitro effects of WF on adipose-derived stem cells (ADSCs). Survival, proliferation, structural integrity, changes in the multidifferentiation potential, and surface markers (cluster of differentiation [CD] 105, CD73, CD90) of ADSCs after cultivation with WF was analyzed. Further, the migration effect of WF on ADSCs was evaluated. The proliferation rate and the migration potential of ADSCs were enhanced significantly by cultivation with WF. There was also a change in the quantity of surface markers after cultivation with WF. In conclusion, in vitro expansion of stem cells with WF proved possible. WF and its cytokines could represent one primary reason for the migration of stem cells toward the wound. Future investigation is warranted to clarify the significance of the shift in surface markers.

Growth behavior of cochlear nucleus neuronal cells on semiconductor substrates.

Auditory brainstem implants provide sound information by direct stimulation of the cochlear nucleus to patients with dysfunctional or absent cranial nerve VIII. In contrast to patients with cochlear implants, the use of the auditory brainstem implants is less successful. This cannot be fully explained by the difference location of stimulation but a rather unspecific neuronal stimulation. The aim of this study was to further examine neuronal cells of the cochlear nucleus and to test their interactions with semiconductor substrates as a potential electrode material for improved auditory brainstem implants. The cochlear nuclei of postnatal day 7 rats were microsurgically dissected. The tissue was dissociated enzymatically and plated on coverslips as control and on the semiconductor substrates silicon or silicon nitride. After 4 days in culture the morphology and growth of dissociated cells was determined by fluorescence and scanning electron microscopy. Dissociated cells of the cochlear nucleus showed reduced cell growth on semiconductor substrates compared with controls. SEM analysis demonstrated close contact of neurons with supporting cells in culture and good adherence of neuronal growth cones on the used materials. These findings present basic knowledge for the development of neuron-electrode interfaces for future auditory brainstem implants.

BMSC enhance the survival of paclitaxel treated squamous cell carcinoma cells in vitro.

The 5-year survival rate of patients suffering from head and neck squamous cell carcinoma (HNSCC) is unsatisfying despite the advances in carcinoma treatment. Recent studies suggest that stem cells can be used as a gene therapy carrier for cancer treatment. Stem cells produce different cytokines such as growth factors in a paracrine manner and cancer cells may show drug resistance in the presence of such growth factors. Reports in the literature concerning treatment of cancer using bone marrow derived stem cells (BMSC) are controversial, which led us to investigate the effects of paclitaxel on human HNSCC cell lines (FaDu and HLaC 78) cultivated simultaneously with BMSC in a transwell system (co-culture). Co-culture and HNSCC cell lines were treated with 10nM of paclitaxel for 24h. Morphology, viability and apoptosis were measured by microscopy, the MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay, and the Annexin V-propidium iodide test. The survival of HNSCC cell lines treated with paclitaxel in co-culture increased significantly compared to control cells. Apoptosis of HNSCC cell lines in co-culture was attenuated significantly. In conclusion, BMSC increase HNSCC resistance to treatment with paclitaxel in vitro. Tumor-stroma interactions are critical components of tumor biology including tumor invasion and metastatic potential. Therefore particular attention must be paid to the complex tumor-stroma interactions to fully understand how tumor cells become chemoresistant.

Cytotoxic, genotoxic and pro-inflammatory effects of zinc oxide nanoparticles in human nasal mucosa cells in vitro.

Despite increasing application of zinc oxide nanoparticles (ZnO-NPs) for industrial purposes, data about potential toxic properties is contradictory. The current study focused on the cyto- and genotoxicity of ZnO-NPs in comparison to ZnO powder in primary human nasal mucosa cells cultured in the air-liquid interface. Additionally, IL-8 secretion as a marker for pro-inflammatory effects was measured. Particle morphology and intracellular distribution were evaluated by transmission electron microscopy (TEM). ZnO-NPs were transferred into the cytoplasm in 10% of the cells, whereas an intranuclear distribution could only be observed in 1.5%. While no cyto- or genotoxicity could be seen for ZnO powder in the dimethylthiazolyl-diphenyl-tetrazolium-bromide (MTT) test, the trypan blue exclusion test, and the single-cell microgel electrophoresis (comet) assay, cytotoxic effects were shown at a ZnO-NP concentration of 50 µg/ml (P<0.01). A significant enhancement in DNA damage was observed starting from ZnO-NP concentrations of 10 µg/ml (P<0.05) in comparison to the control. IL-8 secretion into the basolateral culture medium was increased at ZnO-NP concentrations of 5 µg/ml (P<0.05), as shown by ELISA. Our data indicates cyto- and genotoxic properties as well as a pro-inflammatory potential of ZnO-NPs in nasal mucosa cells. Thus, caution should be taken concerning their industrial and dermatological application. Additionally, further investigation on repetitive NP exposure is needed to estimate the impact of repair mechanisms.