They found negative net precipitation rates of −1.1 and of −3.5 mm day−1 for the WMB and EMB, respectively. Mariotti et al. (2002) estimated different evaporation and precipitation rates using different datasets, and found that the SB203580 Mediterranean Sea had negative net precipitation rates ranging from −1.3 to −1.9 mm day−1, most markedly over EMB. The present calculations (see Table 5) and those presented in these three earlier studies thus differ only slightly. The water balance of the Mediterranean Sea was controlled

by net flow through the Gibraltar Strait and Sicily Channel, net precipitation rates, and freshwater input. The heat balance of the Mediterranean Sea was controlled by heat loss from the water surface, solar radiation into the sea, and heat flow through the Gibraltar Strait and Sicily Channel. Both heat loss and solar radiation display significant (insignificant) trends over the EMB (EMB). This agrees with the previous findings of Shaltout and Omstedt (2012). The annual net heat gain from the WMB (−13 W m−2) was balanced by the heat flow through the Gibraltar Strait and Sicily Channel. The annual net heat loss from the EMB (11 W m−2) was balanced

Ipatasertib by the heat flow through the Sicily Channel. This research was undertaken when Dr. Mohamed Shaltout was a visiting scientist at the Ocean Climate Group, Department of Earth Sciences, University of Gothenburg, Sweden. The work is a contribution to the GEWEX/BALTEX phase II and the newly formed programme “Baltic Earth-Earth System Science for the Baltic Sea region” and the HyMex program. “
“Toxic algal blooms are of a particular concern in eutrophic aquatic Amobarbital ecosystems, where natural or anthropogenically induced nutrient enrichment leads to enhanced algae and cyanobacteria biomass (Sutcliffe and Jones, 1992). About 300 microalgae species were reported as forming so-called algal blooms. Nearly one fourth of these species have a potential to produce

toxic compounds (Hallegraeff et al., 2003). Some of algal toxins may bioaccumulate in aquatic organisms and be transferred through a food chain, reaching critically high concentrations at higher trophic levels (Cazenave et al., 2005, Ferrão-Filho and Kozlowski-Suzuki, 2011, Landsberg, 2002 and Rhodes et al., 2001). Due to the wide toxicological effects of these compounds, including neurotoxicity, hepatoxicity, cytotoxicity and dermatoxicity, there is a risk of health hazard for humans, domestic animals and wildlife related to the toxic algal blooms in aquatic ecosystems (Carmichael, 2001, Kujbida et al., 2006 and Van Dolah, 2000). Among the toxins produced by cyanobacteria microcystins (hepatotoxins) are probably the most hazardous ones in terms of impact on human health (Carmichael, 1994, Chorus and Bartram, 1999 and Funari and Testai, 2008). Microcystins (MC) are very stable (Jones and Orr, 1994 and Tsuji et al., 1994), not destroyed by the common water treatment methods (Keijola et al.

In addition, although the original Report used and recommended the symbols NAD and NADH2 for the oxidized and reduced forms respectively of the coenzyme, they also suggested NAD+ and NADH respectively as alternatives. This latter system has the advantage that it allows the plain symbol NAD to refer to the two forms collectively, but it has the disadvantage that it assigns a+superscript to what

is in reality an anion. In practice the system with NAD+ and NADH has become overwhelmingly the most used, and when it became adopted in Enzyme Nomenclature there was a feeling that the equation looked unbalanced with unequal charges on the left and right-hand sides. In what Alberty in particular

considered as a misguided move, this was then “corrected” by including protons in equations. A suggested way to avoid Selleckchem KU 55933 the problem (Alberty and Cornish-Bowden, 1993), in which the two forms of coenzyme were to be written as NADox and NADred has received no significant adoption in the literature. click here As the entry for acetate kinase considered above is one of the simpler examples, with no comments or specificity information (with the implication that the enzyme catalyses that one reaction only) it is useful to examine a more typical entry: EC 2.7.1.1 Accepted name: hexokinase Reaction: ATP+d-hexose=ADP+d-hexose 6-phosphate Other name(s): hexokinase type IV, glucokinase; IMP dehydrogenase hexokinase d; hexokinase type IV; hexokinase (phosphorylating); ATP-dependent hexokinase; glucose ATP phosphotransferase Comments: d-Glucose, d-mannose, d-fructose, sorbitol and d-glucosamine

can act as acceptors; ITP and dATP can act as donors. The liver isoenzyme has sometimes been called glucokinase. Systematic name: ATP:d-hexose 6-phosphotransferase Links to other databases: BRENDA, EXPASY, GTD, IUBMB, KEGG, METACYC, PDB, UM-BBD, CAS registry number: 9001-51-8 References: 1. Bailey, K. andWebb, E.C. Purification of yeast hexokinase and its reaction with ββ′-dichlorodiethyl sulphide. Biochem. J.42 (1948) 60–68. [PMID: 16748250]. 2. Berger, L., Slein, M.W., Colowick, S.P. and Cori, C.F. Isolation of hexokinase from baker׳s yeast. J. Gen. Physiol.29 (1946) 379–391. 3. Kunitz, M. and McDonald, M.R. Crystalline hexokinase (heterophosphatase). Method of isolation and properties. J. Gen. Physiol.29 (1946) 393–412. 4. Pollard-Knight, D. and Cornish-Bowden, A. Mechanism of liver glucokinase. Mol. Cell. Biochem. 44 (1982) 71–80. [PMID: 7048063]. 5. Ureta, T., Radojković, J., Lagos, R., Guixé, V. and Núñez, L. Phylogenetic and ontogenetic studies of glucose phosphorylating isozymes of vertebrates. Arch. Biol. Med. Exp.12 (1979) 587–604. [PMID: 233226]. 6. Cárdenas, M.L., Rabajille, E. and Niemeyer, H. Fructose: A good substrate for rat-liver ‘glucokinase’ (hexokinase d). Biochem. J. 222 (1984) 363–370.

Thus, the design of novel behavioral test methods started, with the notion that such techniques may enable one to tap into functional alterations induced in the brain of the zebrafish by a variety of ways including mutagenesis or pharmaceutical approaches [9]. The past 15 years has seen a rapid expansion PD0332991 ic50 of this research, an exponential increase of the number of scientific publications in which zebrafish behavior has been the focus [10••]. Notably, the continuous increase of

the number of these publications outpaced behavioral papers on even the most preferred model organisms of biomedical research, the rat and the mouse. In this short review, I will discuss some of the latest developments of this expanding field focusing on a few behavioral methods designed for the zebrafish. I also briefly mention some of the

latest developments in forward genetics as they pertain to the zebrafish. Admittedly, this short and somewhat biased review is far from being exhaustive. Instead, it attempts to illustrate what its author thinks are perhaps the most important issues and advances in the current state of Histone Methyltransferase inhibitor the art of zebrafish phenomics with experimental examples mostly drawn from his laboratory. Fish represent the most species-rich group among vertebrates [11]. Many fish species have been studied from a behavioral Lonafarnib datasheet perspective, but learning from mistakes of the past [12], zebrafish scientists realized that perhaps the best way to design behavioral test paradigms for this newcomer is to try to understand its species-specific features, its ecology and its behavior in nature. Keeping this in mind, a number of

successful behavioral paradigms have been developed that now allows one to test numerous behavioral responses and features of the zebrafish from its cognitive and mnemonic characteristics [13], through fear and anxiety 14 and 15, to social behavior 16 and 17, to mention but a few. Perhaps one of the most important species-specific features of the zebrafish, at least from a behavioral experimentation perspective, is that it is diurnal, that is, active during the day. It has excellent vision and thus visual cues may be employed in the behavioral tests developed for it. This is an important difference compared to laboratory rodents, the rat and the mouse, which are nocturnal species. Although numerous behavioral tasks have been developed for rodents that utilize visual cues [12], the appropriateness of these cues has been debated, and the question of whether one can properly study behavior of these nocturnal animals during their subjective day when they are supposed to be asleep or inactive has been raised. The diurnal zebrafish does not suffer from these controversies.

It is, however, notable that NAPs selleck products by themselves do not exert as much influence as the BoNT/A complex, except for IL-6 which showed equal response (Table 1). MCP-1 and VEGF were two other cytokines which were induced by NAPs alone, albeit not as strongly as BoNT/A complex. BoNT/A complex and NAPs both contain associated proteins for BoNT/A. However, exposition to BoNT/A complex, but not to NAPs,

resulted in significant increase of IP-10, IL-8, IL-15, TNF-α, and RANTES. For the current research, cytokine release was examined after 48 h of incubation. The kinetics of cytokine release have been studied for 24 h to up to one week in lymphocyte (Arva and Andersson, 1999) and kinetics of TNF, IL-6, and IL-8 gene expression after inflammatory stimuli MK-2206 solubility dmso have been shown to have multiple peak at 2–4 h and 24 h (DeForge and Remick, 1991). Although no cytokine release was induced by pure BoNT/A in the current experimental setting, further investigation with different incubation time on complex patterns of cytokine gene expression and production with pure BoNT/A as well as other components of BoNT/A complex is needed. Higher effect of BoNT/A

complex could arise from one or more of the following reasons. One, there is higher level of binding of the BoNT/A in the BoNT/A complex allowing more NAPs to enter the cell. Two, interaction between BoNT/A and NAPs introduce conformational changes which are more critical for triggering cytokine response. Three, there is a physiological link between the effects of BoNT/A and NAPs intracellularly, leading to synergistic host cell response. A previous study on the co-culture of microglia and SH-SY5Y

cells has shown the expression of IL-6, IL-8, and MCP-1 with borrelia burgdorferi stimulation, a spirochete that causes lyme disease, and it is known to potently induce the production of inflammatory mediators in a variety of cells (Myers et al., 2009). Release of MCP-1 from SH-SY5Y has also been reported during the neuroinflammation process (Mitchell et al., 2009). Physiological Celastrol role of cytokine release in neuronal cells can be manifold. The presence and activity of pro-inflammatory cytokines IL-1β and TNF-α were first reported in human and rat brain a decade ago (Breder et al., 1988 and Plata-Salaman et al., 1988). Cytokine release studies enable us to identify cytokines that are produced specifically upon BoNT/A, its complex, or NAPs stimulation. The SH-SY5Y cell line has been proven to be a useful in vitro model for TNF production from neurons and the regulation of that production by alpha2-adrenergic receptor activation (Renauld and Spengler, 2002). Additionally, TNF-α has been shown not only play the critical roles in pathological development and inflammatory induction, but on modulating cell proliferation of neural progenitors in CNS inflammation (Downen et al., 1999 and Wu et al., 2000).

This study aimed to contribute to debates related to European aquaculture development as well as to environmental justice literature by analyzing existing finfish aquaculture conflicts in Europe and by linking them

to the policy level. It underlines that while establishing new strategies for European aquaculture, the focus should not be solely on economic growth, but rather on ecologically, socially and economically sustainable and just development of marine aquaculture. Integration of economic, social and ecological concerns into national and regional aquaculture strategy plans proves to be potentially challenging but necessary in order to ensure social acceptance of fish farms and to control the impacts of new and already existing ones. The article concludes by emphasizing the significance of marine Etoposide manufacturer finfish aquaculture conflicts in Europe and the lessons to be learned in terms of their policy implications. An effective participatory decision-making mechanism should be designed that takes the views and perceptions of all relevant actors into account in order to determine whether or not to construct fish farms; and if yes, where to build them and how many. Best practices safeguarding environmental justice such as the establishment of inclusive decision-making mechanisms, ensuring

access to transparent information Luminespib concentration and an equitable social distribution of burdens, benefits and risks resulting from aquaculture activities should be further investigated and incorporated into future policies. Research for this paper benefited from EC funding under the Marie Curie Actions – Initial

Immune system Training Networks – FP7 – PEOPLE – 2011; Contract no. 289374 – “ENTITLE”. The research would not have been made possible without the support of interviewees who kindly shared their opinions and knowledge. The authors especially desire to acknowledge Seas at Risk network for facilitating contact and the valuable comments and efforts of Begüm Özkaynak, Pınar Ertör Akyazı, Santiago Gorostiza Langa, Melissa Garcia Lamarca and Marien González Hidalgo. “
“The European Marine Strategy Framework Directive (MSFD, 2008/56/EC) aims to achieve and/or maintain Good Environmental Status (GES) of EU marine waters by 2020. The Directive defines GES as: “The environmental status of marine waters where these provide ecologically diverse and dynamic oceans and seas which are clean, healthy and productive” (MSFD Article 3). GES is described by a comprehensive set of 11 qualitative descriptors. Descriptor 5 relates specifically to eutrophication and states that the human-induced eutrophication should be minimized. One of the first steps that had to be finished until July 2012 was the initial assessment of Member States׳ marine waters (Art. 8 MSFD), the determination of GES (Art. 9 MSFD) and the establishment of environmental targets and associated indicators to achieve GES (Art. 10 MSFD).

5 ml) was fed to the rats with the help of a feeding needle. The yield of aqueous curry leaf extract (Cu LE) was 14.72 ± 0.36% (w/w). Male Wistar rats of body weight 160-180 g were used throughout the experiments. The animals were handled as per the

guidelines of institutional animal ethics committee (IAEC) of Department of Physiology, University of Calcutta in accordance with the committee for the purpose of control and supervision of experiment on animals (CPCSEA), Ministry of Environment and Forest, Government of India. All the experimental protocols had the approval (approved under proposal No. IAEC/PROPOSAL/DB-5/2010 dated 05/05/2010, approval date: 16/11/2011) of Institutional Animal Ethics Committee find more (IAEC) of the Department of Physiology, University buy RGFP966 of Calcutta. Prof. P. K. Samanta, M. Sc. (Vet.), Ph. D., Professor and Veterinary Surgeon and CPCSEA Nominee to Department of Physiology, University of Calcutta, acted as the advisor for animal care and handling. For our present study, the animals were housed in galvanized wire cages, in well ventilated, air conditioned rooms of our animal house with 12 hours light/dark cycle, at about 18 °C room temperature for 7 days to get adapted to laboratory condition. All rats had been given a standard diet

containing 18% protein, 71% carbohydrate and vitamins which are considered to be an adequate (normal) dietary protein level [12]. The animals were released from

quarantine and immediately they were kept on fasting condition in specially designed cages for the following 40 hours. After that treatment of rats was carried out as per the schedule mentioned below. The animals were divided into six groups as follows for the dose response study: GROUP I: Control group (C). Rats were allowed to drink water supplied ad libitum. GROUP II: Piroxicam treated group (Px). Rats were orally administered piroxicam at a dose of these 30 mg/kg body weight dose with a feeding needle. The treatment was carried out immediately after 40 hours fasting. GROUP III: Cu LE pre-treated at a dose of 50 mg/kg body weight and piroxicam fed group (Cu LE1). Cu LE was administered at 50 mg/kg body weight at the onset of the experiment and immediately after one hour, the animals were orally fed piroxicam at 30 mg/kg body weight. GROUP IV: Cu LE pre-treated at a dose of 100 mg/kg body weight and piroxicam fed group (Cu LE2). Curry leaf aqueous extract was administered at 100 mg/kg bodyweight at the onset of the experiments and immediately after one hour, the animals were orally fed piroxicam at 30 mg/kg body weight. GROUP V: Cu LE pre-treated at a dose of 200 mg/kg body weight and piroxicam fed group (Cu LE3).

Through a series of downstream effects, the Rho GTPases inhibit NO production in the endothelium by decreasing eNOS expression and activity [4]. Less eNOS activity results in a paucity of the vasodilatory effects of NO. In essence, the Rho GTPases causes arteriolar vasoconstriction. By inhibiting their production, statins “turn off the off switch” and thus promote eNOS activity and vasodilation. Based on this theory, the expected result of statin administration would be increased cerebral blood flow with statins, which has been shown in healthy subjects [3]. In radiation vasculopathy, however, the situation is quite different from healthy controls in that the affected hemisphere has diffuse

arteriolar vasculopathy, while the contralateral hemisphere is spared this pathology. We suggest that the contradictory selleckchem finding we observed is a special result of the rule of vasodilation and an example of Caspase phosphorylation cerebral steal. At baseline, we hypothesize that there is maximal vasodilation in the pathological hemisphere. The addition of the statin could not

improve the already maximally dilated vessels on the pathological hemisphere. Statins and the addition of eNOS activity would, however, increase vessel dilation to the healthy hemisphere, shifting it from a neutral amount of vessel tone to a dilated state. This shift to a more dilated stated on the healthy hemisphere seems to exacerbate the underlying hemodynamic inequity Megestrol Acetate and causes a steal syndrome. By withdraw of statin, the healthy hemisphere tone returns to normal, and the pathological hemisphere was able to shunt more flow and improve. In healthy subjects or those with diffuse bihemispheric cerebrovascular disease, the addition of statins will likely improve vasomotor tone and augment cerebral blood flow. In the special circumstance of a patient with isolated high-grade cerebrovascular disease

in a single vascular bed, the addition of a statin may lead to a cerebral steal phenomenon. “
“The disturbance of cerebrospinal fluid (CSF) circulation in system of CSF pathways owing to the various reasons (impairment of CSF production and absorption, the mechanical block) can cause development of hydrocephalus. Depending on compensating capabilities of brain this pathology may not have clinical symptoms or, being accompanied by increase of intracranial pressure (ICP), can give a clinical picture of intracranial hypertension (ICH) syndrome. In the latter case carrying out surgical treatment – correction of the disturbed CSF circulation by means of shunting or endoscopic intervention – is required. Nevertheless in some cases, when ICH is doubtful or has temporal character, the data of clinical examination, computed tomography/magnetic resonance scanning of the brain appear insufficient for defining indications for operations.

This work was sponsored by Consejo Nacional de Ciencia y Tecnología, México (CONACYT) No. 111941 and Genzyme Corp (now Sanofi). “
“The authorship for the article in Archives of Medical Research 44 (2013) 21-26 should read as follows: Mohamed Kamel Sabry, Mohamed Nazmy Farres, Nermine Abdelnour Melek, Naglaa Ahmed Arafa, and Annie Arek

Ohanessian. We apologize for any confusion or inconvenience this may have caused. “
“1. Kan Saito Division of Pediatric Dentistry, Department Proteases inhibitor of Oral Health and Development Sciences, Tohoku University Graduate School of Dentistry The influence of Sox21 as a novel ameloblast marker on tooth germ differentiation” 2. Hiroyuki Nakamura Nakamura Orthodontic and Pediatric Dental Office Orthopedic treatment using bone-anchored maxillary protraction (BAMP)” 3. Noriko Niizato Department of Pediatric Dentistry, Hiroshima University Graduate School of Biomedical and Health Sciences The dental caries condition of abused children in temporary shelters in Hiroshima” 4. Satoko Oikawa Division of Pediatric Dentistry, Bioactive Compound Library Department of Oral Health and Development Sciences, Tohoku University Graduate School of Dentistry Regulation of dental epithelial cell proliferation and differentiation by laminin” 5. Yuko Nakamura

Department of Pediatric Dentistry, School of Life Dentistry, Nippon Dental University Three-dimensional reconstruction of root malformation in mice by cyclophosphamide 1. Noriko Niizato Department of Pediatric Dentistry, Hiroshima University Graduate School of Biomedical and Health Sciences The Oral Health Condition of Abused see more Children in Temporary Shelters in Japan The Japanese Journal of Pediatric Dentistry; 50 (3) 237–242, 2012 2. Masamichi Ide Department of Pediatric Dentistry,Tsurumi University School of Dental Medicine “Longitudinal Dental Management of Hypophosophatemic Ricketes: Case Report The Japanese Journal of Pediatric Dentistry; 50 (4) 313–319, 2012 3. Aya Yamada Division of Pediatric Dentistry, Department of Oral Health and Development Sciences, Tohoku University Graduate School of Dentistry Epithelial-mesenchymal interaction reduces inhibitory effects of fluoride on proliferation

and enamel matrix expression in dental epithelial cells Pediatric Dental Journal; 22 (1) 55–63, 2012 4. Maiko Bori Department of Pediatric Dentistry, Kyushu Dental University Influence of childhood type II diabetes on bone formation in the growth period Pediatric Dental Journal; 22 (2) 125–139, 2012 5. Hiroshi Sekiguchi Department of Pediatric Dentistry, Tokyo Dental College Missense mutation of EDA1 gene in Japanese family with X-linked anhidrotic ectodermal dysplasia Pediatric Dental Journal; 22 (2) 188–192, 2012 1. Chiaki Yamada-Ito Department of Pediatric Dentistry, Field of Developmental Medicine Course for Health Science, Kagoshima University Graduate School of Medical and Dental Sciences Smoothness of molar movement during gum chewing in children with primary dentition” 2.

8 WE is potentially lethal in a short time if not promptly recognized and treated. In severely malnourished patients, standard doses of thiamine in multivitamin infusion may not be sufficient. This diagnosis should be suspected if neurologic symptoms develop in this context. We believe

that prophylactic additional RO4929097 solubility dmso supplementation of thiamine is reasonable in malnourished patients receiving enteral or parenteral nutrition in order to avoid thiamine deficiency complications. The authors declare that no experiments were performed on humans or animals for this investigation. The authors declare that they have followed the protocols of their work center on the publication of patient data and that all the patients included in the study received sufficient information and gave their written informed consent to participate in the study. The authors have obtained the written informed consent of the patients or subjects mentioned in the article. The corresponding author is in possession of this document. The authors have not received any funding regarding this study. Dr. Paula Ministro has participated in advisory boards of MSD and AbbVie. All the oher authors declare

no conflicts AZD5363 supplier of interest. “
“Foreign body ingestion and food bolus impaction occur commonly,1 and 2 however, most ingested foreign bodies that reach the stomach pass safely through the intestinal tract. Foreign body-induced esophageal perforation is responsible for 16.7% of esophageal perforations and it has been regarded as the most serious injury of the digestive tract,3 particularly if not diagnosed and treated

promptly, being associated with respiratory failure, sepsis or hemorrhage.4 The mortality rate of esophageal perforations hovers close to 20%, especially in cases in which Cell press treatment is delayed for more than 24 h.5 Esophageal perforation management remains controversial and treatment decisions should be individualized depending on the duration of impaction, type of foreign body, size and perforation.6 Surgical primary repair is often the preferred approach, however, there may be a role for interventional endoscopy including the use of stents.7 and 8 Treatments performed before the development of mediastinitis are lifesaving in esophageal perforation patients.9 We report a case of successful endoscopic management in a delayed diagnosis of an esophageal perforation presenting with an associated peri-esophageal abscess. A 57 year-old man was referred to the emergency room due to suspicion of a foreign body impaction. The patient complaints were substernal chest pain, with solid food dyspaghia, fever, progressive prostration and pointed out that he had eaten chicken 5 days before. Blood chemistry revealed leukocytosis and increased C-reactive protein (147 mg/L) and there were no reported abnormalities at the chest X-ray.

The selection was made taking into account the relevance of the target organ and the nature of the test article. The human lung-derived BEAS-2B cell line was first described in 1988, when normal bronchial epithelial cells obtained from autopsy of non-cancerous individuals were isolated, then infected with a replication-defective 12-SV40/adenovirus hybrid and cloned to create an immortalized phenotype (Reddel et al., 1988). The non-cancerous phenotype of BEAS-2B

cells is an advantage this website in the investigation of carcinogenic processes such as DNA damage and cell transformation (Sun et al., 2011). Therefore, BEAS-2B cells have been considered as a relevant cell line for in vitro toxicology testing in the field of pollutants, tobacco products and nanomaterials ( Persoz et al., 2012, Veljkovic et al., 2011 and Haniu et al., 2011). Although, several studies have employed BEAS-2B cells to evaluate the effect of some pro-toxicants such as B[a]P and 4-(N-methylnitrosamino)-1-(3-pyridinyl)-1-butanone (NNK), the metabolic capacity of this particular cell line has not been fully

characterized ( Ovrevik et al., 2010 and Proulx BIBF 1120 supplier et al., 2005). Here, BEAS-2B cells were tested and compared to the lung-derived A549 cells broadly used as pulmonary in vitro system but with no cytochrome P450 expression reported for CYP1A2 and the CYP2A family, and inducibility documented for CYP1A1/1B1 genes ( Hukkanen et al., 2002 and Castell et al., 2005). ADP ribosylation factor Cells derived from hepatocarcinomas considered

to have a more extensive cytochrome P450 enzyme activity (HepG2 and HepaRG) were used as a more comprehensive control for our CYP assays ( Jennen et al., 2010). Moreover, the results were contrasted to those reported in primary human bronchial epithelium culture ( Newland et al., 2011 and Courcot et al., 2012). The results of this study are considered to be useful for in vitro toxicological testing using the cell line BEAS-2B as cell system. Furthermore, we propose that the outlined strategy can be incorporated in the characterization of cell systems used in in vitro testing. The human bronchial epithelial cell line (BEAS-2B), purchased from ATCC (United States), was seeded into culture vessels that had been pre-coated with 0.03 mg/mL PureCol® bovine collagen solution (Nutacon, The Netherlands). Cells were maintained in Bronchial Epithelial Growth Medium (BEGM®) at 37 °C and 5% CO2 in a humidified incubator. BEGM® was prepared by supplementing Bronchial Epithelial Basal Medium with growth supplements provided in the manufacturer’s BEGM® SingleQuot® kit (Lonza Group Ltd., Belgium) containing: bovine pituitary extract, hydrocortisone, human epidermal growth factor, epinephrine, insulin, triiodothyronine, transferrin, gentamicin/amphotericin-B and retinoic acid.