risk factors of food chain 2017

RISK FACTORS OF FOOD CHAIN 2017 PROCEEDIINGS OF XVIII INTERNATIONAL SCIENTIFIC CONFERENCE

September 20 - 22, 2017 Żmiąca, Poland

Risk Factors of Food Chain 2017

20-22 September, Żmiąca, Poland

PROCEEDINGS OF XVIII INTERNATIONAL SCIENTIFIC CONFERENCE

RISK FACTORS OF FOOD CHAIN 2017

September 20 - 22, 2017 Żmiąca, Poland


Reviewers Monika Martiniakova, Professor, Ph.D. Norbert Lukáč, Professor, MSc., Ph.D. Peter Massányi, Professor MVDr. DrSC. Robert Stawarz, Professor, Ph.D.

Editors Renata Muchacka, Ph.D. Eng. Zofia Goc, Ph.D. Agnieszka Greń, Professor, Ph.D. Grzegorz Formicki, Professor, Ph.D.

© Copyright by Pedagogical University of Cracow, Kraków 2017

ISBN 978-83-8084-097-3 DOI 10.24917/9788380840973




Institute of Biology, Faculty of Geography and Biology, Pedagogical University of Cracow, Poland Department of Animal Physiology, Faculty of Biotechnology and Food Sciences, Slovak University of Agriculture in Nitra, Slovak Republic Faculty of Biology and Agriculture, University of Rzeszów, Poland Department of Animal Physiology and Health, Faculty of Agricultural and Environmental Sciences, Szent István University, Gödöllő, Hungary



Scientific Committee of the Conference Agnieszka GREŃ, Professor, Ph.D. Pedagogical University of Cracow, POLAND Peter MASSÁNYI, Professor MVDr. DrSC. Slovak University of Agriculture, Nitra, SLOVAK REPUBLIC Norbert LUKÁČ, Professor, MSc., Ph.D. Slovak University of Agriculture, Nitra, SLOVAK REPUBLIC Robert STAWARZ, Professor, Ph.D. Pedagogical University of Cracow, POLAND Grzegorz FORMICKI, Professor, Ph.D. Pedagogical University of Cracow, POLAND Waldemar SZAROMA, Professor, Ph.D. Pedagogical University of Cracow, POLAND Ján TOMÁŠ, Professor, Ing. C.Sc. Slovak University of Agriculture, Nitra, SLOVAK REPUBLIC Miroslava KAČÁNIOVÁ, Professor, Ph.D. Ing. Slovak University of Agriculture, Nitra, SLOVAK REPUBLIC Monika MARTINIAKOVA, Professor, Ph.D. Constantine the Philosopher University, Nitra, SLOVAK REPUBLIC Radoslav OMELKA, Professor, Ph.D. Constantine the Philosopher University, Nitra, SLOVAK REPUBLIC Eric Rendón SCHNEIR, Professor, Ph.D. The National University Agraria La Molina, Lima, PERU Małgorzata DŻUGAN, Professor, Ph.D. University of Rzeszów, Rzeszów, POLAND Łukasz J. BINKOWSKI, Ph.D. Pedagogical University of Cracow, POLAND Gábor GÉCZI, Ph.D. Szent István University, Gödöllő, HUNGARY Ales PAVLIK, Ph.D. Mendle University, Brno, CZECH REPUBLIC Barbara PINTO, Ph.D. University of Pisa, ITALY Francesco VIZZARRI, Ph.D. University of Molise, Campobasso, ITALY



Organizing Committee Agnieszka GREŃ, Associate professor, Ph.D. (UP Cracow, Poland) Łukasz J. BINKOWSKI, Assistant professor, Ph.D. (UP Cracow, Poland) Zofia GOC, Assistant professor, Ph.D. (UP Cracow, Poland) Renata MUCHACKA, Assistant professor, Ph.D. Eng. (UP Cracow, Poland) Bartłomiej ZYŚK, Assistant professor, Ph.D. (UP Cracow, Poland) Marek GUZIK, Assistant professor, Ph.D. (UP Cracow, Poland) Włodzimierz WOJTAŚ, Ph.D. (UP Cracow, Poland) Tomasz ŁACIAK, Ph.D. (UP Cracow, Poland) Edyta KAPUSTA, MSc. (UP Cracow, Poland) Martyna BŁASZCZYK, MSc. (UP Cracow, Poland) Katarzyna KUCHARSKA, MSc. (UP Cracow, Poland) Thiep VO VAN, MSc. (UP Cracow, Poland) Magdalena SEMLA, MSc. (UP Cracow, Poland) Marta BATORYNA, MSc. (UP Cracow, Poland) Łukasz KOŁODZIEJCZYK, MSc. (UP Cracow, Poland) Eva TVRDÁ, Ph.D. (SUA Nitra, Slovakia) Tomáš SLANINA, Ph.D. (SUA Nitra, Slovakia) Anna PASTERNAKIEWICZ, Ph.D. (UR Rzeszów, Poland) Péter KORZENSZKY, Dr., Ph.D. (SzIU Gödöllő, Hungary)

Secretary Łukasz J. BINKOWSKI, Ph.D. (UP Cracow, Poland) Martyna BŁASZCZYK, MSc. (UP Cracow, Poland) Zofia GOC, Ph.D. (UP Cracow, Poland) Renata MUCHACKA, Ph.D. Eng. (UP Cracow, Poland)



CONTENT EFFECTS OF T-2 TOXIN ON HUMAN OSTEOBLAST VIABILITY AND MORPHOLOGY Adamkovicova M., Mondockova V., Lukacova M., Babosova R., Sranko P., Omelka R., Martiniakova M., Bauerova M. ........................................................................................... 8 QUERCETIN HAS INSIGNIFICANT EFFECT ON QUANTITATIVE 3D CHARACTERISTICS OF FEMORAL BONE IN ADULT RABBITS Babosova R., Mondockova V., Adamkovicova M., Omelka R., Goc Z., Kalafova A., Capcarova M., Martiniakova M. ............................................................................................................. 13 DIFFERENCES BETWEEN CAFFEINE CONTENTS IN VARIOUS COMMERCIAL COFFEES ACCORDING TO GEOGRAPHICAL ORIGIN Bobkova A., Arvay J., Snirc M., Belej L., Bobko M. ............................................................. 19 METAL CONTENT AND ANTIOXIDANTS IN SEMINAL PLASMA OF SMOKING MEN Formicki G., Greń A., Muchacka R., Goc Z., Kapusta E., Massanyi P., Stawarz R. ................... 25 THE EFFECT OF ANTIOXIDANTS ON SERUM GLUCOSE LEVEL AND LIVER ENZYMES ACTIVITY IN MICE WITH INDUCED OBESITY Goc Z., Kapusta E., Formicki G., Greń A., Muchacka R., Massányi P., Lukáč N., Slanina T., Jambor T. ..................................................................................................................... 37 BIOCHEMICAL MARKERS OF INFLAMMATION IN DAIRY CATTLE Greifová H., Tvrdá E., Červeňanská P., Tušimová E., Kováčik A., Zbyňovská K., Lukáč N. ....... 44 INFLUENCE OF COENZYME Q10 AND QUERCETIN IN ANTIOXIDANT ENZYME ACTIVITY IN MOUSE BRAIN INFLAMMATION Kapusta E., Goc Z., Muchacka R., Greń A., Šarocká A., Babosová R., Martiniaková M., Omelka R., Formicki G., Szaroma W. ........................................................................................... 50 POTENTIALLY TOXIC ALGAL BLOOMS IN THE BUFFER ZONE OF THE OJCOWSKI NATIONAL PARK, SOUTHERN POLAND Kołodziejczyk Ł. M. ........................................................................................................ 61 CHANGES IN FEMORAL BONE STRUCTURE OF MALE RABBITS AFTER APRICOT SEEDS ADMINISTRATION Kováčová V., Ďúranová H., Šarocká A., Omelka R., Halenár M., Kolesárová A., Martiniaková M. ................................................................................................................................... 66 DOES THE TYPE OF HEN HOUSING AFFECT CALCIUM AND LEAD CONCENTRATIONS IN EGG SHELLS? Kucharska K., Gęca A., Binkowski Ł. J., Błaszczyk M., Stawarz R. ........................................ 73 RISK FACTORS OF FOOD CHAIN – THE CONFERENCE HISTORY Massányi P., Binkowski Ł., Formicki G., Greń A., Stawarz R., Golian J., Lukáč N., Toman R..... 79 REPRODUCTIVE TOXICITY OF MERCURY IN VIVO AND IN VITRO Massányi P., Formicki G., Lukáč N., Slivková J., Kováčik J., Toman R., Binkowski Ł., Greń A., Stawarz R. .................................................................................................................... 84

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EFFECT OF ADDING COMMON NETTLE EXTRACTS TO DRINKING WATER ON THE LIPID PEROXIDATION, ANTIOXIDANT ENZYMES ACTIVITY AND GLUTATHIONE LEVEL IN EGGS OF LAYING HENS EXPOSED TO ELEVATED AIR TEMPERATURE Muchacka R., Sosnówka-Czajka E., Skomorucha I., Kapusta E., Greń A., Goc Z. .................... 90 INFLUENCE OF SMOKING ON THE EXPRESSION OF p53 AND Ki-67 PROTEINS IN PATIENTS WITH NON-SMALL CELL LUNG CANCER Rogoziński P., Binkowski Ł., Semla M., Kucharzewski M., Wandzel P., Bruliński K. ................. 96 THE IMPACT OF ETHANOL ON FEMORAL BONE STRUCTURE IN MICE Sarocka A., Kovacova V., Omelka R., Kapusta E., Goc Z., Formicki G., Martiniakova M. ........ 109 HOW ENVIRONMENTAL CONTAMINANTS AFFECT OVARIAN FUNCTIONS. PROTECTIVE EFFECT OF TWO DIFFRENT EXTENDERS ON THE SPERM MOTILITY OF ENDANGERED ORAVKA CHICKEN Svoradová A., Kuželová L., Baláži A., Vašíček J., Tomková M., Chrenek P. .......................... 115 LEAD AND CADMIUM CONCENTRATIONS IN SHEEP (OVIS ARIES) BLOOD SAMPLES FROM POLLUTED AREA AND THEIR ECOTOXICOLOGICAL INTERACTIONS WITH HEPATIC PROFILE Tirpak F., Kovacik A., Arvay J., Tusimova E., Zbynovska K., Halo M. Jr., Massanyi P. ........... 119 ANATOMY OF DIGESTIVE SYSTEM CHINCHILLA (CHINCHILLA LANIGERA) Urynowicz W. T., Batoryna M. ....................................................................................... 125 INDEX ........................................................................................................................... 131

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DOI 10.24917/9788380840973.1

EFFECTS OF T-2 TOXIN ON HUMAN OSTEOBLAST VIABILITY AND MORPHOLOGY

Adamkovicova M.1, Mondockova V.1, Lukacova M.1, Babosova R. 2, Sranko P.1, Omelka R.1, Martiniakova M.2, Bauerova M.1 1

Department of Botany and Genetics, Constantine the Philosopher University in Nitra, Slovak Republic

2

Department of Zoology and Anthropology, Constantine the Philosopher University in Nitra, Slovak Republic

[email protected]

Abstract

Fusarium mycotoxins have various acute and chronic effects on humans and animals. However, there are limited data on their cytotoxic potential on bone cells. Thus, the effects of T-2 toxin on osteoblasts growth were investigated in vitro. Primary human osteoblasts were exposed to T-2 toxin (10 and 100 ng/mL) for 48 hr. Osteoblast viability was determined by the colorimetric MTT assay and measured spectrophotometrically. Cell morphology was described both qualitatively and quantitatively. Osteoblasts viability significantly decreased with the increase of T-2 toxin concentration. Moreover, T-2 toxin had a profound effect on osteoblast morphology. Exposure to 100 ng/mL T-2 toxin resulted in significantly decreased osteoblast surface area. Strong increase of irregularly shaped and dead floating cells was found. The results suggest that T-2 toxin may affect bone formation differently depending on exposure conditions.

Keywords: T-2 toxin, osteoblast, morphology, viability Introduction T-2 toxins are mycotoxins of the group trichothecenes type A produced by fungi of the Fusarium genus (Wu et al., 2010). Trichothecenes are agriculturally among the most important mycotoxins that are present in cereal commodities such as wheat, barley and maize (Edlayne et al., 2009). Common manifestations of trichothecene toxicity are growth retardation, reproductive disorders, immuno-compromization, feed refusal and vomiting (Peraica et al., 1999; Hussein, Brasel, 2001; Kimura et al., 2007). There are limited data on the potential toxic effects of trichothecenes on bone cells. Therefore, the aim of the present study was to evaluate the effects of T-2 toxin on osteoblasts viability and morphology in vitro.

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Materials and Methods Primary human osteoblasts and media were purchased from PromoCell GmbH (Heidelberg, Germany). Cells were cultured according to protocol at 37°C in air containing 5% CO2. Cell viability was investigated in a 96-well plate format using the colorimetric MTT reduction assay. Cells were plated at a density of 5x103 cells per well. Culture medium was changed and treatment was initiated with T-2-toxin (ROMER LABS Division Holding GmbH, Tulln) 24 hr after plating. Cells were exposed to T-2 toxin at concentration10 ng/ml and 100 ng/ml for 48 hr. Medium containing 10% MTT (3-(4, 5-dimethylthiazol-2-yl)- 2, 5-diphenyltetrazolium bromide) solution (PromoKine, Heidelberg, Germany) was added to each well, and the tissue culture plates were incubated at 37°C for 4 hr. Afterwards, the MTT containing medium was entirely removed. After adding DMSO (Sigma, USA), the optical density was measured by a microplate reader (Thermo Scientific Varioskan) at 570 nm with a reference filter at 630 nm. Untreated control was regarded as 100% cell viability. The effects of different T-2 toxin concentrations on cell morphology were observed under a phase contrast microscope. Photographs were taken at representative areas of culture plates. The surface area of individual cells was measured using ImageJ 1.46r software (National Institute of Mental Health, Maryland, USA). At least ten representative cells were measured from each independent experiment. Data are presented as the mean ± standard deviation of three independent experiments. Statistical analysis was performed by analysis of variance (ANOVA) followed by the Tukey-test using SPSS Statistics 17.0 (Chicago, IL, USA). Results and Discussion T-2 toxin is the most potent mycotoxin of the trichothecenes group that pose risk to human health (Peraica et al., 1999). In the current study, the cytotoxicity of T-2 toxin on human osteoblast was evaluated. Several studies suggested the significant concentration-dependent reduction of viability cells with T-2 toxin (Königs et al., 2009; Yang et al., 2017). No significant cytoxicity was observed in osteoblasts exposed to 10 ng/mL T-2 toxin. Cell viability assay showed that cells maintained their growth rate in comparison with control group. However, osteoblasts viability significantly decreased with the increase of T-2 concentration after 48 hr exposure (Fig. 1). Simultaneously with the decrease of cell viability, the prominent cell alterations observed were obtained at a higher concentration of the T-2 toxin. There was a significant increase of irregularly shaped and dead floating cells in group exposed to 100 ng/mL of T-2 toxin (Fig. 2). Cell shape changed from spread, spindle and stellate to a contracted, circular morphology. On the other hand, the shape of the osteoblast exposed to 10 ng/mL T-2 toxin did not changed compared to the osteoblast from control 9



group (Fig. 3). The results concerning effects of T-2 toxin on cell morphology agree with several literature data which showed that acute exposure to trichothecene mycotoxins caused ultrastructural alterations and growth inhibition (Trusal, 1985; Reubel et al., 1987).

Figure 1. Concentration-dependent effects of T-2 toxin treatment on cell viability. Cell viability was determined by MTT assay. The percentage of viability was calculated as the following formula: (viable cells) % = (OD of drug-treated sample/OD of untreated sample)×100. C – control group, E1 – group exposed to 10 ng/mL T-2 toxin, E2 - 100ng/mL T-2 toxin. Values were expressed as the mean ± SD. *p 5 μm/s), percentage of progressive motile spermatozoa (motility > 20 μm/s), DCL (distance curved line; μm), DAP (distance average path, μm), DSL (distance straight line, μm), VCL (velocity curved line, μm/s), VAP (velocity average path, μm/s), VSL (velocity straight line, μm/s), LIN (linearity – VSL:VCL), STR (straightness – VSL:VAP), WOB (wobble – VAP:VCL), ALH (amplitude of lateral head displacement, μm) and BCF (beat cross frequency, Hz). Obtained data were statistically analyzed by a spreadsheet (Microsoft Office Excel) and SAS package using t-test and Scheffe's test. For the Annexin V analysis semen samples were centrifuged at 5000 rpm for 6 min and resuspended in equal volume of HEPES–buffered saline. Semen suspension (5 μL) was mixed with 100 μL working solution of Annexin V-Fluos (Annexin-V-Fluos staining kit, Roche Diagnostics, Germany) and incubated for 15 min at 37°C. Afterwards aliquot of the semen suspension was placed between microslide and coverslip into 5 μL of the anti-fade medium Vectashield containing DAPI fluorescent dye. Staining with Annexin V was checked under the Leica fluorescent microscope (Leica Microsystem, Germany) using 488 nm wavelength filter. Spermatozoa with disordered membrane exhibited green fluorescence, while live spermatozoa were unstained.

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Results and Discussion

In vivo experiment Qualitative examination showed undulation of basal membrane, dilatation of blood vessels in interstitium and occurrence of empty spaces in germinal epithelium in testes. Quantitative microscopic analysis found decreased relative volume of germinal epithelium, increased relative volume of interstitium and increased apoptosis occurrence suggesting damaged interstitium and revealed occurrence of edemas. The relative volume of seminiferous tubules showed higher luminization. The number of nuclei was decreased in all experimental groups what is in positive relation with occurrence of empty spaces. Also other evaluated criteria demonstrated significant differences between control group and experimental groups. In vivo part of study reports a negative effect of mercury on the structure and function of testes.

In vitro experiment In in vitro study the effects of mercury (HgCl2) on the motility and structural integrity of rabbit spermatozoa were investigated. At Time 0 the highest motility was detected in the control group (67.09±8.72%). Motility in groups with mercury administration was lower in comparison with control. Significant differences were detected in groups with 50.0–83.3 µg HgCl2/mL (p