Libor Vojtek

Biological activities of selected polyphenol-rich fruits related to immunity and gastrointestinal health

Small fruits are a rich source of bioactive substances, including polyphenols, and are therefore suitable raw materials for the production of functional foods. In the current work, we studied the antioxidative properties of six fruits: rosehip, chokeberry, hawthorn, blackcurrant, blueberry and rowanberry via different methods (ORAC, TRAP, HORAC and inhibition of lipid peroxidation). Their effect on the production of reactive oxygen species (ROS) by phagocytes, antimicrobial properties against 11 human pathogens, and mitogenic effect on hamster spleen lymphocytes were also tested. Rosehip extract showed the highest antioxidant activity via ORAC, TRAP and HORAC assays, whereas blueberry extract was the most potent inhibitor of lipid peroxidation. All extracts inhibited ROS production of opsonized zymosan-activated phagocytes, indicating that extracts interfere with the signaling cascade of phagocyte activation upstream to the protein kinase C activation. Chokeberry, blackcurrant and rowanberry extracts revealed strong antimicrobial properties against a broad spectrum of microorganisms and also had the highest mitogenic activity.

Escherichia coli K-12 (pEGFPluxABCDEamp): a tool for analysis of bacterial killing by antibacterial agents and human complement activities on a real-time basis

Photorhabdus luminescens luxCDABE genes were integrated into E. coli K-12 using a high copy number plasmid containing modified luxABCDE genes under the control of the powerful Lac promoter. This strain emitted 10 times higher bioluminescence (BL) than P. luminescens. BL production under different growth conditions was studied. In both bacterial strains, the increase in BL signal correlated with the increase in optical density (OD) in a rich growth medium. However, at the logarithmic growth phase, the BL signal was roughly constant. By contrast, in minimal growth media, there was no substantial growth and the BL/cell was approximately five times higher than in the rich medium. The dynamic measurement range of BL was 10(2) -10(7) colony-forming units (CFU) in E. coli and 10(3) -10(7)  CFU in P. luminescens. Because the decrease in the BL signal correlated with the decrease in CFU and OD, i.e. the number of bacterial cells killed, it proved to be very suitable for assessing the antibacterial effects of different antimicrobial agents. Unlike with plate counting, the kinetics of killing can be monitored on a real-time basis using BL measurements. Complement activities in different samples can be estimated using only one serum dilution. The transformed E. coli strain appeared to be superior to P. luminescens in these applications because E. coli was complement sensitive, the detection limit of E. coli was one order lower and the BL-producing system of P. luminescens appeared to be quite unstable.

Rank-based biomarker index to assess cadmium ecotoxicity on the earthworm Eisenia andrei.

A proper soil risk assessment needs to estimate the processes that affect the fate and the behaviour of a contaminant, which are influenced by soil biotic and abiotic components. For this reason, the measurement of biomarkers in soil bioindicator organisms, such as earthworms, has recently received increasing attention. In this study, the earthworm Eisenia andrei was used to assess the pollutant-induced stress syndrome after exposure to sublethal concentrations of Cd (10 or 100 μg g(-1)) in OECD soil, after 14 d of exposure. Cadmium bioaccumulation and potential biomarkers such as catalase (CAT), hydrogen peroxide (H2O2), glutathione-S-transferase (GST), malondialdehyde (MDA), phenoloxidase (PO), metallothioneins (MTs) and genotoxic damage were determined. Results suggested that the exposure to 10 and 100 μg g(-1) Cd significantly increased Cd bioaccumulation, MTs and MDA; 100 μg g(-1) Cd contamination evidenced significantly higher values of H2O2 content and PO activity; CAT activity was inhibited at the higher concentration while GST and Comet assay did not show any significant differences from the control. Rank-based biomarker index showed that both different contaminated soils had an effect on the earthworms and allowed to validate the ecotoxicological relevance of this battery of biomarkers for a promising integrated multi-marker approach in soil monitoring and assessment.

A novel in situ silver/hyaluronan bio-nanocomposite fabrics for wound and chronic ulcer dressing: In vitro and in vivo evaluations

In-situ formed hyaluronan/silver (HA/Ag) nanoparticles (NPs) were used to prepare composite fibers/fabrics for the first time. Different concentrations of silver nitrate (1, 2mg/100ml) were added at ambient temperature to sodium hyaluronate solution (40mg/ml), then the pH was increased to 8 by adding sodium hydroxide. The in-situ formed HA/Ag-NPs were used to prepare fibers/nonwoven fabrics by wet-dry-spinning technique (WDST). UV/vis spectroscopy, SEM, TEM, DLS, XPS, XRD and TGA were employed to characterize the structure and composition of the nanocomposite, surface morphology of fiber/fabrics, particle size of Ag-NPs, chemical interactions of Ag0 and HA functional groups, crystallinity and thermal stability of the wound dressing, respectively. The resultant HA/Ag-NPs1 and HA/Ag-NPs2 composite showed uniformly dispersed throughout HA fiber/fabrics (SEM), an excellent distribution of Ag-NPs with 25±2, nm size (TEM, DLS) and acceptable mechanical properties. The XRD analysis showed that the in-situ preparation of Ag-NPs increased the crystallinity of the resultant fabrics as well as the thermal stability. The antibacterial performance of medical HA/Ag-NPs fabrics was evaluated against gram negative bacteria E. coli K12, exhibiting significant bactericidal activity. The fibers did not show any cytotoxicity against human keratinocyte cell line (HaCaT). In-vivo animal tests indicated that the prepared wound dressing has strong healing efficacy (non-diabetics/diabetics rat model) compared to the plain HA fabrics and greatly accelerated the healing process. Based on our results, the new HA/Ag-NPs-2mg nonwoven wound dressing fabrics can be used in treating wounds and chronic ulcers as well as cell carrier in different biological research and tissue engineering.

Novel electrospun gelatin/oxycellulose nanofibers as a suitable platform for lung disease modeling

Novel hydrolytically stable gelatin nanofibers modified with sodium or calcium salt of oxycellulose were prepared by electrospinning method. The unique inhibitory effect of these nanofibers against Escherichia coli bacteria was examined by luminometric method. Biocompatibility of these gelatin/oxycellulose nanofibers with eukaryotic cells was tested using human lung adenocarcinoma cell line NCI-H441. Cells firmly adhered to nanofiber surface, as determined by scanning electron microscopy, and no signs of cell dying were detected by fluorescent live/dead assay. We propose that the newly developed gelatin/oxycellulose nanofibers could be used as promising scaffold for lung disease modeling and anti-cancer drug testing.

Consequent effects of the great cormorant (Phalacrocorax carbo sinensis) predation on parasite infection and body condition of common carp (Cyprinus carpio)

Lesions ranging from surface wounds to deep tissue wounds caused by cormorant predation were observed on several species of the farmed fish in Pohořelice, Czech Republic. Two-year-old stocked common carp Cyprinus carpio harvested in late March were examined for ectoparasites and endoparasites, injuries extent, and lysozyme concentration in skin mucus. Additionally, three body condition indices were measured. Endoparasite infection occurred only scarcely. Wounded fish were more susceptible to the ectoparasites Gyrodactylus spp. and Dactylogyrus spp. (Monogenea), and Ichthyophthirius multifiliis (Ciliophora). The intensity of infection of other ectoparasites Eudiplozoon nipponicum (Monogenea), Argulus spp. (Branchiura) and trichodinids (Ciliophora) did not significantly differ between wounded and control groups of fish. Lysozyme concentration in fish mucus was significantly higher in wounded fish and was positively associated with both the extent of damaged epithelium and Gyrodactylus spp. abundance. There were no differences in Fulton’s condition factor and lipid content in muscle and liver tissues between wounded and non-wounded fish. Higher values of spleen-somatic index in wounded fish corresponded to increased intensity of parasite infection, most likely reflecting changes in immune system of infected fish. Although our results did not show any significant effect of cormorant attacks on fish condition, the wounded fish had significantly higher parasite numbers which could impact the growth or survival of the fish throughout the production season.

Associations of urban environmental pollution with health-related physiological traits in a free-living bird species

Urban environmental pollution results in contamination of the tissues of synanthropic organisms by toxic trace elements with potential impacts on human health. Passerine birds may serve as convenient indicators of such contamination. In this study we investigated the effect of blood and plumage contamination with heavy metals (lead Pb, cadmium Cd, copper Cu, chromium Cr) and arsenic metalloid (As) on condition, health and ornamental colour in free-living great tit (Parus major) males from 13 cities across the Czech Republic (EU), mist netted during the early breading season (April-May). Our results showed a significant association of heavy metal tissue contamination with immune function, namely leukocyte composition in the avian blood circulation. High heavy metal contamination in bird feathers was linked to a high heterophil/lymphocyte (H/L) ratio, indicating long-term stress in individuals inhabiting heavily polluted environments. In contrast, males with higher concentrations of heavy metals in blood had a lower H/L ratio, assumingly due to the direct toxicity of heavy metals in certain cell types. This is also supported by traits indicative of anaemia-like haemolytic conditions (decreased absolute erythrocyte count) and increased haematopoiesis (a tendency for increased frequencies of immature erythrocytes). We did not find any association of heavy metal contamination with the bacteriolytic activity of plasma complement, feather growth or ornamentation (black breast stripe area and yellow colouration). There was no significant relationship between heavy metal contamination in blood or feathers and PM10 pollution at the study sites. Our correlational study is the first to show on a large geographic scale that despite strict European air pollution regulations and regular monitoring that have allowed general improvements in atmospheric contamination, non-degradable heavy metals persistently contaminate animal blood and feathers in anthropogenic environments at levels that may have subclinical yet physiological effects with varied influence on health.

Effect of T-2 toxin-contaminated diet on common carp (Cyprinus carpio L.).

ABSTRACT The T‐2 toxin, a fungal metabolite produced by Fusarium molds, occurs in a range of agriculture products. Reduced availability of fish meal has led to increasing use of cereals as a source of protein in commercial aquaculture feeds, which has increased the potential for mycotoxin contamination. The purpose of this study was to investigate toxicity of T‐2 toxin intake in common carp (Cyprinus carpio L.) using haematological, biochemical and immunological parameters and oxidative stress indices. In a four‐week feeding trial, fish were fed a commercial diet with 5.3 mg/kg T‐2 toxin added. Ingestion of contaminated diet did not lead to mortality of fish, probably due to lower feed intake. On the other hand, it significantly affected haematological variables such as haematocrit, haemoglobin, red blood cell counts leading to anemia and white blood cell counts leading to leukopenia due to lymphopenia. Plasma glucose concentration and alanine amino transferase activity showed a significant increase while triglycerides concentration decreased. Activity of ceruloplasmin was significantly decreased in plasma. Further, liver glutathione S‐transferase activity was significantly increased and catalase activity decreased, in parallel with a significant increase in caudal kidney catalase activity and a decrease in glutathione peroxidase activity. Finally, lipid peroxidation (detected as malondialdehyde) was significantly increased in the liver and caudal kidney. Changes in non‐specific immune response and cytokine levels in head kidney indicated immune system sensitivity to T‐2 toxin. Overall, the results demonstrate that this feed‐borne mycotoxin is able to induce anaemia and oxidative stress and cause changes in the immune response of common carp. HIGHLIGHTSThe aim of the study was to investigate toxicity of T‐2 toxin intake in common carp.Feeding trial led to changes in haematological and biochemical parameter.Lipid peroxidation was significantly increased in the liver and caudal kidney.Exposition to T2 led to changes in non‐specific immune response.Exposition to T2 led to changes in mRNA expression for selected cytokines.

Plasma-chemical modifications of cellulose for biomedical applications

Abstract A 6-carboxycellulose (in medicine known as “oxidized cellulose” or “oxycellulose”) is one of the cellulose derivatives popular in the field of surgery. Health products based on oxidized cellulose are great local hemostatics with unique bactericidal and fully bioabsorbable effects. Traditional process of native cellulose oxidation is described as a complex radical reaction in strong acidic liquid medium doped by toxic nitrous radicals (NO*). Our plasma-chemical reaction demonstrates a new synthesis method of oxidized cellulose with unique bactericidal effect. This plasma-chemical treatment is based on atmospheric plasma discharge in liquid medium leading to the oxidation of polysaccharide molecules resulting in oxycellulose. Final oxycellulose properties were evaluated by infrared spectroscopy and carboxyl content determination. The biological impact showed a strong germicidal effect. Graphical Abstract

Effect of the mycotoxin deoxynivalenol on the immune responses of rainbow trout (Oncorhynchus mykiss).

515 Veterinarni Medicina, 60 , 2015 (9): 515–521 Original Paper doi: 10.17221/8443-VETMED Effect of the mycotoxin deoxynivalenol on the immune responses of rainbow trout ( Oncorhynchus mykiss ) I. Matejova 1,2 , M. Vicenova 2 , L. Vojtek 3 , H. Kudlackova 2 , K. Nedbalcova 2 , M. Faldyna 2 , E. Sisperova 1 , H. Modra 1 , Z. Svobodova 1 1 University of Veterinary and Pharmaceutical Sciences, Brno, Czech Republic 2 Veterinary Research Institute, Brno, Czech Republic 3 Masaryk University, Brno, Czech Republic ABSTRACT : The trichothecene mycotoxin deoxynivalenol (DON) is commonly found as a natural contaminant in cereals such as wheat, barley, and corn, and exhibits various toxicological effects when present in animal feeds. The effects of DON at a nominal 2 mg/kg feed on immune responses of rainbow trout were investigated, includ - ing relative gene expression of important cytokines (TNF-alfa, IL-8, IL-1beta, IL-10), lysozyme concentration in skin mucus, and level of antigen-specific IgM in blood plasma after vaccination with the commercial vaccine AquaVac ERM containing Yersinia ruckeri type 1 (Hagerman strain). Twenty one-year-old rainbow trout Oncorhynchus mykiss were randomly divided into two groups. The control received a commercial feed with a naturally occurring low level of DON (225 ug /kg feed), while an experimental group was fed the same formulation with DON added to 1964 ug /kg feed. The trial continued for 23 days. Consumption of feed with added DON showed a significant effect on the immune system, as indicated by a higher level of pro-inflammatory cytokine TNF-alfa ( P < 0.05) and of IL-8 (non-significant) in head kidney. Expression of the pro-inflammatory gene IL-1beta and the expression of a gene encoding anti-inflammatory cytokine (IL-10) were not influenced by DON treatment. Effects on the concentration of skin mucus lysozyme and specific IgM antibody levels were not observed during this experiment. These results suggest that prolonged ingestion of low doses of DON may influence the immune responses of rainbow trout.