Robust expansion of human hepatocytes in Fah-/-/Rag2-/-/Il2rg-/- mice

Mice that could be highly repopulated with human hepatocytes would have many potential uses in drug development and research applications. The best available model of liver humanization, the uroplasminogen-activator transgenic model, has major practical limitations. To provide a broadly useful hepatic xenorepopulation system, we generated severely immunodeficient, fumarylacetoacetate hydrolase (Fah)-deficient mice. After pretreatment with a urokinase-expressing adenovirus, these animals could be highly engrafted (up to 90%) with human hepatocytes from multiple sources, including liver biopsies. Furthermore, human cells could be serially transplanted from primary donors and repopulate the liver for at least four sequential rounds. The expanded cells displayed typical human drug metabolism. This system provides a robust platform to produce high-quality human hepatocytes for tissue culture. It may also be useful for testing the toxicity of drug metabolites and for evaluating pathogens dependent on human liver cells for replication.     

Effect of simvastatin in different concentrations on free radicals in A-2058 human melanoma malignum cells—EPR studies

The influence of concentration of simvastatin (SIM) on free radicals in A-2058 human melanoma malignum cells was studied. The proliferation assay for melanoma A-2058 cells with SIM in concentration range from 0.1 to 20 µM was performed. SIM in the concentrations of 0.1, 0.3, and 1 μM only slightly changed the growth of A-2058 cells, but the growth of the cells considerably decreased for higher concentrations of SIM. Free radicals in the cells were examined by an X-band (9.3 GHz) electron paramagnetic resonance (EPR) spectroscopy. o-Semiquinone free radicals with g-factors in the range of 2.0060 to 2.0065 were found in A-2058 cells. The asymmetric broad EPR spectra with linewidths (ΔB_pp) from 0.87 to 1.25 mT were measured. The fast spin-lattice relaxation processes characterized all the tested cells. The free radical concentrations in the all A-2058 cells cultured with SIM were lower than in the control cells. The quenching of free radicals in A-2058 cells depended on concentration of SIM. This effect was the weakest for concentration of SIM of 3 μM. The strongest decrease of free radical concentration caused SIM in concentration of 1 μM.     

Effect of Zinc Sulfate and Zinc Glycine Chelate on Concentrations of Acute Phase Proteins in Chicken Serum and Liver Tissue

Biological Trace Element Research - The aim of the study was to determine how inorganic and organic forms of zinc affect the concentrations of C-reactive protein (CRP), serum amyloid A (SAA),...     

Long-term field study on stabilization of contaminated wastes by growing clonally reproduced Silene vulgaris calamine ecotype

Plant and Soil - Pseudo-metallophyte Silene vulgaris frequently colonizes polluted areas. We investigated whether plants obtained under in vitro conditions can be used to form long-term communities...     

Relationships between the properties of Spitsbergen soil,number and biodiversity of rhizosphere microorganisms,and heavy metal concentration in selected plant species

Plant and Soil - Ethylene-insensitive mutation (ein)-conferred Arabidopsis tolerance to Cd has been reported. However, the mechanisms involved are far from clear. This study explores possible...     

Pertactin-deficient Bordetella pertussis isolates in Poland—a country with whole-cell pertussis primary vaccination

The introduction of pertussis vaccination in the 1950s resulted in a significant decrease in the incidence of disease. However, since the 1990s many highly vaccinated countries have observed the re-emergence of the disease. One of the causes of this phenomenon might be related to the adaptation of Bordetella pertussis to vaccination. The purpose of the presented study was an investigation of the emergence and spread of vaccine antigen-deficient B. pertussis isolates in Poland and genomic characterization of the currently circulating pathogen population using PFGE, MLVA and MAST. The results revealed that all tested isolates expressed Ptx, FHA and ACT antigens but 15.4% (4/26) of isolates from 2010 to 2016 were Prn-deficient. Moreover, one TcfA-deficient isolate was collected in 2015. The genotyping showed a genetic distinction between the isolates circulating in 2010–2016 and isolates from previous periods. The majority of currently circulating isolates belonged to PFGE group IV (96.2%), type MT27 (73.1%), and carried ptxA1-ptxC2-ptxP3-prn2-tcfA2-fim2-1-fim3-1 alleles (61.5%). The unique genetic structure of the B. pertussis population in Poland has changed since 2010 and became similar to that observed in countries with aP vaccination. This could be a result of increasing use of aP vaccines (60% of primary vaccination in 2013) over wP vaccines, which have been broadly used for primary vaccination in Poland for decades.     

Affilin Molecules Selected against the Human Papillomavirus E7 Protein Inhibit the Proliferation of Target Cells

Intracellular binding proteins can be applied as research tools for target validation and study of protein function in cells and potentially as therapeutics. The success of intracellular binding reagents depends on their affinity and specificity for target molecules, although their stability and functionality in the intracellular environment actually determine their usefulness for such application. Alternative binding proteins derived from scaffolds devoid of disulfide bonds are well suited for intracellular use, as their folding and stability are usually not impaired under reducing conditions. Here, we describe the generation of intracellular binding reagents called Affilin, based on the human γB-crystallin scaffold. The target was human papillomavirus E7 protein implicated in the development of cervical cancer. E7 binders were selected from the combinatorial γB-crystallin library by conventional phage display technique. Affilin variants specifically bound the E7 protein with affinities in the nanomolar range. Intracellular expression of Affilin molecules in E7-positive cells led to inhibition of cellular proliferation. The effect was specific, as the growth of E7-negative cells or cells expressing the wild-type γB-crystallin scaffold remained unaffected. These results demonstrate that the γB-crystallin scaffold allows the de novo generation of alternative binding proteins, which are suitable for intracellular applications as they retain their functionality in the reducing environment of mammalian cells.     

The influence of abscisic acid on different polysomal populations in embryonal tissue during pea seeds germination

The influence of abscisic acid (ABA) on the processes of formation of different polysomal populations, their structures and stability in embryonal tissue during pea seeds germination was studied. The contents of total ribosomal fraction increased in all samples up to 72 h of germination and then decreased. The contents of polysomal population (FP, MBP, CBP and CMBP) extracted from the embryonal tissue after 72 hrs of germination of pea seeds were then quantified. It turned out that in examined tissue of control sample, fraction of free polysomes (FP) was the most abounded. This population of polysomes in sprouts decreased after ABA treatment. FP content decreased even more when the higher ABA concentration was applied during germination. Similar changes were observed in the fraction of membrane-bound polysomes (MBP). Quite different tendencies were found, however, in forming population of the cytoskeleton-membrane-bound polysomes (CMBP). The CMBP population content in embryonal tissue increased in a dosage dependent manner with increasing concentration of ABA applied during seed germination. This indicates the important role of CMBP fraction in synthesis of specific proteins in embryos in the time when processes of seeds germination are retarded by ABA. In the final part we examined the stability of polysomes isolated from sprouts of germinating seeds in water and sprouts isolated from seeds treated with ABA (100 μM) during germination. Total polysomes isolated from embryonal tissue of germinating seeds treated with ABA showed much higher resistance to exogenous ribonuclease digestion than total polysomes of control sample. The obtained results suggest that ABA influence on different polysomal population formation also controls their stability.     

Seasonal Changes of Flavonoid Content in Melittis melissophyllum L. (Lamiaceae)

Melittis melissophyllum L., a medicinal plant currently used in the folk medicine, was analyzed for the content of flavonoid compounds. The plants were collected in two locations in Poland in May and September. MeOH Extracts from the leaves and flowers (separately) were analyzed by HPLC‐DAD. Eight compounds were identified in all the samples and quantitatively analyzed as cinaroside (=luteolin 7‐O‐glucoside), rutin, myricetin, quercitrin, quercetin, luteolin, kaempferol, and apigenin. M. melissophyllum accumulated the highest total amounts of flavonoids in May (flowers: from 258 to 271 mg/100 g dry weight (dw); leaves: from 143 to 155 mg/100 g dw) and significantly lower ones in September (leaves: from 83 to 92 mg/100 g dw). The main compound was cinaroside (May: up to 249 mg/100 g dw; September: up to 43 mg/100 g dw). Advanced multivariate statistical techniques (cluster analysis (CA) and principal component analysis (PCA)) were used to characterize the sample populations and to analyze the data. We report, for the first time, the results of the quantitative analysis of M. melissophyllum flavonoids and seasonal changes in their accumulation. Our results show that the time of harvesting has a significant influence on the flavonoid content in M. melissophyllum, while the geographical location does not have such an effect.