EXPRESSION OF mRNA FOR CYTOKINES COMPARED TO THEIR CONCENTRATIONS IN CULTURE SUPERNATES OF U937 CELLS EXPOSED TO POLYCLONAL ACTIVATORS

Investigation of the cytokine expression dynamics as well as the cytokine-producing potential of immune-competent cells allows extensive studies of their functional characteristics. mRNAs encoding a number of cytokine genes are relatively stable, thus their level may be used as a marker for assessing the levels of activation and proliferation of immunocompetent cells as well as for evaluating the cytokine-producing potential of immunocompetent cells.In our work, we assessed correlations between the levels of mRNA expression specific for IL-10, TNF-α, GM-CSF cytokines determined in a culture of differentiated macrophage U937 cells, and protein concentrations of the same cytokines as measured in supernates of U937 cell cultures, without and after exposure to polyclonal activators. The IL-10, TNF-α, GM-CSF mRNA expression was determined by real-time quantitative polymerase chain reaction. Protein concentrations of IL-10, TNF-α, GM-CSF cytokines in the culture supernatant of U937 cells were measured by an enzyme immunoassay. The use of an initially homogeneous cell culture in the study is convenient due to the identical conditions in all experimental variants.The most pronounced effect of polyclonal activators is exerted upon production of GM-CSF mRNA, as well as protein concentration of this cytokine in the cell culture supernatants, thus actually coinciding with RT-qPCR results.  The TNF-α mRNA level decreased under the influence of polyclonal activators, whereas concentration of this cytokine was decreased in the cell supernate. The TNF-α protein in a culture medium did not reflect temporal changes in the cellular TNF-α mRNA expression, probably, due to potential decrease of cellular mRNA occurring by the feedback inhibitory mechanism. While the cytokines can accumulate and remain in the supernatant, the mRNA-related events leading to cytokine formation may be completed earlier. Therefore, the signalling pathways and cytokine release kinetics should be studied after establishing the time dependence at short time intervals, which may be individual for each cytokine.Thus, the results of a study using polyclonal activators suggest that polyclonal activators applied as mitogens, have a significant effect upon the concentration of secreted IL-10, TNFα and GM-CSF. In this case, polyclonal activators affect the levels of mRNA encoded by cytokine genes, thus indicating transcriptional mechanisms of its action. But, in view of the fact that the data are ambiguous, in order to achieve greater correspondence between the changes in the studied proteins and specific mRNAs, a detailed description of the time dependence is required for the changes in mRNA contents

Methanosarcinaceae and acetate-oxidizing pathways dominate in high-rate thermophilic anaerobic digestion of waste-activated sludge

This study investigated the process of high-rate, high-temperature methanogenesis to enable very-high-volume loading during anaerobic digestion of waste-activated sludge. Reducing the hydraulic retention time (HRT) from 15 to 20 days in mesophilic digestion down to 3 days was achievable at a thermophilic temperature (55°C) with stable digester performance and methanogenic activity. A volatile solids (VS) destruction efficiency of 33 to 35% was achieved on waste-activated sludge, comparable to that obtained via mesophilic processes with low organic acid levels

Surplus Photosynthetic Antennae Complexes Underlie Diagnostics of Iron Limitation in a Cyanobacterium

Chlorophyll fluorescence from phytoplankton provides a tool to assess iron limitation in the oceans, but the physiological mechanism underlying the fluorescence response is not understood. We examined fluorescence properties of the model cyanobacterium Synechocystis PCC6803 and a ΔisiA knock-out mutant of the same species grown under three culture conditions which simulate nutrient conditions found in the open ocean: (1) nitrate and iron replete, (2) limiting-iron and high-nitrate, representative of natural high-nitrate, low-chlorophyll regions, and (3) iron and nitrogen co-limiting. We show that low variable fluorescence, a key diagnostic of iron limitation, results from synthesis of antennae complexes far in excess of what can be accommodated by the iron-restricted pool of photosynthetic reaction centers. Under iron and nitrogen co-limiting conditions, there are no excess antennae complexes and variable fluorescence is high. These results help to explain the well-established fluorescence characteristics of phytoplankton in high-nutrient, low-chlorophyll ocean regions, while also accounting for the lack of these properties in low-iron, low-nitrogen regions. Importantly, our results complete the link between unique molecular consequences of iron stress in phytoplankton and global detection of iron stress in natural populations from space

Biomethanation potential of biological and other wastes

Anaerobic technology has been traditionally applied for the treatment of carbon rich wastewater and organic residues. Anaerobic processes can be fully integrated in the biobased economy concept for resource recovery. After a brief introduction about applications of anaerobic processes to industrial wastewater treatment, agriculture feedstock and organic fraction of municipal solid waste, the position of anaerobic processes in biorefinery concepts is presented. Integration of anaerobic digestion with these processes can help in the maximisation of the economic value of the biomass used, while reducing the waste streams produced and mitigating greenhouse gases emissions. Besides the integration of biogas in the existing full-scale bioethanol and biodiesel production processes, the potential applications of biogas in the second generation lignocellulosic, algae and syngas-based biorefinery platforms are discussed.(undefined

Genotoxic effect induced by hydrogen peroxide in human hepatoma cells using comet assay

Background: Hydrogen peroxide is a common reactive oxygen intermediate generated by variousforms of oxidative stress. Aims: The aim of this study was to investigate the DNA damage capacity ofH2O2 in HepG2 cells. Methods: Cells were treated with H2O2 at concentrations of 25 μM or 50 μM for5 min, 30 min, 40 min, 1 h or 24 h in parallel. The extent of DNA damage was assessed by the cometassay. Results: Compared to the control, DNA damage by 25 μM and 50 μM H2O2 increasedsignificantly with increasing incubation time up to 1 h, but it was not increased at 24 h. Conclusions:Our Findings confirm that H2O2 is a typical DNA damage inducing agent and thus is a good modelsystem to study the effects of oxidative stress. DNA damage in HepG2 cells increased significantlywith H2O2 concentration and time of incubation but later decreased likely due to DNA repairmechanisms and antioxidant enzyme