Early changes in ornithine decarboxylase activity during Friend leukemia cell differentiation.

Agents such as dimethylsulfoxide, N,N'-dimethylformamide and bisacetyldiaminopentane that induce erythroid differentiation of Friend leukemia cells, cause a rapid increase in ornithine decarboxylase (EC 4.1.1.17) activity in intact cells during the 'latent' period preceding the accumulation of hemoglobin-containing cells. Blockage of erythroid differentiation with 5-bromo-2'-deoxyuridine did not prevent these alterations in enzyme activity. Addition of each chemical inducer in the extracts of these cells stimulate the basal levels of ornithine decarboxylase activity. These data indicate that the chemical inducers of differentiation modify the normal pattern of ornithine decarboxylase activity in this system.     

The Impact of Biofumigation and Chemical Fumigation Methods on the Structure and Function of the Soil Microbial Community

Biofumigation (BIOF) is carried out mainly by the incorporation of brassica plant parts into the soil, and this fumigation activity has been linked to their high glucosinolate (GSL) content. GSLs are hydrolyzed by the endogenous enzyme myrosinase to release isothiocyanates (ITCs). A microcosm study was conducted to investigate the effects induced on the soil microbial community by the incorporation of broccoli residues into soil either with (BM) or without (B) added myrosinase and of chemical fumigation, either as soil application of 2-phenylethyl ITC (PITC) or metham sodium (MS). Soil microbial activity was evaluated by measuring fluorescein diacetate hydrolysis and soil respiration. Effects on the structure of the total microbial community were assessed by phospholipid fatty acid analysis, while the impact on important fungal (ascomycetes (ASC)) and bacterial (ammonia-oxidizing bacteria (AOB)) guilds was evaluated by denaturating gradient gel electrophoresis (DGGE). Overall, B, and to a lesser extent BM, stimulated microbial activity and biomass. The diminished effect of BM compared to B was particularly evident in fungi and Gram-negative bacteria and was attributed to rapid ITC release following the myrosinase treatment. PITC did not have a significant effect, whereas an inhibitory effect was observed in the MS-treated soil. DGGE analysis showed that the ASC community was temporarily altered by BIOF treatments and more persistently by the MS treatment, while the structure of the AOB community was not affected by the treatments. Cloning of the ASC community showed that MS application had a deleterious effect on potential plant pathogens like Fusarium, Nectria, and Cladosporium compared to BIOF treatments which did not appear to inhibit them. Our findings indicate that BIOF induces changes on the structure and function of the soil microbial community that are mostly related to microbial substrate availability changes derived from the soil amendment with fresh organic materials.     

F₂-isoprostane formation, measurement and interpretation: the role of exercise

The level of F₂-isoprostanes (F₂-IsoP) in blood or urine is widely regarded as the reference marker for the assessment of oxidative stress. As a result, nowadays, F₂-IsoP is the most frequently measured oxidative stress marker. Nevertheless, determining F₂-IsoP is a challenging task and the measurement is neither free of mishaps nor straightforward. This review presents for the first time the effect of acute and chronic exercise on F₂-IsoP levels in plasma, urine and skeletal muscle, placing emphasis on the origin, the methodological caveats and the interpretation of F₂-IsoP alterations. From data analysis, the following effects of exercise have emerged: (i) acute exercise clearly increases F₂-IsoP levels in plasma and this effect is generally short-lived, (ii) acute exercise and increased contractile activity markedly increase F₂-IsoP levels in skeletal muscle, (iii) chronic exercise exhibits trend for decreased F₂-IsoP levels in urine but further research is needed. Theoretically, it seems that significant amounts of F₂-IsoP can be produced not only from phospholipids but from neutral lipids as well. The origin of F₂-IsoP detected in plasma and urine (as done by almost all studies in humans) remains controversial, as a multitude of tissues (including skeletal muscle and plasma) can independently produce F₂-IsoP.     

Interruption of triacylglycerol synthesis in the endoplasmic reticulum is the initiating event for saturated fatty acid-induced lipotoxicity in liver cells

The aim of the present study was to investigate in detail the molecular mechanisms by which free fatty acids induce liver toxicity in liver cells. HepG2 and Huh7 human liver cell lines were exposed to varying concentrations of stearate (18:0), oleate (18:1), or mixtures of the two fatty acids, and the effects on cell proliferation, lipid droplet accumulation and induction of endoplasmic reticulum stress and apoptosis were evaluated. It was observed that: (a) stearate, but not oleate, inhibited cell proliferation and induced cell death; (b) stearate-induced cell death had the characteristics of endoplasmic reticulum stress-mediated and mitochondrial-mediated apoptosis; (c) the activation of stearate in the form of stearoyl-CoA was a necessary step for the lipotoxic effect; (d) the capacity of cells to produce and accumulate triacylglycerols in the form of lipid droplets was interrupted following exposure to stearate, whereas it proceeded normally in oleate-treated cells; and (e) the presence of relatively low amounts of oleate protected cells from stearate-induced toxicity and restored the ability of the cells to accumulate triacylglycerols. Our data suggest that interruption of triacylglycerol synthesis in the endoplasmic reticulum, apparently because of the formation of a pool of oversaturated intermediates, represents the key initiating event in the mechanism of saturated fatty acid-induced lipotoxicity.     

Modelling the Delta1/Notch1 pathway: in search of the mediator(s) of neural stem cell differentiation

The Notch1 signalling pathway has been shown to control neural stem cell fate through lateral inhibition of mash1, a key promoter of neuronal differentiation. Interaction between the Delta1 ligand of a differentiating cell and the Notch1 protein of a neighbouring cell results in cleavage of the trans-membrane protein, releasing the intracellular domain (NICD) leading to the up regulation of hes1. Hes1 homodimerisation leads to down regulation of mash1. Most mathematical models currently represent this pathway up to the formation of the HES1 dimer. Herein, we present a detailed model ranging from the cleavage of the NICD and how this signal propagates through the Delta1/Notch1 pathway to repress the expression of the proneural genes. Consistent with the current literature, we assume that cells at the self renewal state are represented by a stable limit cycle and through in silico experimentation we conclude that a drastic change in the main pathway is required in order for the transition from self-renewal to differentiation to take place. Specifically, a model analysis based approach is utilised in order to generate hypotheses regarding potential mediators of this change. Through this process of model based hypotheses generation and testing, the degradation rates of Hes1 and Mash1 mRNA and the dissociation constant of Mash1-E47 heterodimers are identified as the most potent mediators of the transition towards neural differentiation.     

Mitotic catastrophe occurs in the absence of apoptosis in p53-null cells with a defective G1 checkpoint

Cell death occurring during mitosis, or mitotic catastrophe, often takes place in conjunction with apoptosis, but the conditions in which mitotic catastrophe may exhibit features of programmed cell death are still unclear. In the work presented here, we studied mitotic cell death by making use of a UV-inactivated parvovirus (adeno-associated virus; AAV) that has been shown to induce a DNA damage response and subsequent death of p53-defective cells in mitosis, without affecting the integrity of the host genome. Osteosarcoma cells (U2OSp53DD) that are deficient in p53 and lack the G1 cell cycle checkpoint respond to AAV infection through a transient G2 arrest. We found that the infected U2OSp53DD cells died through mitotic catastrophe with no signs of chromosome condensation or DNA fragmentation. Moreover, cell death was independent of caspases, apoptosis-inducing factor (AIF), autophagy and necroptosis. These findings were confirmed by time-lapse microscopy of cellular morphology following AAV infection. The assays used readily revealed apoptosis in other cell types when it was indeed occurring. Taken together the results indicate that in the absence of the G1 checkpoint, mitotic catastrophe occurs in these p53-null cells predominantly as a result of mechanical disruption induced by centrosome overduplication, and not as a consequence of a suicide signal.     

Isokinetic knee joint evaluation in track and field events

The purpose of this study was to evaluate maximal torque of the knee flexors and extensors, flexor/extensor ratios, and maximal torque differences between the 2 lower extremities in young track and field athletes. Forty male track and field athletes 13-17 years old and 20 male nonathletes of the same age participated in the study. Athletes were divided into 4 groups according to their age and event (12 runners and 10 jumpers 13-15 years old, 12 runners and 6 jumpers 16-17 years old) and nonathletes into 2 groups of the same age. Maximal torque evaluation of knee flexors and extensors was performed on an isokinetic dynamometer at 60°·s(-1). At the age of 16-17 years, jumpers exhibited higher strength values at extension than did runners and nonathletes, whereas at the age of 13-15 years, no significant differences were found between events. Younger athletes were weaker than older athletes at flexion. Runners and jumpers were stronger than nonathletes in all relative peak torque parameters. Nonathletes exhibited a higher flexor/extensor ratio compared with runners and jumpers. Strength imbalance in athletes was found between the 2 lower extremities in knee flexors and extensors and also at flexor/extensor ratio of the same extremity. Young track and field athletes exhibit strength imbalances that could reduce their athletic performance, and specific strength training for the weak extremity may be needed.