TGF-β1 suppresses IL-6-induced STAT3 activation through regulation of Jak2 expression in prostate epithelial cells

Chronic inflammation plays an important role in the initiation and progression of various human diseases including benign prostatic hyperplasia or prostate cancer. Here we show that the proinflammatory cytokine interleukin-6 (IL-6) has prosurvival effects and chronically activates the Jak2/STAT3 signalling pathway in a model of benign prostatic hyperplasia (BPH-1). We demonstrate that the antiinflammatory cytokine transforming growth factor-β1 (TGF-β1), which also permanently activates its canonical signalling pathway through SMAD proteins in BPH-1 cells, modifies the effects of IL-6 on cell proliferation. Importantly, TGF-β1 inhibits IL-6 signal transduction by decreasing the phosphorylation levels of STAT3. This effect is associated with decreased expression of Jak2 at both mRNA and protein levels. Moreover, we showed that TGF-β1 inhibits IL-6-induced expression of the cancer-associated gene MUC1. These observations demonstrated a novel interaction between TGF-β1 and IL-6 signalling and suggested another mechanism of how defects in TGF-β signalling, frequently associated with prostate pathologies, can contribute to the disruption of tissue homeostasis.     

Genetic diversity and its effect on fitness in an endangered plant species, <Emphasis Type="Italic">Dracocephalum austriacum</Emphasis> L.

The aim of this study was to estimate genetic diversity and assess its importance for plant fitness in a species belonging to the most endangered species in Europe, Dracocephalum austriacum L., and to select the most valuable populations for conservation of genetic diversity within the species in the studied regions. We analyzed allozyme variation of 12 populations in three distinct regions (Czech Karst, Moravia and Slovak Karst) in Central Europe. The results showed high genetic diversity within populations (80.14%) and relatively low differentiation among populations within regions (9.42%) and between regions (10.45%). Seed production was significantly higher in larger, genetically more diverse and less inbred populations. The results suggest that genetic diversity has important effect on seed production in this species and thus can be expected to have strong direct consequences for plant fitness and vitality of the whole populations. They also show large variation in genetic diversity between populations and indicate which populations should get a priority in attempts to conserve all the genetic diversity within the region.     

Phosphorylation pattern of the NDUFS4 subunit of complex I of the mammalian respiratory chain

The NDUFS4 subunit of complex I of the mammalian respiratory chain has a fully conserved carboxy-terminus with a canonical RVSTK phosphorylation site. Immunochemical analysis with specific antibodies shows that the serine in this site of the protein is natively present in complex I in both the phosphorylated and non-phosphorylated state. Two-dimensional IEF/SDS–PAGE electrophoresis, ³²P labelling and immunodetection show that “in vitro” PKA phosphorylates the serine in the C-terminus of the NDUFS4 subunit in isolated bovine complex I. ³²P labelling and TLC phosphoaminoacid mapping show that PKA phosphorylates serine and threonine residues in the purified heterologous human NDUFS4 protein.     

4Pi microscopy reveals an impaired three-dimensional mitochondrial network of pancreatic islet β-cells,an experimental model of type-2 diabetes

Insulin production in pancreatic β-cells is critically linked to mitochondrial oxidative phosphorylation. Increased ATP production triggered by blood glucose represents the β-cells' glucose sensor. Type-2 diabetes mellitus results from insulin resistance in peripheral tissues and impaired insulin secretion. Pathology of diabetic β-cells might be reflected by the altered morphology of mitochondrial network. Its characterization is however hampered by the complexity and density of the three-dimensional (3D) mitochondrial tubular networks in these cell types. Conventional confocal microscopy does not provide sufficient axial resolution to reveal the required details; electron tomography reconstruction of these dense networks is still difficult and time consuming. However, mitochondrial network morphology in fixed cells can also be studied by 4Pi microscopy, a laser scanning microscopy technique which provides an ～ 7-fold improved axial resolution (～ 100 nm) over conventional confocal microscopy. Here we present a quantitative study of these networks in insulinoma INS-1E cells and primary β-cells in Langerhans islets. The former were a stably-transfected cell line while the latter were transfected with lentivirus, both expressing mitochondrial matrix targeted redox-sensitive GFP. The mitochondrial networks and their partial disintegration and fragmentation are revealed by carefully created iso-surface plots and their quantitative analysis. We demonstrate that β-cells within the Langerhans islets from diabetic Goto Kakizaki rats exhibited a more disintegrated mitochondrial network compared to those from control Wistar rats and model insulinoma INS-1E cells. Standardization of these patterns may lead to development of morphological diagnostics for Langerhans islets, for the assessment of β-cell condition, before their transplantations.     

Ploidy-specific interactions of three host plants with arbuscular mycorrhizal fungi: Does genome copy number matter?

? Premise of the study: Polyploidy has been shown to affect different plant traits and modulate interactions between plants and other organisms, such as pollinators and herbivores. However, no information is available on whether it can also shape the functioning of mycorrhizal symbiosis. ? Methods: The mycorrhizal growth response was assessed for three angiosperms with intraspecific ploidy variation. Different cytotypes of Aster amellus, Campanula gentilis, and Pimpinella saxifraga were either left uninoculated or were inoculated with arbuscular mycorrhizal (AM) fungi in a pot experiment. After 3 mo of cultivation in a greenhouse, plant growth, phosphorus concentration in the shoot biomass, and development of the AM symbiosis were evaluated. ? Key results: No significant ploidy-specific differences in AM development were recorded. The inoculation led to consistently greater phosphorus uptake; however, the effect on plant growth differed considerably among plant species, populations, ploidy levels, and AM species. A salient ploidy-specific response was observed in A. amellus. Whereas diploid plants benefited from AM inoculation, the hexaploids consistently showed negative or no-growth responses (depending on the AM species). In contrast to A. amellus, no interactions between inoculation and ploidy were observed in C. gentilis and P. saxifraga. ? Conclusions: The first evidence is provided of a ploidy-specific response of a mycotrophic plant to AM fungi. Our results demonstrate the complexity of interaction between plants and associated AM fungi, with the ploidy level of the host plant being one component that may modulate the functioning of the symbiosis.     

<Emphasis Type="Italic">TRAF1/C5</Emphasis>polymorphism is not associated with increased mortality in rheumatoid arthritis: two large longitudinal studies

Introduction  

Recently an association between a genetic variation in TRAF1/C5 and mortality from sepsis or cancer was found in rheumatoid arthritis (RA). The most prevalent cause of death, cardiovascular disease, may have been missed in that study, since patients were enrolled at an advanced disease stage. Therefore, we used an inception cohort of RA patients to investigate the association between TRAF1/C5 and cardiovascular mortality, and replicate the findings on all-cause mortality. As TRAF1/C5 associated mortality may not be restricted to RA, we also studied a large cohort of non-RA patients.     

Inhibition of anion channels derived from mitochondrial membranes of the rat heart by stilbene disulfonate—DIDS

The objective of this work was to characterize in more detail the inhibition effect of diisothiocyanatostilbene-2′,2-disulfonic acid (DIDS) on anion channels isolated from the rat heart mitochondria. The channels reconstituted into a planar lipid membrane displayed limited powers of discrimination between anions and cations and the ion conductance measured under asymmetric (250/50 mM KCl, cis/trans) and symmetric (150 mM KCl) conditions was ∼100 pS. DIDS caused a dramatic decrease in the channel activity (IC₅₀ = 11.7 ± 3.1 μM) only when it was added to the cis side of a planar lipid membrane. The inhibition was accompanied by the significant prolongation of closings and the shortening of openings within the burst as well as gaps between bursts were prolonged and durations of bursts were reduced. The blockade was complete and irreversible when concentration of DIDS was increased up to 200 μM. Our data indicate that DIDS is an allosteric blocker of mitochondrial anion channels and this specific effect could be used as a tool for reliable identification of anion channels on the functional level.     

Suppressive potential of bean (Phaseolus vulgaris) flour against five species of stored-product mites (Acari: Acarididae)

Previous research has demonstrated that legume proteins have insecticidal activity against stored-product pests, but activity against stored-product mites has not been tested. A study was therefore conducted to explore the potential of bean, Phaseolus vulgaris L., flour as novel botanical acaricide against five species of storage and dust mites: Acarus siro L., Aleuroglyphus ovatus (Troupeau), Caloglyphus redickorzevi (Zachvatkin), Lepidoglyphus destructor (Schrank), and Tyrophagus putrescentiae (Schrank). The effect of wheat, Triticum aestivum L., grain enriched with bean flour to eight concentrations (0, 0.01, 0.1, 0.5, 1, 2.5, 5, and 10%) on population growth initiating from the density of 50 mites per 100 g of wheat was recorded for 21 d under laboratory conditions (grain moisture 14.6% moisture content and 25 degree C in darkness). The enrichment of grain with bean flour suppressed the population growth of all tested species: 0.01% concentration reduced population growth of all tested species to >50% in comparison with the control population. The most sensitive species were A. siro and L. destructor, followed by T. putrescentiae and C. redickorzevi. The least sensitive species was A. ovatus. The terminal (i.e., after 21 d) density of mites positively correlated with bean flour concentration. The suppressive effect of bean flour was not linear but rather asymptotic. The results of this study are discussed in the context of the application of bean flour in integrated control of stored-product mites and the elimination of stored-product mite allergens.     

An exodeoxyribonuclease from Streptomyces coelicolor: Expression,purification and biochemical characterization

Streptomyces coelicolor A3(2) produces several intra and extracellular enzymes with deoxyribonuclease activities. The examined N-terminal amino acid sequence of one of extracellular DNAases (TVTSVNVNGLL) and database search on S. coelicolor genome showed a significant homology to the putative secreted exodeoxyribonuclease. The corresponding gene (exoSc) was amplified, cloned, expressed in Escherichia coli, purified to homogeneity and characterized. Exonuclease recExoSc degraded chromosomal, linear dsDNA with 3′-overhang ends, linear ssDNA and did not digest linear dsDNA with blunt ends, supercoiled plasmid ds nor ssDNA. The substrate specificity of recExoSc was in the order of dsDNA > ssDNA > 3′-dAMP. The purified recExoSc was not a metalloprotein and exhibited neither phosphodiesterase nor RNase activity. It acted as 3′-phosphomonoesterase only at 3′-dAMP as a substrate. The optimal temperature for its activity was 57 °C in Tris–HCl buffer at optimal pH = 7.5 for either ssDNA or dsDNA substrates. It required a divalent cation (Mg²⁺, Co²⁺, Ca²⁺) and its activity was strongly inhibited in the presence of Zn²⁺, Hg²⁺, chelating agents or iodoacetate.