Vascular endothelial cells express a functional fas-receptor due to lack of hemodynamic forces

The fas system is present in atherosclerotic lesions. However, its role in the initiation and progression is still unclear. Here we show that in endothelial cells (EC) the expression of the fas receptor is regulated by flow conditions. The EC of the vascular system are regularly exposed to a range of hemodynamic forces with great impact on cellular structures and functions. Recently it was reported that in endothelial cells the lack of hemodynamic forces as well as irregular flow conditions trigger apoptosis by induction of a mechanosensitive autocrine loop of thrombospondin-1 and the _V3 integrin/integrin-associated protein complex. Here we show that EC cultivated under regular laminar flow conditions are devoid of the fas-receptor whereas cultivation under static conditions as well as under turbulence leads to its expression. Stimulation of the fas-receptor by its ligand increases the amount of apoptotic cells by twofold; the increase can be prevented by blocking the fas-receptor. The availability of the expressed fas receptor for stimulation by its ligand hints at a role as a tool for progression of atherosclerosis.     

Acute and sub-acute effects of 2-butenoic acid-3-(diethoxy phosphinothioyl) methyl ester (RPR-II) on testis of albino rat

Acute and sub-acute toxic effects of a novel phosphorothionate coded as RPR-II on testis of albino rats were studied. In acute study rats received a single dose of 12.3 mg/kg of RPR-II and sacrificed after 24 hr. For sub-acute study 0.58 mg/kg/day was administered orally to rats for 10 and 21 days. Acute exposure of rats to RPR-II brought no change either in the gonadosomatic index (GSI) or in the structure of testis or in the serum levels of testosterone. Testis glutathione (GSH) level and glutathione S-transferase (GST) activity was significantly decreased whereas, acid phosphatase (AcP) levels increased significantly at 24 hr post-treatment. On 7th day (withdrawal period) after the cessation of the treatment the GSH, GST, AcP, and AkP levels reached to near control. The sub-acute study revealed a significant decrease in GSI on 10th and 21st day of the treatment. In contrast, a time-dependent and significant increased in GSH level and GST activity was observed on 100th and 21st day of post-treatment, except GSH level on 10th day, which was declined. Due to RPR-II treatment the testis AcP and alkaline phosphatase (AkP) levels were significant at both 10th and 21st day of medication but AcP levels were increased whereas AkP levels decreased. The histopathological studies on day 10th showed considerable loss of spermatozoids in testis and at 21st day complete derangement of cellular organization was observed. Testosterone levels decreased significantly on 10th day and remained significantly low at 21st day. However, withdrawal studies showed a recovery in testis of rat treated with RPR-II. GST, GSH, GSI, AcP and AkP values recovered, testosterone levels were also well recovered but recovery in testis structure remained at a low profile. The present study suggests that RPR-II may cause testicular toxicity in rats affecting the normal functioning of testis and it also gave some new information in withdrawal studies.     

Mechanosensitive induction of apoptosis in fibroblasts is regulated by thrombospondin-1 and integrin associated protein (CD47)

Fibroblasts are cultured in three-dimensional collagen matrices to investigate the effect of mechanical tension on the regulation of apoptosis. Under the influence of mechanical loading, the cells show little apoptosis whereas releasing of tension leads to an increase up to tenfold during the first 24 h and remains constant for further 48 h. An autocrine loop of the integrin _V3/CD47 receptor complex and thrombospondin-1 is identified as the molecular coupling device between mechanical loading and apoptosis: The integrin _V3 is expressed under mechanical loading as well as unloading whereas the CD47 could only be identified after the release of tension. The secreted thrombospondin binds to the active receptor and induces apoptosis. The presented mechanosensitive regulation of apoptosis in fibroblast cultures could be an essential mechanism for the regression of the granulation tissue by apoptosis in the process of wound healing.     

Identification of Arabidopsis rat mutants

Limited knowledge currently exists regarding the roles of plant genes and proteins in the Agrobacterium tumefaciens-mediated transformation process. To understand the host contribution to transformation, we carried out root-based transformation assays to identify Arabidopsis mutants that are resistant to Agrobacterium transformation (rat mutants). To date, we have identified 126 rat mutants by screening libraries of T-DNA insertion mutants and by using various “reverse genetic” approaches. These mutants disrupt expression of genes of numerous categories, including chromatin structural and remodeling genes, and genes encoding proteins implicated in nuclear targeting, cell wall structure and metabolism, cytoskeleton structure and function, and signal transduction. Here, we present an update on the identification and characterization of these rat mutants.     

Real-time PCR for chloroquine sensitivity assay and for pfmdr1-pfcrt single nucleotide polymorphisms in Plasmodium falciparum

Plasmodium falciparum drug resistance is a major problem in malaria endemic areas. Molecular markers and in vitro tests have been developed to study and monitor drug resistance. However, none, used alone, can provide sufficient data concerning the level of drug resistance and to issue precise guidelines for drug use policies in endemic areas. We propose real-time PCR for the simultaneous detection of pfcrt and pfmdr1 genes mutations and to determine the half-maximal inhibitory response (IC(50)) of antimalarial drug. Using hybridization probes and SybrGreen technology on LightCycler instrument, point mutations of pfcrt and pfmdr1 genes have been successfully detected in 161 human blood samples and determination of IC values was applied to chloroquine-sensitive and chloroquine-resistant strains. Moreover, mixed infections caused by P. falciparum clones with wild-type or mutant alleles could be efficiency separated. The aim of this study was not to provide definitive data concerning the rate of mutations in an endemic area, but to describe a powerful method allowing the quantification of DNA for IC(50) determination and the detection of major pfmdr1 and pfcrt mutations.     

Whole genome shotgun sequencing guided by bioinformatics pipelines--an optimized approach for an established technique

While the sequencing of bacterial genomes has become a routine procedure at major sequencing centers, there are still a number of genome projects at small- or medium-size facilities. For these facilities a maximum of control over sequencing, assembling and finishing is essential. At the same time, facilities have to be able to co-operate at minimum costs for the overall project. We have established a pipeline for the distributed sequencing of Alcanivorax borkumensis SK2, Azoarcus sp. BH72, Clavibacter michiganensis subsp. michiganensis NCPPB382, Sorangium cellulosum So ce56 and Xanthomonas campestris pv. vesicatoria 85-10. Our pipeline relies on standard tools (e.g. PHRED/PHRAP, CAP3 and Consed/Autofinish) wherever possible, supplementing them with new tools (BioMake and BACCardI) to achieve the aims described above.     

Coupling cell movement to multicellular development in myxobacteria

The myxobacteria are Gram-negative organisms that are capable of multicellular, social behaviour. In the presence of nutrients, swarms of myxobacteria feed cooperatively by sharing extracellular digestive enzymes, and can prey on other bacteria. When the food supply runs low, they initiate a complex developmental programme that culminates in the production of a fruiting body. Myxobacteria move by gliding and have two, polarly positioned engines to control their motility. The two engines undergo coordinated reversals, and changes in the reversal frequency and speed are responsible for the different patterns of movement that are seen during development. The myxobacteria communicate with each other and coordinate their movements through a cell-contact-dependent signal. Here, the cell movements that culminate in the development of the multicellular fruiting body are reviewed.     

Neurofeedback treatment for attention-deficit/hyperactivity disorder in children: a comparison with methylphenidate

Clinical trials have suggested that neurofeedback may be efficient in treating attention-deficit/hyperactivity disorder (ADHD). We compared the effects of a 3-month electroencephalographic feedback program providing reinforcement contingent on the production of cortical sensorimotor rhythm (12–15 Hz) and beta1 activity (15–18 Hz) with stimulant medication. Participants were N = 34 children aged 8–12 years, 22 of which were assigned to the neurofeedback group and 12 to the methylphenidate group according to their parents' preference. Both neurofeedback and methylphenidate were associated with improvements on all subscales of the Test of Variables of Attention, and on the speed and accuracy measures of the d2 Attention Endurance Test. Furthermore, behaviors related to the disorder were rated as significantly reduced in both groups by both teachers and parents on the IOWA-Conners Behavior Rating Scale. These findings suggest that neurofeedback was efficient in improving some of the behavioral concomitants of ADHD in children whose parents favored a nonpharmacological treatment.     

Organic phosphorus in soil water under a European beech (Fagus sylvatica L.) stand in northeastern Bavaria, Germany: seasonal variability and changes with soil depth

Organically bound phosphorus (P) is a mobile form of phosphorus in many soils and thus its dynamics relevant for the leaching and cycling of this element. Despite its importance, little is known about the chemical composition of dissolved organic P. We studied the concentrations, fluxes, and chemical composition of organic P in forest floor leachates and soil solutions in a Rendzic Leptosol under a 90-year-old European beech (Fagus sylvatica L.) forest over a 27-month period (1997–1999). The chemical composition of organic P was analysed using XAD-8 fractionation and ³¹P-nuclear magnetic resonance (NMR) spectroscopy. Organic P was the dominant P form in forest floor leachates as well as in porewaters of the mineral soil. The largest concentrations of organic P were observed during summer and peaked (330–400 g dissolved organic P l^–1) after rain storms following short dry periods, concurrently with the concentrations of organic carbon (OC). Because of high rainfall, fluxes of organic P (and C) were greatest in autumn although concentrations of organic C and P were lower than in summer. In forest floor leachates, the hydrophilic fraction of dissolved organic matter contained 83 ± 13% of the bulk organic P. In soil solutions from 90 cm depth, organic P was almost exclusively in the hydrophilic fraction. Because of the low retention of the hydrophilic fraction of dissolved organic matter in the mineral soils, concentrations of organic P in soil water remained almost constant with depth. Consequently, organic P contributed > 95% of the total P leached into deeper subsoils. The overall retention of organic P in the weakly developed mineral soils was little and so the average annual fluxes of organic P in subsoils at 90 cm depth (38 mg m^–2) comprised 67% of those from the forest floors (57 mg m^–2) during the study period. Hence, organic P proved to be mobile in the studied soil. ³¹P-NMR spectroscopy confirmed the dominance of organic P species in soil water. Signals due to inorganic P occurred only in spectra of samples collected in winter and spring months. Spectra of samples from summer and autumn revealed traces of condensed phosphates. Due to low P contents, identification of organic P species in samples from winter and spring was not always possible. In summer and autumn, monoester and diester phosphates were the dominant organic species and varied little in their relative distributions. The distribution of organic species changed little from forest floor leachates to the subsoil solutions indicating that the composition of P-containing compounds was not influenced by sorptive interactions or biological transformation.     

Changes in ectonucleotidase activities in rat Sertoli cells during sexual maturation

Sertoli cell maturation is a complex process involving both morphological and biochemical changes. These cells have previously been shown to be targets for extracellular purine structures such as ATP and adenosine. These compounds evoke responses in rat Sertoli cells through the purinoceptor families, P₂X and P₂Y and PA₁. The signals to purinoceptors are usually terminated by the action of ectonucleotidases. In a previous work, we demonstrated that rat Sertoli cells have ecto-ATPdiphosphohydrolase (EC 3.6.1.5), ecto-5-nucleotidase (EC 3.1.3.5) and ecto-adenosine deaminase (ecto-ADA) (EC 3.5.4.4) activities. Here we investigated whether some changes occur during rat Sertoli cell maturation in these activities. Rat Sertoli cells obtained from rats of different ages representing the pre pubertal, mid pubertal and young adult (10-, 18- and 35-day-old, respectively) were cultured and used for different assays. The nucleotide hydrolysis was estimated by measuring the Pi released using a colorimetric method and by HPLC analysis. ATP and ADP hydrolysis was increased 3-fold during sexual maturation. AMP hydrolysis increased 4-fold in 10- to 35-day-old Sertoli cells. Similar results were obtained when we used other substrates to measure the extracellular hydrolysis of nucleotides (GTP, GDP, GMP and IMP). The ecto-ADA activity showed a 2-fold increase in the specific activity (18- to 35-day-old Sertoli cells). The termination of the purine cascade by adenosine degradation was faster in the 35- than in 18-day-old Sertoli cells. Follicle Stimulating Hormone (FSH) influences on the ectonucleotidase activities were investigated in 10- and 18-day-old Sertoli cells and a significant increase in the ATP and ADP hydrolysis was observed. Our results show an increase in the extracellular purine cascade during the Sertoli cell development, indicating a rise in the purine communication inside the seminiferous tubules with rat sexual maturation.