Extracellular vesicles swarm the cancer microenvironment: from tumor-stroma communication to drug intervention.

Intercellular communication sets the pace for transformed cells to survive and to thrive. Extracellular vesicles (EVs), such as exosomes, microvesicles and large oncosomes, are involved in this process shuttling reciprocal signals and other molecules between transformed and stromal cells, including fibroblasts, endothelial and immune cells. As a result, these cells are adapted or recruited to a constantly evolving cancer microenvironment. Moreover, EVs take part in the response to anticancer therapeutics not least by promoting drug resistance throughout the targeted tumor. Finally, circulating EVs can also transport important molecules to remote destinations in order to prime metastatic niches in an otherwise healthy tissue. Although the understanding of EV biology remains a major challenge in the field, their characteristics create new opportunities for advances in cancer diagnostics and therapeutics.

Plantar pressures increase with raising body weight: A standardised approach with paired sample using neutral shoes.

BACKGROUND: Plantar pressure leads to stress on plantar tissue and can be seen as risk factor for metatarsal stress fractures or plantar ulcers and is associated with prolonged and complicated recurrence of existing tissue damages. A clear demarcation of a systematic raise of body load regarding its effect on plantar pressure has not been described. OBJECTIVE: Assessing plantar pressure patterns in different conditions of body weight, comparing data to initial body weight. METHODS: Seventeen healthy volunteers were asked to participate. Peak pressure values were assessed during walking with dynamic pedobarography and analysed from three foot sections. Body weight was loaded up gradually with 10%, 20% and 30% of the individual initial weight by using a weighted vest. RESULTS: We were able to detect a statistically significant increase of plantar pressure for all foot regions in case of loaded body weight of 20% and 30% comparing to initial weight (p< 0.05). The midfoot area displays a significant increase for peak pressure for the preferred foot even for 10% body load. CONCLUSIONS: Peak plantar pressure increases with loaded body weight. The midfoot area seems to be a sensitive area in case of adapting increasing foot load. Considering the clinical relevance, loaded body weight has to be seen as risk factor for increasing plantar pressure patterns and should be considered in recurrence of plantar ulcers or stress fractures.

Prefrontal dopamine and the dynamic control of human long-term memory.

Dopaminergic projections to the prefrontal cortex support higher-order cognitive functions, and are critically involved in many psychiatric disorders that involve memory deficits, including schizophrenia. The role of prefrontal dopamine in long-term memory, however, is still unclear. We used an imaging genetics approach to examine the hypothesis that dopamine availability in the prefrontal cortex selectively affects the ability to suppress interfering memories. Human participants were scanned via functional magnetic resonance imaging while practicing retrieval of previously studied target information in the face of interference from previously studied non-target information. This retrieval practice (RP) rendered the non-target information less retrievable on a later final test-a phenomenon known as retrieval-induced forgetting (RIF). In total, 54 participants were genotyped for the catechol-O-methyltransferase (COMT) Val(108/158)Met polymorphism. The COMT Val(108/158)Met genotype showed a selective and linear gene-dose effect on RIF, with the Met allele, which leads to higher prefrontal dopamine availability, being associated with greater RIF. Mirroring the behavioral pattern, the functional magnetic resonance imaging data revealed that Met allele carriers, compared with Val allele carriers, showed a greater response reduction in inhibitory control areas of the right inferior frontal cortex during RP, suggesting that they more efficiently reduced interference. These data support the hypothesis that the cortical dopaminergic system is centrally involved in the dynamic control of human long-term memory, supporting efficient remembering via the adaptive suppression of interfering memories.

N-Methylmorpholine-N-oxide ring cleavage registration by ESR under heating conditions of the Lyocell process.

Thermal cleavage processes of N-methylmorpholine-N-oxide monohydrate (NMMO) were observed in pure NMMO as well as in cellulose/NMMO solutions by ESR at temperatures of the industrial Lyocell process ( approximately 370K). Generated radicals were attributed to the alkylnitroxyl type radicals -CH(2)-NO-CH(3) in NMMO and additional (and dominated) -CH(2)-NO-CH(2)- in cellulose/NMMO solutions. Formation of both radical types formed due to NMMO ring scission is suggested.

Electron spin resonance study of radicals generated in cellulose/N-methylmorpholine solutions after flash photolysis at 77 K.

The processes of radical formation in N-methylmorpholine-N-oxide monohydrate (NMMO) and cellulose/NMMO solutions were studied by ESR at 77 K under high-power UV (lambda = 248 nm) excimer laser flash photolysis. Radicals mainly generated were attributed to the nitroxide type radicals -CH2-NO*-CH2- and -CH2-NO*-CH3 at the first step and methyl *CH3 and formyl *CHO radicals at the second step of the photoreaction. Kinetic studies of these radicals revealed that formation and recombination rates of the radicals depend on the cellulose concentration in cellulose/NMMO solutions and the concentration of additional ingredients, e.g. Fe(II) and propyl gallate. Even at frozen state temperature, acceleration or quenching of radical reaction processes was found. The proposed scheme of UV light-induced NMMO degradation during irradiation based on ESR data correlates well with independently obtained results based on high-performance liquid chromatography (HPLC). The analysis of degradation products by HPLC, e.g. aminoethanol and acetaldehyde, supports the assumption concerning a radical-initiated ring opening of NMMO.

Jahn-Teller stabilization of a "polar" metal oxide surface: Fe3O4(001).

Using ab initio thermodynamics we compile a phase diagram for the surface of Fe3O4(001) as a function of temperature and oxygen pressures. A hitherto ignored polar termination with octahedral iron and oxygen forming a wavelike structure along the [110] direction is identified as the lowest energy configuration over a broad range of oxygen gas-phase conditions. This novel geometry is confirmed via x-ray diffraction analysis. The stabilization of the Fe3O4(001) surface goes together with dramatic changes in the electronic and magnetic properties, e.g., a half metal to metal transition.

Syntaxin 6: the promiscuous behaviour of a SNARE protein.

Vesicle flow within the cell is responsible for the dynamic maintenance of and communication between intracellular compartments. In addition, vesicular transport is crucial for communication between the cell and its surrounding environment. The ability of a vesicle to recognise and fuse with an appropriate compartment or vesicle is determined by its protein and lipid composition as well as by proteins in the cytosol. SNARE proteins present on both vesicle as well as target organelle membranes provide one component necessary for the process of membrane fusion. While in mammalian cells the main focus of interest about SNARE function has centred on those involved in exocytosis, recent data on SNAREs involved in intracellular membrane-trafficking steps have provided a deeper insight into the properties of these proteins. We take, as an example, the promiscuous SNARE syntaxin 6, a SNARE involved in multiple membrane fusion events. The properties of syntaxin 6 reveal similarities but also differences in the behaviour of intracellular SNAREs and the highly specialised exocytotic SNARE molecules.

Homotypic fusion of immature secretory granules during maturation requires syntaxin 6.

Homotypic fusion of immature secretory granules (ISGs) gives rise to mature secretory granules (MSGs), the storage compartment in endocrine and neuroendocrine cells for hormones and neuropeptides. With the use of a cell-free fusion assay, we investigated which soluble N-ethylmaleimide-sensitive fusion protein attachment receptor (SNARE) molecules are involved in the homotypic fusion of ISGs. Interestingly, the SNARE molecules mediating the exocytosis of MSGs in neuroendocrine cells, syntaxin 1, SNAP-25, and VAMP2, were not involved in homotypic ISG fusion. Instead, we have identified syntaxin 6 as a component of the core machinery responsible for homotypic ISG fusion. Subcellular fractionation studies and indirect immunofluorescence microscopy show that syntaxin 6 is sorted away during the maturation of ISGs to MSGs. Although, syntaxin 6 on ISG membranes is associated with SNAP-25 and SNAP-29/GS32, we could not find evidence that these target (t)-SNARE molecules are involved in homotypic ISG fusion. Nor could we find any involvement for the vesicle (v)-SNARE VAMP4, which is known to be associated with syntaxin 6. Importantly, we have shown that homotypic fusion requires the function of syntaxin 6 on both donor as well as acceptor membranes, which suggests that t-t-SNARE interactions, either direct or indirect, may be required during fusion of ISG membranes.

Diazaborine resistance in yeast involves the efflux pumps Ycf1p and Flr1p and is enhanced by a gain-of-function allele of gene YAP1.

We have found that YAP1-mediated diazaborine resistance in the yeast Saccharomyces cerevisiae requires two efflux pumps, i.e. the major-facilitator-superfamily transporter Flr1p, which is located in the cytoplasmic membrane and the ATP-binding-cassette transporter Ycf1p which is present in the vacuolar membrane. Both these transporters are known to be under the control of the transcriptional transactivator Yap1p which explains our earlier finding that overexpression of YAP1 mediates diazaborine resistance. Overexpression of YAP1 in a Deltaflr1Deltaycf1 double disruptant strain does not mediate any diazaborine resistance, showing that these pumps are the only ones involved in detoxification of this drug. We also found a new mechanism of diazaborine resistance which is caused by an allelic form of YAP1, designated YAP1-11. This allele of YAP1 carries a mutation that leads to a C620F exchange in the C-terminal cysteine-rich-domain region and is the first mutant of YAP1 that was isolated by a conventional genetic screen for drug resistance. The protein encoded by the gain-of-function allele may transactivate by a different mechanism from the wild-type protein when overexpressed because it does not enhance YCF1 mRNA and still mediates diazaborine resistance in a Deltaflr1Deltaycf1 background.

Diazaborine resistance in the yeast Saccharomyces cerevisiae reveals a link between YAP1 and the pleiotropic drug resistance genes PDR1 and PDR3.

We have investigated the mechanisms underlying resistance to the drug diazaborine in Saccharomyces cerevisiae. We used UV mutagenesis to generate resistant mutants, which were divided into three different complementation groups. The resistant phenotype in these groups was found to be caused by allelic forms of the genes AFG2, PDR1, and PDR3. The AFG2 gene encodes an AAA (ATPases associated to a variety of cellular activities) protein of unknown function, while PDR1 and PDR3 encode two transcriptional regulatory proteins involved in pleiotropic drug resistance development. The isolated PDR1-12 and PDR3-33 alleles carry mutations that lead to a L1044Q and a Y276H exchange, respectively. In addition, we report that overexpression of Yap1p, the yeast homologue of the transcription factor AP1, results in a diazaborine-resistant phenotype. The YAP1-mediated diazaborine resistance is dependent on the presence of functional PDR1 and PDR3 genes, although PDR3 had a more pronounced effect. These results provide the first evidence for a functional link between the Yap1p-dependent stress response pathway and Pdr1p/Pdr3p-dependent development of pleiotropic drug resistance.

Distinct functional properties of three human paired-box-protein, PAX8, isoforms generated by alternative splicing in thyroid, kidney and Wilms' tumors.

The mammalian paired box (Pax) genes encode a family of transcription factors involved in embryogenesis. The murine and human Pax8 genes are expressed in developing and adult thyroid as well as in the developing secretory system and at the lower level in adult kidney. In the secretory system expression is localized to the induced, extensively differentiating parts that undergo a transition from mesenchyme to epithelium. The human PAX8 gene generates at least five different alternatively spliced transcripts encoding different PAX8 isoforms. These isoforms differ in their carboxy-terminal regions downstream of the paired domain that has been shown previously to be responsible for the DNA binding. The PAX8a isoform contains a 63 amino-acid serine-rich region that is absent in the isoform PAX8b whereas PAX8c reveals a novel 99-amino-acid proline-rich region. This proline-rich region arises due to an unusual reading-frame shift in the PAX8 transcript. RNAse protection and RT(reverse transcription)-PCR analysis show the expression of all three PAX8 transcripts in human thyroid, kidney and five Wilms' tumors. Band-shift assay indicates a greatly reduced binding affinity of the isoform PAX8c to a DNA sequence from the promoter of the thyroperoxidase gene compared to the binding of PAX8a and PAX8b to this sequence. Deletion analysis of murine PAX8a indicates that its activating domain residues at the carboxy terminus of the protein which is shared by isoforms PAX8a and PAX8b. In accordance with these data PAX8a and PAX8b activate transcription from a thyroglobulin promoter as well as from a cotransfected synthetic PAX8-specific promoter/chlorampericol acetyltransferase (CAT) reporter containing a Pax8-binding oligonucleotide in front of the basal herpes simplex virus thymidine kinase (HSV-TK) promoter (P11/12-TK-CAT). However if the basal HSV-TK promoter of this reporter is substituted by a minimal adenovirus E1b TATA element, PAX8a and PAX8b fail to activate transcription. Of the three chimaeric forms containing the GAL4 DNA-binding domain at the amino-terminal end fused to the corresponding carboxy-terminal regions of the PAX8 isoforms beginning immediately downstream of the paired domain only a GAL4-PAX8b fusion significantly activates transcription from a cotransfected GAL4-specific upstream-activating-sequence (UAS)-TK-CAT reporter. Substitution of the basal HSV-TK promoter in this reporter by the minimal E1b TATA element does not affect this activation. These results indicate that the PAX8 isoforms display different functional properties and may also function differently in vivo.(ABSTRACT TRUNCATED AT 400 WORDS)