Estradiol increases VEGF in human breast studied by whole-tissue culture

Sex steroid exposure constitutes a risk factor for breast cancer, but little is known about the effects of sex steroids on the normal breast, largely because of the lack of convenient models. We have developed a method of culturing normal breast tissue ex vivo. We have applied this method to investigate the effects of estradiol and progesterone on the key angiogenic mediator, vascular endothelial growth factor (VEGF), in the breast. Whole breast tissue was obtained from routine reduction mammoplasty. Tissue biopsies were cultured in vitro for 1-3 weeks, and the expression of luminal cytokeratin 18 was determined by immunohistochemistry. As an application, tissue biopsies were treated in vitro for 1 week with or without estradiol or estradiol and progesterone. Estrogen receptor, progesterone receptor, and Ki-67 were analyzed, and VEGF levels were examined by quantitative immunoassay and immunohistochemistry. Whole breast tissue was cultured ex vivo for 1 week with preserved morphology. Increased detachment of the luminal epithelium was observed after 2 weeks. Estradiol increased extracellular levels of VEGF in normal breast tissue biopsy medium. The addition of progesterone had neither stimulatory nor inhibitory effects on secreted VEGF. The method of whole breast tissue culturing thus provide a means by which to explore the biology of normal breast tissue. Our results suggest that estradiol exerts pro-angiogenic effects in normal breast by increasing levels of biologically active VEGF.     

Proteome profiling of Populus euphratica Oliv. upon heat stress

BACKGROUND AND AIMS: Populus euphratica is a light-demanding species ecologically characterized as a pioneer. It grows in shelter belts along riversides, being part of the natural desert forest ecosystems in China and Middle Eastern countries. It is able to survive extreme temperatures, drought and salt stress, marking itself out as an important plant species to study the mechanisms responsible for survival of woody plants under heat stress. METHODS: Heat effects were evaluated through electrolyte leakage on leaf discs, and LT(50) was determined to occur above 50 degrees C. Protein accumulation profiles of leaves from young plants submitted to 42/37 degrees C for 3 d in a phytotron were determined through 2D-PAGE, and a total of 45 % of up- and downregulated proteins were detected. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF)/TOF analysis, combined with searches in different databases, enabled the identification of 82 % of the selected spots. KEY RESULTS: Short-term upregulated proteins are related to membrane destabilization and cytoskeleton restructuring, sulfur assimilation, thiamine and hydrophobic amino acid biosynthesis, and protein stability. Long-term upregulated proteins are involved in redox homeostasis and photosynthesis. Late downregulated proteins are involved mainly in carbon metabolism. CONCLUSIONS: Moderate heat response involves proteins related to lipid biogenesis, cytoskeleton structure, sulfate assimilation, thiamine and hydrophobic amino acid biosynthesis, and nuclear transport. Photostasis is achieved through carbon metabolism adjustment, a decrease of photosystem II (PSII) abundance and an increase of PSI contribution to photosynthetic linear electron flow. Thioredoxin h may have a special role in this process in P. euphratica upon moderate heat exposure.     

Peak exercise capacity estimated from incremental shuttle walking test in patients with COPD: a methodological study

Background

In patients with COPD, both laboratory exercise tests and field walking tests are used to assess physical performance. In laboratory tests, peak exercise capacity in watts (W peak) and/or peak oxygen uptake (VO₂ peak) are assessed, whereas the performance on walking tests usually is expressed as distance walked. The aim of the study was to investigate the relationship between an incremental shuttle walking test (ISWT) and two laboratory cycle tests in order to assess whether W peak could be estimated from an ISWT.

Methods

Ninety-three patients with moderate or severe COPD performed an ISWT, an incremental cycle test (ICT) to measure W peak and a semi-steady-state cycle test with breath-by-breath gas exchange analysis (CPET) to measure VO₂ peak. Routine equations for conversion between cycle tests were used to estimate W peak from measured VO₂ peak (CPET). Conversion equation for estimation of W peak from ISWT was found by univariate regression.

Results

There was a significant correlation between W peak and distance walked on ISWT × body weight (r = 0.88, p < 0.0001). The agreement between W peak measured by ICT and estimated from ISWT was similar to the agreement between measured W peak (ICT) and W peak estimated from measured VO₂ peak by CPET.

Conclusion

Peak exercise capacity measured by an incremental cycle test could be estimated from an ISWT with similar accuracy as when estimated from peak oxygen uptake in patients with COPD.

     

Substituent distribution and clouding behavior of hydroxypropyl methyl cellulose analyzed using enzymatic degradation

The distribution of substituents along the polymer backbone will have a strong influence on the properties of modified cellulose. Endoglucanases were used to degrade three different batches of hydroxypropyl methyl cellulose (HPMC) derivatives with similar chemical properties. The phase separation of the HPMCs as a function of temperature, i.e., the clouding behavior, was analyzed prior to degradation. The total amount of unsubstituted glucose was determined using total acid hydrolysis followed by high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD). The products after enzymatic degradation were analyzed with size-exclusion chromatography with online multiangle light scattering and refractive index detection and also with reducing end determination. To further characterize the formed products, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry was employed for analysis of short-chained oligosaccharides. The different endoglucanases showed varying degradation capability of HPMC derivatives, depending on structure of the active site. The investigated HPMCs had different susceptibility to degradation by the endoglucanases. The results showed a difference in substituent distribution between HPMC batches, which could explain the differing clouding behaviors. The batch with the lowest cloud point was shown to contain a higher number of non-degradable, highly substituted regions.     

pyewacket, a new zebrafish fin pigment pattern mutant

Many mutants that disrupt zebrafish embryonic pigment pattern have been isolated, and subsequent cloning of the mutated genes causing these phenotypes has contributed to our understanding of pigment cell development. However, few mutants have been identified that specifically affect development of the adult pigment pattern. Through a mutant screen for adult pigment pattern phenotypes, we identified pyewacket (pye), a novel zebrafish mutant in which development of the adult caudal fin pigment pattern is aberrant. Specifically, pye mutants have fin melanocyte pigment pattern defects and fewer xanthophores than wild-type fins. We mapped pye to an interval where a single gene, the zebrafish ortholog of the human gene DHRSX, is present. pye will be an informative mutant for understanding how xanthophores and melanocytes interact to form the pigment pattern of the adult zebrafish fin.     

Importance of spatial habitat structure on establishment of host defenses against brood parasitism

We used metapopulation dynamics to develop a mathematical simulationmodel for brood parasites and their hosts in order to investigatethe validity of the "spatial habitat structure hypothesis,"which states that a low level of parasite egg rejection in hostpopulations is due to the immigration of acceptor individualsfrom nonparasitized populations. In our model, we varied dispersalrate and the relative carrying capacity of host individualsin parasitized and unparasitized patches. When both the relativecarrying capacity in the parasite-free patch and the dispersalrate increase, the nonparasitized patch will provide more acceptorindividuals to the parasite-prone patch. As the relative carryingcapacity in the parasite-free patch increases, the equilibriumfrequency of rejecters both in the parasite-prone and in theparasite-free patch decreases toward zero for intermediate levelsof the dispersal rate. Although the rejecter strategy is moreadaptive than the acceptor strategy in the parasite-prone patch,large numbers of acceptors are produced in the parasite-freepatch dispersing to the parasitized patch. As the number ofindividuals in the parasite-free patch increases, parasitismrate can be maintained stable at a high equilibrium level inthe parasite-prone patch.     

A study of long-term effects on plasmid-containing Escherichia coli in carbon-limited chemostat using 2D-fluorescence spectrofluorimetry

Strain stability of plasmid-containing recombinant organisms is clearly important for industrial applications. Stability is normally assessed by methods such as selective colony forming units or by simply measuring the recombinant product. These methods are typically performed off-line, are time-consuming, and do not give detailed information on the changes in the metabolism. In the current work, long-term stability of a plasmid-containing strain of Escherichia coli (W3110.shik1) capable of shikimic acid overproduction was studied by means of a 2D-fluorescence sensor (BioView) able to emit and detect light in ranges of 260-560 nm and 300-600 nm, respectively. Long-term carbon-limited chemostat experiments were made under both selective (tetracycline-containing medium) and nonselective conditions. It is shown that the fluorescence spectra provide information about metabolic changes at an earlier stage, thereby giving a noninvasive method for monitoring of strain stability. Further, the fluorescence measurements showed that (i) the metabolic changes in the strain W3110.shik1 with time were qualitatively different in selective and nonselective environment, (ii) plasmid recombination resulted primarily in increased biomass yield, and (iii) a change in metabolism probably involving FAD/FMN and pyridoxal-5-P occurred in all experiments. It was concluded that the strain was not stable in any growth condition for more than about 25 growth generations and even less if plasmid recombination took place.     

Genome-wide mapping of susceptibility to coronary artery disease identifies a novel replicated locus on chromosome 17

Coronary artery disease (CAD) is a leading cause of death world-wide, and most cases have a complex, multifactorial aetiology that includes a substantial heritable component. Identification of new genes involved in CAD may inform pathogenesis and provide new therapeutic targets. The PROCARDIS study recruited 2,658 affected sibling pairs (ASPs) with onset of CAD before age 66 y from four European countries to map susceptibility loci for CAD. ASPs were defined as having CAD phenotype if both had CAD, or myocardial infarction (MI) phenotype if both had a MI. In a first study, involving a genome-wide linkage screen, tentative loci were mapped to Chromosomes 3 and 11 with the CAD phenotype (1,464 ASPs), and to Chromosome 17 with the MI phenotype (739 ASPs). In a second study, these loci were examined with a dense panel of grid-tightening markers in an independent set of families (1,194 CAD and 344 MI ASPs). This replication study showed a significant result on Chromosome 17 (MI phenotype; p = 0.009 after adjustment for three independent replication tests). An exclusion analysis suggests that further genes of effect size λ_sib > 1.24 are unlikely to exist in these populations of European ancestry. To our knowledge, this is the first genome-wide linkage analysis to map, and replicate, a CAD locus. The region on Chromosome 17 provides a compelling target within which to identify novel genes underlying CAD. Understanding the genetic aetiology of CAD may lead to novel preventative and/or therapeutic strategies.