Intratumor versus intertumor heterogeneity in gene expression profiles of soft-tissue sarcomas

Soft-tissue sarcomas (STSs) constitute more than 30 histologic entities. In addition, within each entity, tumors are often heterogeneous in macroscopic features, genetic alterations, microscopic appearance, and clinical course. Therefore, there has been concern about whether a single tumor sample can provide a gene expression profile representative of the entire tumor. We used 27-k cDNA microarray slides to assess the importance of intratumor versus intertumor heterogeneity of the gene expression profiles of 2 morphologically heterogeneous STSs. Multiple pieces of tumor (8 and 10 pieces) were obtained from a myxoid variant of malignant fibrous histiocytoma (MFH) and a leiomyosarcoma (LMS), respectively, and the expression patterns were compared with single tumor samples from 20 MFHs and 16 LMSs. Hierarchical clustering analysis of the expression profiles showed that samples from the same tumor clustered together. The average intratumor distance was considerably shorter than the average intertumor distance in both LMS and MFH. In addition, tumor subclusters that distinguished different macroscopic parts of the tumor could be discerned. We concluded that intratumor variability exists but that accurate gene expression profiling also could be obtained using single samples from a large STS.     

Effect of tissue fixatives on telomere length determination by quantitative PCR

Telomere length is a well established marker of cellular senescence and thus biological age. Quantitative PCR allows the determination even from very low amounts of tissue by using telomere specific and single copy gene primers. Comparing a directly processed tissue sample to a 4% formaldehyde fixed one showed a significantly reduced efficiency of PCR reactions (mainly in single copy gene experiments) in a storage time-dependent manner resulting in an artificial increase in reported relative telomere length. This effect was not seen when the tissue was stored in RNA later solution. In summary, telomere length determination from formaldehyde fixed material by quantitative PCR is not a reliable method. Unfortunately therefore, many easily accessible tissue samples from pathology laboratories are unsuitable for this technique.     

Effects of active recovery on power output during repeated maximal sprint cycling

The effects of active recovery on metabolic and cardiorespiratory responses and power output were examined during repeated sprints. Male subjects (n = 13) performed two maximal 30-s cycle ergometer sprints, 4 min apart, on two separate occasions with either an active [cycling at 40 (1)% of maximal oxygen uptake; mean (SEM)] or passive recovery. Active recovery resulted in a significantly higher mean power output ( ) during sprint 2, compared with passive recovery [ ] 603 (17) W and 589 (15) W, P < 0.05]. This improvement was totally attributed to a 3.1 (1.0)% higher power generation during the initial 10 s of sprint 2 following the active recovery (P < 0.05), since power output during the last 20 s sprint 2 was the same after both recoveries. Despite the higher power output during sprint 2 after active recovery, no differences were observed between conditions in venous blood lactate and pH, but peak plasma ammonia was significantly higher in the active recovery condition [205 (23) vs 170 (20) μmol · 1⁻¹;P < 0.05]. No differences were found between active and passive recovery in terms of changes in plasma volume or arterial blood pressure throughout the test. However, heart rate between the two 30-s sprints and oxygen uptake during the second sprint were higher for the active compared with passive recovery [148 (3) vs 130 (4) beats · min⁻¹;P < 0.01) and 3.3 (0.1) vs 2.8 (0.1) 1 · min⁻¹;P < 0.01]. These data suggest that recovery of power output during repeated sprint exercise is enhanced when low-intensity exercise is performed between sprints. The beneficial effects of an active recovery are possibly mediated by an increased blood flow to the previously exercised muscle.     

The nucleotide sequence of a Polish isolate of Tomato torrado virus

A new virus was isolated from greenhouse tomato plants showing symptoms of leaf and apex necrosis in Wielkopolska province in Poland in 2003. The observed symptoms and the virus morphology resembled viruses previously reported in Spain called Tomato torrado virus (ToTV) and that in Mexico called Tomato marchitez virus (ToMarV). The complete genome of a Polish isolate Wal’03 was determined using RT-PCR amplification using oligonucleotide primers developed against the ToTV sequences deposited in Genbank, followed by cloning, sequencing, and comparison with the sequence of the type isolate. Phylogenetic analyses, performed on the basis of fragments of polyproteins sequences, established the relationship of Polish isolate Wal’03 with Spanish ToTV and Mexican ToMarV, as well as with other viruses from Sequivirus, Sadwavirus, and Cheravirus genera, reported to be the most similar to the new tomato viruses. Wal’03 genome strands has the same organization and very high homology with the ToTV type isolate, showing only some nucleotide and deduced amino acid changes, in contrast to ToMarV, which was significantly different. The phylogenetic tree clustered aforementioned viruses to the same group, indicating that they have a common origin. The nucleotide sequence of the Wal’03 isolate reported in this article has been submitted to the GenBank and is available under accession numbers: EU563948 (RNA1) and EU563947 (RNA2).     

Oligonucleotide microarray-based detection and genotyping of Plum pox virus

Plum pox virus (PPV) is the most damaging viral pathogen of stone fruits. The detection and identification of its strains are therefore of critical importance to plant quarantine and certification programs. Existing methods to screen strains of PPV suffer from significant limitations such as the simultaneous detection and genotyping of several strains of PPV in samples infected with different isolates of the virus. A genomic strategy for PPV screening based on the viral nucleotide sequence was developed to enable the detection and genotyping of the virus from infected plant tissue or biological samples. The basis of this approach is a long 70-mer oligonucleotide DNA microarray capable of simultaneously detecting and genotyping PPV strains. Several 70-mer oligonucleotide probes were specific for the detection and genotyping of individual PPV isolates to their strains. Other probes were specific for the detection and identification of two or three PPV strains. One probe (universal), derived from the genome highly conserved 3' non-translated region, detected all individual strains of PPV. This universal PPV probe, combined with probes specific for each known strain, could be used for new PPV strain discovery. Finally, indirect fluorescent labeling of cDNA with cyanine after cDNA synthesis enhanced the sensitivity of the virus detection without the use of the PCR amplification step. The PPV microarray detected and identified efficiently the PPV strains in PPV-infected peach, apricot and Nicotiana benthamiana leaves. This PPV detection method is versatile, and enables the simultaneous detection of plant pathogens.     

Efficacy and safety of a new apple-flavoured oral rehydration solution in children with acute gastroenteritis: a double-blind randomized controlled trial

Aim: To assess the efficacy and safety of a new oral rehydration solution (ORS) with improved flavour in the management of children with acute gastroenteritis (AGE). Methods: Children 4 to 48 months of age with AGE (≥3 loose or watery stools per day for >1 but <5 days) with mild‐to‐moderate dehydration (3% to 9% loss of body weight) according to the World Health Organization criteria randomly received regular hypotonic ORS (Na 60 mmol/L, glucose 78 mmol/L) or the same hypotonic ORS with an apple taste. Results: Of the 147 children randomized, 130 (88.4%) were available for intention‐to‐treat analysis. The proportion of children with the resolution of signs of dehydration in the experimental group compared with the control group was similar at 24 h (49/63 vs. 57/67, respectively, p = 0.28). There were also no significant differences in adequate weight gain (p = 0.48) and urine production at 24 h (p = 0.95) between groups. There were no differences between groups in any of the secondary outcome measures, including ORS intake. No adverse events were observed in the study groups. Conclusions: In an outpatient setting, there was no difference in efficacy between the study products. Both ORSs were equally effective and may be used interchangeably.     

Effect of exogenous opioid peptides on TNF-alpha-induced human neutrophil apoptosis in vitro

Polymorphonuclear leukocyte (neutrophil) apoptosis is an important mechanism regulating the life span and some functions of neutrophils at inflamed sites. Opioid peptides are present in the peripheral circulation and their concentrations rapidly increase as a result of stress and inflammation. The effect of opioid peptides such as met-enkephalin (M-ENK) and beta-endorphin (beta-END) on tumor necrosis factor (TNF)-alpha-induced apoptosis in human neutrophils in vitro was investigated. Neutrophils isolated from peripheral blood were cultured in the absence or presence of 10(-6)-10(-10) M of opioid peptides for 8, 12 and 18 h. Features of apoptotic neutrophils were measured by a flow cytometric method based on analysis of the apoptotic nuclei (DNA content). We found that M-ENK and beta-END enhanced both uninduced and TNF-alpha-induced neutrophil apoptosis in vitro in a dose-dependent manner. The effect of opioid peptides on the modulation of neutrophil apoptosis was not reversed by the opioid-receptor antagonist naloxone. The results suggest that M-ENK and beta-END can regulate neutrophil life span via apoptosis and in this way may participate in the resolution of inflammation.