Alpha-Toxin Induces Programmed Cell Death of Human T cells, B cells, and Monocytes during USA300 Infection

This investigation examines the influence of alpha-toxin (Hla) during USA300 infection of human leukocytes. Survival of an USA300 isogenic deletion mutant of hla (USA300Δhla) in human blood was comparable to the parental wild-type strain and polymorphonuclear leukocyte (PMN) plasma membrane permeability caused by USA300 did not require Hla. Flow cytometry analysis of peripheral blood mononuclear cells (PBMCs) following infection by USA300, USA300Δhla, and USA300Δhla transformed with a plasmid over-expressing Hla (USA300Δhla Comp) demonstrated this toxin plays a significant role inducing plasma membrane permeability of CD14+, CD3+, and CD19+ PBMCs. Rapid plasma membrane permeability independent of Hla was observed for PMNs, CD14+ and CD19+ PBMCs following intoxication with USA300 supernatant while the majority of CD3+ PBMC plasma membrane permeability induced by USA300 required Hla. Addition of recombinant Hla to USA300Δhla supernatant rescued CD3+ and CD19+ PBMC plasma membrane permeability generated by USA300 supernatant. An observed delay in plasma membrane permeability caused by Hla in conjunction with Annexin V binding and ApoBrdU Tunel assays examining PBMCs intoxicated with recombinant Hla or infected with USA300, USA300Δhla, USA300Δhla Comp, and USA300ΔsaeR/S suggest Hla induces programmed cell death of monocytes, B cells, and T cells that results in plasma membrane permeability. Together these findings underscore the importance of Hla during S. aureus infection of human tissue and specifically demonstrate Hla activity during USA300 infection triggers programmed cell death of human monocytes, T cells and B cells that leads to plasma membrane permeability

Human skeletal muscle plasmalemma alters its structure to change its Ca2+-handling following heavy-load resistance exercise

High-force eccentric exercise results in sustained increases in cytoplasmic Ca2+ levels ([Ca2+]cyto), which can cause damage to the muscle. Here we report that a heavy-load strength training bout greatly alters the structure of the membrane network inside the fibres, the tubular (t-) system, causing the loss of its predominantly transverse organization and an increase in vacuolation of its longitudinal tubules across adjacent sarcomeres. The transverse tubules and vacuoles displayed distinct Ca2+-handling properties. Both t-system components could take up Ca2+ from the cytoplasm but only transverse tubules supported store-operated Ca2+ entry. The retention of significant amounts of Ca2+ within vacuoles provides an effective mechanism to reduce the total content of Ca2+ within the fibre cytoplasm. We propose this ability can reduce or limit resistance exercise-induced, Ca2+-dependent damage to the fibre by the reduction of [Ca2+]cyto to help maintain fibre viability during the period associated with delayed onset muscle soreness

Use of Non-Amplified RNA Samples for Microarray Analysis of Gene Expression

Demand for high quality gene expression data has driven the development of revolutionary microarray technologies. The quality of the data is affected by the performance of the microarray platform as well as how the nucleic acid targets are prepared. The most common method for target nucleic acid preparation includes in vitro transcription amplification of the sample RNA. Although this method requires a small amount of starting material and is reported to have high reproducibility, there are also technical disadvantages such as amplification bias and the long, laborious protocol. Using RNA derived from human brain, breast and colon, we demonstrate that a non-amplification method, which was previously shown to be inferior, could be transformed to a highly quantitative method with a dynamic range of five orders of magnitude. Furthermore, the correlation coefficient calculated by comparing microarray assays using non-amplified samples with qRT-PCR assays was approximately 0.9, a value much higher than when samples were prepared using amplification methods. Our results were also compared with data from various microarray platforms studied in the MicroArray Quality Control (MAQC) project. In combination with micro-columnar 3D-Gene™ microarray, this non-amplification method is applicable to a variety of genetic analyses, including biomarker screening and diagnostic tests for cancer

Thiopurine Methyltransferase Predicts the Extent of Cytotoxicty and DNA Damage in Astroglial Cells after Thioguanine Exposure

Thiopurine methyltransferase (Tpmt) is the primary enzyme responsible for deactivating thiopurine drugs. Thiopurine drugs (i.e., thioguanine [TG], mercaptopurine, azathioprine) are commonly used for the treatment of cancer, organ transplant, and autoimmune disorders. Chronic thiopurine therapy has been linked to the development of brain cancer (most commonly astrocytomas), and Tpmt status has been associated with this risk. Therefore, we investigated whether the level of Tpmt protein activity could predict TG-associated cytotoxicity and DNA damage in astrocytic cells. We found that TG induced cytotoxicity in a dose-dependent manner in Tpmt+/+, Tpmt+/− and Tpmt−/− primary mouse astrocytes and that a low Tpmt phenotype predicted significantly higher sensitivity to TG than did a high Tpmt phenotype. We also found that TG exposure induced significantly more DNA damage in the form of single strand breaks (SSBs) and double strand breaks (DSBs) in primary astrocytes with low Tpmt versus high Tpmt. More interestingly, we found that Tpmt+/− astrocytes had the highest degree of cytotoxicity and genotoxicity (i.e., IC50, SSBs and DSBs) after TG exposure. We then used human glioma cell lines as model astroglial cells to represent high (T98) and low (A172) Tpmt expressers and found that A172 had the highest degree of cytoxicity and SSBs after TG exposure. When we over-expressed Tpmt in the A172 cell line, we found that TG IC50 was significantly higher and SSB's were significantly lower as compared to mock transfected cells. This study shows that low Tpmt can lead to greater sensitivity to thiopurine therapy in astroglial cells. When Tpmt deactivation at the germ-line is considered, this study also suggests that heterozygosity may be subject to the greatest genotoxic effects of thiopurine therapy

Family structure and posttraumatic stress reactions: a longitudinal study using multilevel analyses

<p>Abstract</p> <p>Background</p> <p>There is limited research on the relevance of family structures to the development and maintenance of posttraumatic stress following disasters. We longitudinally studied the effects of marital and parental statuses on posttraumatic stress reactions after the 2004 Southeast Asian tsunami and whether persons in the same households had more shared stress reactions than others.</p> <p>Method</p> <p>The study included a tourist population of 641 Norwegian adult citizens, many of them from families with children. We measured posttraumatic stress symptoms with the Impact of Event Scale-Revised at 6 months and 2 years post-disaster. Analyses included multilevel methods with mixed effects models.</p> <p>Results</p> <p>Results showed that neither marital nor parental status was significantly related to posttraumatic stress. At both assessments, adults living in the same household reported levels of posttraumatic stress that were more similar to one another than adults who were not living together. Between households, disaster experiences were closely related to the variance in posttraumatic stress symptom levels at both assessments. Within households, however, disaster experiences were less related to the variance in symptom level at 2 years than at 6 months.</p> <p>Conclusions</p> <p>These results indicate that adult household members may influence one another's posttraumatic stress reactions as well as their interpretations of the disaster experiences over time. Our findings suggest that multilevel methods may provide important information about family processes after disasters.</p

New Developments in Brief Interventions to Treat Problem Drinking in Nonspecialty Health Care Settings

The delivery of brief interventions (BIs) in health care settings to reduce problematic alcohol consumption is a key preventive strategy for public health. However, evidence of effectiveness beyond primary care is inconsistent. Patient populations and intervention components are heterogeneous. Also, evidence for successful implementation strategies is limited. In this article, recent literature is reviewed covering BI effectiveness for patient populations and subgroups, and design and implementation of BIs. Support is evident for short-term effectiveness in hospital settings, but long-term effects may be confounded by changes in control groups. Limited evidence suggests effectiveness with young patients not admitted as a consequence of alcohol, dependent patients, and binge drinkers. Influential BI components include high-quality change plans and provider characteristics. Health professionals endorse BI and feel confident in delivering it, but training and support initiatives continue to show no significant effects on uptake, prompting calls for systematic approaches to implementing BI in health care

The genomes of two key bumblebee species with primitive eusocial organization

Background: The shift from solitary to social behavior is one of the major evolutionary transitions. Primitively eusocial bumblebees are uniquely placed to illuminate the evolution of highly eusocial insect societies. Bumblebees are also invaluable natural and agricultural pollinators, and there is widespread concern over recent population declines in some species. High-quality genomic data will inform key aspects of bumblebee biology, including susceptibility to implicated population viability threats. Results: We report the high quality draft genome sequences of Bombus terrestris and Bombus impatiens, two ecologically dominant bumblebees and widely utilized study species. Comparing these new genomes to those of the highly eusocial honeybee Apis mellifera and other Hymenoptera, we identify deeply conserved similarities, as well as novelties key to the biology of these organisms. Some honeybee genome features thought to underpin advanced eusociality are also present in bumblebees, indicating an earlier evolution in the bee lineage. Xenobiotic detoxification and immune genes are similarly depauperate in bumblebees and honeybees, and multiple categories of genes linked to social organization, including development and behavior, show high conservation. Key differences identified include a bias in bumblebee chemoreception towards gustation from olfaction, and striking differences in microRNAs, potentially responsible for gene regulation underlying social and other traits. Conclusions: These two bumblebee genomes provide a foundation for post-genomic research on these key pollinators and insect societies. Overall, gene repertoires suggest that the route to advanced eusociality in bees was mediated by many small changes in many genes and processes, and not by notable expansion or depauperation

The prevalence of pelvic organ prolapse symptoms and signs and their relation with bladder and bowel disorders in a general female population

Contains fulltext : 81191.pdf (publisher's version ) (Closed access)INTRODUCTION AND HYPOTHESIS: In selected populations, pelvic organ prolapse (POP) was associated with bladder/bowel symptoms, but data on the general female population are lacking. Our aim was to obtain normative data on the prevalence of POP and pelvic floor dysfunction (PFD) symptoms and signs and to identify associations. METHODS: Validated questionnaires on POP and PFD (urogenital distress inventory, (UDI) and defaecation distress inventory (DDI)) were sent to a general population of 2,979 women (aged 45-85 years). Data were analysed using the Kruskal-Wallis test, chi square test and Spearman's rank correlation coefficient. RESULTS: Response rate was 62.7%. Associations between POP stage and parity (0.002) and vaginal bulging (<0.001) are significant. Anatomical locations of POP and PFD symptoms correlated significantly with incontinence of flatus, feeling anal prolapse, manual evacuation of stool, vaginal bulging, constipation and pain during faecal urge (p < or = 0.005). CONCLUSIONS: Strategies should be developed to alleviate obstructive bowel disorders associated with POP