Srf1 Is a Novel Regulator of Phospholipase D Activity and Is Essential to Buffer the Toxic Effects of C16:0 Platelet Activating Factor

During Alzheimer's Disease, sustained exposure to amyloid-β42 oligomers perturbs metabolism of ether-linked glycerophospholipids defined by a saturated 16 carbon chain at the sn-1 position. The intraneuronal accumulation of 1-O-hexadecyl-2-acetyl-sn-glycerophosphocholine (C16:0 PAF), but not its immediate precursor 1-O-hexadecyl-sn-glycerophosphocholine (C16:0 lyso-PAF), participates in signaling tau hyperphosphorylation and compromises neuronal viability. As C16:0 PAF is a naturally occurring lipid involved in cellular signaling, it is likely that mechanisms exist to protect cells against its toxic effects. Here, we utilized a chemical genomic approach to identify key processes specific for regulating the sensitivity of Saccharomyces cerevisiae to alkyacylglycerophosphocholines elevated in Alzheimer's Disease. We identified ten deletion mutants that were hypersensitive to C16:0 PAF and five deletion mutants that were hypersensitive to C16:0 lyso-PAF. Deletion of YDL133w, a previously uncharacterized gene which we have renamed SRF1 (Spo14 Regulatory Factor 1), resulted in the greatest differential sensitivity to C16:0 PAF over C16:0 lyso-PAF. We demonstrate that Srf1 physically interacts with Spo14, yeast phospholipase D (PLD), and is essential for PLD catalytic activity in mitotic cells. Though C16:0 PAF treatment does not impact hydrolysis of phosphatidylcholine in yeast, C16:0 PAF does promote delocalization of GFP-Spo14 and phosphatidic acid from the cell periphery. Furthermore, we demonstrate that, similar to yeast cells, PLD activity is required to protect mammalian neural cells from C16:0 PAF. Together, these findings implicate PLD as a potential neuroprotective target capable of ameliorating disruptions in lipid metabolism in response to accumulating oligomeric amyloid-β42

Expression of Stem Cell Markers in the Human Fetal Kidney

In the human fetal kidney (HFK) self-renewing stem cells residing in the metanephric mesenchyme (MM)/blastema are induced to form all cell types of the nephron till 34th week of gestation. Definition of useful markers is crucial for the identification of HFK stem cells. Because wilms' tumor, a pediatric renal cancer, initiates from retention of renal stem cells, we hypothesized that surface antigens previously up-regulated in microarrays of both HFK and blastema-enriched stem-like wilms' tumor xenografts (NCAM, ACVRIIB, DLK1/PREF, GPR39, FZD7, FZD2, NTRK2) are likely to be relevant markers. Comprehensive profiling of these putative and of additional stem cell markers (CD34, CD133, c-Kit, CD90, CD105, CD24) in mid-gestation HFK was performed using immunostaining and FACS in conjunction with EpCAM, an epithelial surface marker that is absent from the MM and increases along nephron differentiation and hence can be separated into negative, dim or bright fractions. No marker was specifically localized to the MM. Nevertheless, FZD7 and NTRK2 were preferentially localized to the MM and emerging tubules (<10% of HFK cells) and were mostly present within the EpCAMneg and EpCAMdim fractions, indicating putative stem/progenitor markers. In contrast, single markers such as CD24 and CD133 as well as double-positive CD24+CD133+ cells comprise >50% of HFK cells and predominantly co-express EpCAMbright, indicating they are mostly markers of differentiation. Furthermore, localization of NCAM exclusively in the MM and in its nephron progenitor derivatives but also in stroma and the expression pattern of significantly elevated renal stem/progenitor genes Six2, Wt1, Cited1, and Sall1 in NCAM+EpCAM- and to a lesser extent in NCAM+EpCAM+ fractions confirmed regional identity of cells and assisted us in pinpointing the presence of subpopulations that are putative MM-derived progenitor cells (NCAM+EpCAM+FZD7+), MM stem cells (NCAM+EpCAM-FZD7+) or both (NCAM+FZD7+). These results and concepts provide a framework for developing cell selection strategies for human renal cell-based therapies

Prospective study on severe malaria among in-patients at Bombo regional hospital, Tanga, north-eastern Tanzania

In Tanzania, malaria is the major cause of morbidity and mortality, accounting for about 30% of all hospital admissions and around 15% of all hospital deaths. Severe anaemia and cerebral malaria are the two main causes of death due to malaria in Tanga, Tanzania. This was a prospective observational hospital-based study conducted from October 2004 to September 2005. Consent was sought from study participants or guardians in the wards. Finger prick blood was collected from each individual for thick and thin smears, blood sugar levels and haemoglobin estimations by Haemocue machine after admission. A total of 494 patients were clinically diagnosed and admitted as cases of severe malaria. Majority of them (55.3%) were children below the age of 5 years. Only 285 out of the total 494 (57.7%) patients had positive blood smears for malaria parasites. Adults aged 20 years and above had the highest rate of cases with fever and blood smear negative for malaria parasites. Commonest clinical manifestations of severe malaria were cerebral malaria (47.3%) and severe anaemia (14.6%), particularly in the under-fives. Case fatality was 3.2% and majority of the deaths occurred in the under-fives and adults aged 20 years and above with negative blood smears. Proper laboratory diagnosis is crucial for case management and reliable data collection. The non-specific nature of malaria symptomatologies limits the use of clinical diagnosis and the IMCI strategy. Strengthening of laboratory investigations to guide case management is recommended

Six RNA Viruses and Forty-One Hosts: Viral Small RNAs and Modulation of Small RNA Repertoires in Vertebrate and Invertebrate Systems

We have used multiplexed high-throughput sequencing to characterize changes in small RNA populations that occur during viral infection in animal cells. Small RNA-based mechanisms such as RNA interference (RNAi) have been shown in plant and invertebrate systems to play a key role in host responses to viral infection. Although homologs of the key RNAi effector pathways are present in mammalian cells, and can launch an RNAi-mediated degradation of experimentally targeted mRNAs, any role for such responses in mammalian host-virus interactions remains to be characterized. Six different viruses were examined in 41 experimentally susceptible and resistant host systems. We identified virus-derived small RNAs (vsRNAs) from all six viruses, with total abundance varying from “vanishingly rare” (less than 0.1% of cellular small RNA) to highly abundant (comparable to abundant micro-RNAs “miRNAs”). In addition to the appearance of vsRNAs during infection, we saw a number of specific changes in host miRNA profiles. For several infection models investigated in more detail, the RNAi and Interferon pathways modulated the abundance of vsRNAs. We also found evidence for populations of vsRNAs that exist as duplexed siRNAs with zero to three nucleotide 3′ overhangs. Using populations of cells carrying a Hepatitis C replicon, we observed strand-selective loading of siRNAs onto Argonaute complexes. These experiments define vsRNAs as one possible component of the interplay between animal viruses and their hosts

Optimizing suicide prevention programs and their implementation in Europe (OSPI Europe): an evidence-based multi-level approach

Background Suicide and non-fatal suicidal behaviour are significant public health issues in Europe requiring effective preventive interventions. However, the evidence for effective preventive strategies is scarce. The protocol of a European research project to develop an optimized evidence based program for suicide prevention is presented. Method The groundwork for this research has been established by a regional community based intervention for suicide prevention that focuses on improving awareness and care for depression performed within the European Alliance Against Depression (EAAD). The EAAD intervention consists of (1) training sessions and practice support for primary care physicians,(2) public relations activities and mass media campaigns, (3) training sessions for community facilitators who serve as gatekeepers for depressed and suicidal persons in the community and treatment and (4) outreach and support for high risk and self-help groups (e.g. helplines). The intervention has been shown to be effective in reducing suicidal behaviour in an earlier study, the Nuremberg Alliance Against Depression. In the context of the current research project described in this paper (OSPI-Europe) the EAAD model is enhanced by other evidence based interventions and implemented simultaneously and in standardised way in four regions in Ireland, Portugal, Hungary and Germany. The enhanced intervention will be evaluated using a prospective controlled design with the primary outcomes being composite suicidal acts (fatal and non-fatal), and with intermediate outcomes being the effect of training programs, changes in public attitudes, guideline-consistent media reporting. In addition an analysis of the economic costs and consequences will be undertaken, while a process evaluation will monitor implementation of the interventions within the different regions with varying organisational and healthcare contexts. Discussion This multi-centre research seeks to overcome major challenges of field research in suicide prevention. It pools data from four European regions, considerably increasing the study sample, which will be close to one million. In addition, the study will gather important information concerning the potential to transfer this multilevel program to other health care systems. The results of this research will provide a basis for developing an evidence-based, efficient concept for suicide prevention for EU-member states

Involvement of components of the phospholipid-signaling pathway in wound-induced phenylpropanoid metabolism in lettuce (Lactuca sativa) leaf tissue

11 pages, 8 figures.In plant tissue, a wound signal is produced at the site of injury and propagates or migrates into adjacent tissue where it induces increased phenylalanine ammonia lyase (PAL, EC 4.3.1.5) activity and phenylpropanoid metabolism. We used excised mid-rib leaf tissue from Romaine lettuce (Lactuca sativa L., Longifolia) as a model system to examine the involvement of components of the phospholipid-signaling pathway in wound-induced phenolic metabolism. Exposure to 1-butanol vapors or solutions inhibited wound-induced increase in PAL activity and phenolic metabolism. Phospholipases D (EC 3.1.4.4), an enzyme involved in the phospholipid-signaling pathway is specifically inhibited by 1-butanol. Re-wounding tissue, in which an effective 1-butanol concentration had declined below active levels by evaporation, did not elicit the normal wound response. It appears the 1-butanol-treated tissue developed resistance to wound-induced increases in phenylpropanoid metabolism that persisted even when active levels of 1-butanol were no longer present. However, a metabolic product of 1-butanol, rather than 1-butanol itself, may be the active compound eliciting persistence resistance. Inhibiting a subsequent enzyme in the phospholipid-signaling pathway, lipoxygenase (LOX; EC 1.13.11.12) with 1-phenyl-3-pyrazolidinone (1P3P) or reducing the product of LOX activity with diethyldithio-carbamic acid (DIECA) also inhibited wound-induced PAL activity and phenolic accumulation. The effectiveness of 1-butanol, DIECA, and 1P3P declined as the beginning of the 1-h immersion period was delayed from 0 to 4 h after excision. This decline in effectiveness is consistent with involvement of the inhibitors in the production or propagation of a wound signal. The wound signal in lettuce moves into adjacent tissue at 0.5 cm h−1, so delaying application would allow the signal to move into and induce the wound response in adjacent tissue before the delayed application inhibited synthesis of the signal. Salicylic acid (SA) inhibits allene oxide synthase (AOS, EC 4.2.1.92), another enzyme in the phospholipid-signaling pathway. Exposure to 1 or 10 mM SA for 60 min reduced wound-induced phenolic accumulation by 26 or 56%, respectively. However, 1 mM SA lost its effectiveness if applied 3 h after excision, while 10 mM SA remained effective even when applied 4 h after excision. At 1 mM, SA may be perturbing the wound signal through inhibition of AOS, while at 10 mM it appears to have some generally inhibitory effect on subsequent phenolic metabolism. These data further implicate the phospholipid-signaling pathway in the generation of a wound signal that induces phenolic metabolism in wounded leaf tissue.Peer reviewe