DNAemia detection by multiplex PCR and biomarkers for infection in systemic inflammatory response syndrome patients

Fast and reliable assays to precisely define the nature of the infectious agents causing sepsis are eagerly anticipated. New molecular biology techniques are now available to define the presence of bacterial or fungal DNA within the bloodstream of sepsis patients. We have used a new technique (VYOO?) that allows the enrichment of microbial DNA before a multiplex polymerase chain reaction (PCR) for pathogen detection provided by SIRS-Lab (Jena, Germany). We analyzed 72 sepsis patients and 14 non-infectious systemic inflammatory response syndrome (SIRS) patients. Among the sepsis patients, 20 had a positive blood culture and 35 had a positive microbiology in other biological samples. Of these, 51.4% were positive using the VYOO? test. Among the sepsis patients with a negative microbiology and the non-infectious SIRS, 29.4% and 14.2% were positive with the VYOO? test, respectively. The concordance in bacterial identification between microbiology and the VYOO? test was 46.2%. This study demonstrates that these new technologies offer great hopes, but improvements are still needed.     

Targeted overexpression of osteoactivin in cells of osteoclastic lineage promotes osteoclastic resorption and bone loss in mice

This study sought to test whether targeted overexpression of osteoactivin (OA) in cells of osteoclastic lineage, using the tartrate-resistant acid phosphase (TRAP) exon 1B/C promoter to drive OA expression, would increase bone resorption and bone loss in vivo. OA transgenic osteoclasts showed ～2-fold increases in OA mRNA and proteins compared wild-type (WT) osteoclasts. However, the OA expression in transgenic osteoblasts was not different. At 4, 8, and 15.3 week-old, transgenic mice showed significant bone loss determined by pQCT and confirmed by μ-CT. In vitro, transgenic osteoclasts were twice as large, had twice as much TRAP activity, resorbed twice as much bone matrix, and expressed twice as much osteoclastic genes (MMP9, calciton receptor, and ADAM12), as WT osteoclasts. The siRNA-mediated suppression of OA expression in RAW264.7-derived osteoclasts reduced cell size and osteoclastic gene expression. Bone histomorphometry revealed that transgenic mice had more osteoclasts and osteoclast surface. Plasma c-telopeptide (a resorption biomarker) measurements confirmed an increase in bone resorption in transgenic mice in vivo. In contrast, histomorphometric bone formation parameters and plasma levels of bone formation biomarkers (osteocalcin and pro-collagen type I N-terminal peptide) were not different between transgenic mice and WT littermates, indicating the lack of bone formation effects. In conclusion, this study provides compelling in vivo evidence that osteoclast-derived OA is a novel stimulator of osteoclast activity and bone resorption.     

The Phase Behavior and Organization of Sphingomyelin/Cholesterol Membranes: A Deuterium NMR Study

We have studied the dependence of the phase and domain characteristics of sphingomyelin (SM)/cholesterol model membranes on sterol content and temperature using deuterium nuclear magnetic resonance. NMR spectra of N-palmitoyl(D31)-D-erythro-sphingosylphosphorylcholine (PSM-d31) were taken for temperatures from 25 to 70°C and cholesterol concentrations of 0–40%. Analogous experiments were performed using 1-palmitoyl,2-palmitoyl(D31)-sn-glycero-3-phosphocholine (DPPC-d31)/cholesterol membranes to carefully compare the data obtained using palmitoyl chains that have similar “kinked” conformations. The constructed phase diagrams exhibit both solid-ordered (so) + liquid-ordered (lo) and liquid-disordered (ld) + lo phase-coexistence regions with a clear three-phase line. Macroscopic (micron-sized) coexistence of ld and lo phases was not observed; instead, line-broadening in the ld+lo region was characterized by intermediate exchange of lipids between the two types of domains. The length scales associated with the domains were estimated to be 75–150 nm for PSM-d31/cholesterol and DPPC-d31/cholesterol model membranes.     

The GUDMAP database--an online resource for genitourinary research

The GenitoUrinary Development Molecular Anatomy Project (GUDMAP) is an international consortium working to generate gene expression data and transgenic mice. GUDMAP includes data from large-scale in situ hybridisation screens (wholemount and section) and microarray gene expression data of microdissected, laser-captured and FACS-sorted components of the developing mouse genitourinary (GU) system. These expression data are annotated using a high-resolution anatomy ontology specific to the developing murine GU system. GUDMAP data are freely accessible at www.gudmap.org via easy-to-use interfaces. This curated, high-resolution dataset serves as a powerful resource for biologists, clinicians and bioinformaticians interested in the developing urogenital system. This paper gives examples of how the data have been used to address problems in developmental biology and provides a primer for those wishing to use the database in their own research.     

Inhibition of MLC phosphorylation restricts replication of influenza virus--a mechanism of action for anti-influenza agents

Influenza A viruses are a severe threat worldwide, causing large epidemics that kill thousands every year. Prevention of influenza infection is complicated by continuous viral antigenic changes. Newer anti-influenza agents include MEK/ERK and protein kinase C inhibitors; however, the downstream effectors of these pathways have not been determined. In this study, we identified a common mechanism for the inhibitory effects of a significant group of anti-influenza agents. Our studies showed that influenza infection activates a series of signaling pathways that converge to induce myosin light chain (MLC) phosphorylation and remodeling of the actin cytoskeleton. Inhibiting MLC phosphorylation by blocking RhoA/Rho kinase, phospholipase C/protein kinase C, and HRas/Raf/MEK/ERK pathways with the use of genetic or chemical manipulation leads to the inhibition of influenza proliferation. In contrast, the induction of MLC phosphorylation enhances influenza proliferation, as does activation of the HRas/Raf/MEK/ERK signaling pathway. This effect is attenuated by inhibiting MLC phosphorylation. Additionally, in intracellular trafficking studies, we found that the nuclear export of influenza ribonucleoprotein depends on MLC phosphorylation. Our studies provide evidence that modulation of MLC phosphorylation is an underlying mechanism for the inhibitory effects of many anti-influenza compounds.     

Down syndrome’s brain dynamics: analysis of fractality in resting state

To the best knowledge of the authors there is no study on nonlinear brain dynamics of down syndrome (DS) patients, whereas brain is a highly complex and nonlinear system. In this study, fractal dimension of EEG, as a key characteristic of brain dynamics, showing irregularity and complexity of brain dynamics, was used for evaluation of the dynamical changes in the DS brain. The results showed higher fractality of the DS brain in almost all regions compared to the normal brain, which indicates less centrality and higher irregular or random functioning of the DS brain regions. Also, laterality analysis of the frontal lobe showed that the normal brain had a right frontal laterality of complexity whereas the DS brain had an inverse pattern (left frontal laterality). Furthermore, the high accuracy of 95.8 % obtained by enhanced probabilistic neural network classifier showed the potential of nonlinear dynamic analysis of the brain for diagnosis of DS patients. Moreover, the results showed that the higher EEG fractality in DS is associated with the higher fractality in the low frequencies (delta and theta), in broad regions of the brain, and the high frequencies (beta and gamma), majorly in the frontal regions.     

A Proline/Arginine-Rich End Leucine-Rich Repeat Protein (PRELP) Variant Is Uniquely Expressed in Chronic Lymphocytic Leukemia Cells

Proline/arginine-rich end leucine-rich repeat protein (PRELP) belongs to the small leucine-rich proteoglycan (SLRP) family, normally expressed in extracellular matrix of collagen-rich tissues. We have previously reported on another SLRP, fibromodulin (FMOD) in patients with chronic lymphocytic leukemia (CLL). PRELP is structurally similar to FMOD with adjacent localization on chromosome 1 (1q32.1). As cluster-upregulation of genes may occur in malignancies, the aim of our study was to analyze PRELP expression in CLL. PRELP was expressed (RT-PCR) in all CLL patients (30/30), as well as in some patients with mantle cell lymphoma (3/5), but not in healthy donor leukocytes (0/20) or tumor samples from other hematological malignancies (0/35). PRELP was also detected in CLL cell-lines (4/4) but not in cell-lines from other hematological tumors (0/9). PRELP protein was detected in all CLL samples but not in normal leukocytes. Deglycosylation experiments revealed a CLL-unique 38 kDa core protein, with an intact signal peptide. This 38 kDa protein was, in contrast to the normal 55 kDa size, not detected in serum which, in combination with the uncleaved signal peptide, suggests cellular retention. The unique expression of a 38 kDa PRELP in CLL cells may suggest involvement in the pathobiology of CLL and merits further studies.     

Crosstalk between MMP-13, CD44, and TWIST1 and its role in regulation of EMT in patients with esophageal squamous cell carcinoma

Molecular and Cellular Biochemistry - Matrix metalloproteinases (MMPs) play key roles in epithelial-mesenchymal transition (EMT) for the development of cancer cell invasion and metastasis. MMP-13...