Rfx6 Maintains the Functional Identity of Adult Pancreatic β Cells.

SummaryIncreasing evidence suggests that loss of β cell characteristics may cause insulin secretory deficiency in diabetes, but the underlying mechanisms remain unclear. Here, we show that Rfx6, whose mutation leads to neonatal diabetes in humans, is essential to maintain key features of functionally mature β cells in mice. Rfx6 loss in adult β cells leads to glucose intolerance, impaired β cell glucose sensing, and defective insulin secretion. This is associated with reduced expression of core components of the insulin secretion pathway, including glucokinase, the Abcc8/SUR1 subunit of KATP channels and voltage-gated Ca2+ channels, which are direct targets of Rfx6. Moreover, Rfx6 contributes to the silencing of the vast majority of “disallowed” genes, a group usually specifically repressed in adult β cells, and thus to the maintenance of β cell maturity. These findings raise the possibility that changes in Rfx6 expression or activity may contribute to β cell failure in humans

Use of the Internet for health information by physicians for patient care in a teaching hospital in Ibadan, Nigeria

BACKGROUND: The Internet is the world's largest network of information, communication and services. Although the Internet is widely used in medicine and has made significant impact in research, training and patient care, few studies had explored the extent to which Nigerian physicians use Internet resources for patient care. The objective of this study was to assess physicians' use of the Internet for health information for patient care. METHOD: 172 physicians at the University College hospital (UCH) Ibadan, Nigeria; completed a 31-item, anonymous, standardized questionnaire. The Epi-Info software was used for data analysis. RESULTS: The mean age of the respondents was 31.95 years (SD 4.94). Virtually all (98%) the respondents had used the Internet; 76% accessed it from cyber cafes. E-mail was the most commonly used Internet service (64%). Ninety percent of the respondents reported they had obtained information from the Internet for patient care; of this number, 76.2% had searched a database. The database most recently searched was MEDLINE/PubMed in 99% of cases. Only 7% of the respondents had ever searched the Cochrane Library. More than half (58.1%) perceived they had no confidence to download full-text articles from online sources such as the Health Internetwork Access to Research Initiative (HINARI). Multiple barriers to increased use of the Internet were identified including poor availability of broadband (fast connection speed) Internet access, lack of information searching skills, cost of access and information overload. CONCLUSION: Physicians' use of the Internet for health information for patient care was widespread but use of evidenced-based medicine resources such as Cochrane Library, Up-to-date and Clinical Evidence was minimal. Awareness and training in the use of EBM resources for patient care is needed. Introduction of EBM in the teaching curriculum will enhance the use of EBM resources by physicians for patient care

Human Mas-related G protein-coupled receptors-X1 induce chemokine receptor 2 expression in rat dorsal root ganglia neurons and release of chemokine ligand 2 from the human LAD-2 mast cell line

Primate-specific Mas-related G protein-coupled receptors-X1 (MRGPR-X1) are highly enriched in dorsal root ganglia (DRG) neurons and induce acute pain. Herein, we analyzed effects of MRGPR-X1 on serum response factors (SRF) or nuclear factors of activated T cells (NFAT), which control expression of various markers of chronic pain. Using HEK293, DRG neuron-derived F11 cells and cultured rat DRG neurons recombinantly expressing human MRGPR-X1, we found activation of a SRF reporter gene construct and induction of the early growth response protein-1 via extracellular signal-regulated kinases-1/2 known to play a significant role in the development of inflammatory pain. Furthermore, we observed MRGPR-X1-induced up-regulation of the chemokine receptor 2 (CCR2) via NFAT, which is considered as a key event in the onset of neuropathic pain and, so far, has not yet been described for any endogenous neuropeptide. Up-regulation of CCR2 is often associated with increased release of its endogenous agonist chemokine ligand 2 (CCL2). We also found MRGPR-X1-promoted release of CCL2 in a human connective tissue mast cell line endogenously expressing MRGPR-X1. Thus, we provide first evidence to suggest that MRGPR-X1 induce expression of chronic pain markers in DRG neurons and propose a so far unidentified signaling circuit that enhances chemokine signaling by acting on two distinct yet functionally co-operating cell types. Given the important role of chemokine signaling in pain chronification, we propose that interruption of this signaling circuit might be a promising new strategy to alleviate chemokine-promoted pain

NeuriteQuant: An open source toolkit for high content screens of neuronal Morphogenesis

<p>Abstract</p> <p>Background</p> <p>To date, some of the most useful and physiologically relevant neuronal cell culture systems, such as high density co-cultures of astrocytes and primary hippocampal neurons, or differentiated stem cell-derived cultures, are characterized by high cell density and partially overlapping cellular structures. Efficient analytical strategies are required to enable rapid, reliable, quantitative analysis of neuronal morphology in these valuable model systems.</p> <p>Results</p> <p>Here we present the development and validation of a novel bioinformatics pipeline called NeuriteQuant. This tool enables fully automated morphological analysis of large-scale image data from neuronal cultures or brain sections that display a high degree of complexity and overlap of neuronal outgrowths. It also provides an efficient web-based tool to review and evaluate the analysis process. In addition to its built-in functionality, NeuriteQuant can be readily extended based on the rich toolset offered by ImageJ and its associated community of developers. As proof of concept we performed automated screens for modulators of neuronal development in cultures of primary neurons and neuronally differentiated P19 stem cells, which demonstrated specific dose-dependent effects on neuronal morphology.</p> <p>Conclusions</p> <p>NeuriteQuant is a freely available open-source tool for the automated analysis and effective review of large-scale high-content screens. It is especially well suited to quantify the effect of experimental manipulations on physiologically relevant neuronal cultures or brain sections that display a high degree of complexity and overlap among neurites or other cellular structures.</p

Membrane Surface Nanostructures and Adhesion Property of T Lymphocytes Exploited by AFM

The activation of T lymphocytes plays a very important role in T-cell-mediated immune response. Though there are many related literatures, the changes of membrane surface nanostructures and adhesion property of T lymphocytes at different activation stages have not been reported yet. However, these investigations will help us further understand the biophysical and immunologic function of T lymphocytes in the context of activation. In the present study, the membrane architectures of peripheral blood T lymphocytes were obtained by AFM, and adhesion force of the cell membrane were measured by acquiring force–distance curves. The results indicated that the cell volume increased with the increases of activation time, whereas membrane surface adhesion force decreased, even though the local stiffness for resting and activated cells is similar. The results provided complementary and important data to further understand the variation of biophysical properties of T lymphocytes in the context of in vitro activation

Restricted Morphological and Behavioral Abnormalities following Ablation of β-Actin in the Brain

The local translation of β-actin is one mechanism proposed to regulate spatially-restricted actin polymerization crucial for nearly all aspects of neuronal development and function. However, the physiological significance of localized β-actin translation in neurons has not yet been demonstrated in vivo. To investigate the role of β-actin in the mammalian central nervous system (CNS), we characterized brain structure and function in a CNS-specific β-actin knock-out mouse (CNS-ActbKO). β-actin was rapidly ablated in the embryonic mouse brain, but total actin levels were maintained through upregulation of other actin isoforms during development. CNS-ActbKO mice exhibited partial perinatal lethality while survivors presented with surprisingly restricted histological abnormalities localized to the hippocampus and cerebellum. These tissue morphology defects correlated with profound hyperactivity as well as cognitive and maternal behavior impairments. Finally, we also identified localized defects in axonal crossing of the corpus callosum in CNS-ActbKO mice. These restricted defects occurred despite the fact that primary neurons lacking β-actin in culture were morphologically normal. Altogether, we identified novel roles for β-actin in promoting complex CNS tissue architecture while also demonstrating that distinct functions for the ubiquitously expressed β-actin are surprisingly restricted in vivo

Regulation of N-WASP and the Arp2/3 Complex by Abp1 Controls Neuronal Morphology

Polymerization and organization of actin filaments into complex superstructures is indispensable for structure and function of neuronal networks. We here report that knock down of the F-actin-binding protein Abp1, which is important for endocytosis and synaptic organization, results in changes in axon development virtually identical to Arp2/3 complex inhibition, i.e., a selective increase of axon length. Our in vitro and in vivo experiments demonstrate that Abp1 interacts directly with N-WASP, an activator of the Arp2/3 complex, and releases the autoinhibition of N-WASP in cooperation with Cdc42 and thereby promotes N-WASP-triggered Arp2/3 complex-mediated actin polymerization. In line with our mechanistical studies and the colocalization of Abp1, N-WASP and Arp2/3 at sites of actin polymerization in neurons, we reveal an essential role of Abp1 and its cooperativity with Cdc42 in N-WASP-induced rearrangements of the neuronal cytoskeleton. We furthermore show that introduction of N-WASP mutants lacking the ability to bind Abp1 or Cdc42, Arp2/3 complex inhibition, Abp1 knock down, N-WASP knock down and Arp3 knock down, all cause identical neuromorphological phenotypes. Our data thus strongly suggest that these proteins and their complex formation are important for cytoskeletal processes underlying neuronal network formation

CD34 marks angiogenic tip cells in human vascular endothelial cell cultures

The functional shift of quiescent endothelial cells into tip cells that migrate and stalk cells that proliferate is a key event during sprouting angiogenesis. We previously showed that the sialomucin CD34 is expressed in a small subset of cultured endothelial cells and that these cells extend filopodia: a hallmark of tip cells in vivo. In the present study, we characterized endothelial cells expressing CD34 in endothelial monolayers in vitro. We found that CD34-positive human umbilical vein endothelial cells show low proliferation activity and increased mRNA expression of all known tip cell markers, as compared to CD34-negative cells. Genome-wide mRNA profiling analysis of CD34-positive endothelial cells demonstrated enrichment for biological functions related to angiogenesis and migration, whereas CD34-negative cells were enriched for functions related to proliferation. In addition, we found an increase or decrease of CD34-positive cells in vitro upon exposure to stimuli that enhance or limit the number of tip cells in vivo, respectively. Our findings suggest cells with virtually all known properties of tip cells are present in vascular endothelial cell cultures and that they can be isolated based on expression of CD34. This novel strategy may open alternative avenues for future studies of molecular processes and functions in tip cells in angiogenesis