Melanoma cells break down LPA to establish local gradients that drive chemotactic dispersal.

The high mortality of melanoma is caused by rapid spread of cancer cells, which occurs unusually early in tumour evolution. Unlike most solid tumours, thickness rather than cytological markers or differentiation is the best guide to metastatic potential. Multiple stimuli that drive melanoma cell migration have been described, but it is not clear which are responsible for invasion, nor if chemotactic gradients exist in real tumours. In a chamber-based assay for melanoma dispersal, we find that cells migrate efficiently away from one another, even in initially homogeneous medium. This dispersal is driven by positive chemotaxis rather than chemorepulsion or contact inhibition. The principal chemoattractant, unexpectedly active across all tumour stages, is the lipid agonist lysophosphatidic acid (LPA) acting through the LPA receptor LPAR1. LPA induces chemotaxis of remarkable accuracy, and is both necessary and sufficient for chemotaxis and invasion in 2-D and 3-D assays. Growth factors, often described as tumour attractants, cause negligible chemotaxis themselves, but potentiate chemotaxis to LPA. Cells rapidly break down LPA present at substantial levels in culture medium and normal skin to generate outward-facing gradients. We measure LPA gradients across the margins of melanomas in vivo, confirming the physiological importance of our results. We conclude that LPA chemotaxis provides a strong drive for melanoma cells to invade outwards. Cells create their own gradients by acting as a sink, breaking down locally present LPA, and thus forming a gradient that is low in the tumour and high in the surrounding areas. The key step is not acquisition of sensitivity to the chemoattractant, but rather the tumour growing to break down enough LPA to form a gradient. Thus the stimulus that drives cell dispersal is not the presence of LPA itself, but the self-generated, outward-directed gradient

Formin 1-Isoform IV Deficient Cells Exhibit Defects in Cell Spreading and Focal Adhesion Formation

Background: Regulation of the cytoskeleton is a central feature of cell migration. The formin family of proteins controls the rate of actin nucleation at its barbed end. Thus, formins are predicted to contribute to several important cell processes such as locomotion, membrane ruffling, vesicle endocytosis, and stress fiber formation and disassociation. Methodology/Principal Findings: In this study we investigated the functional role of Formin1-isoform4 (Fmn1-IV) by using genetically null primary cells that displayed augmented protrusive behaviour during wound healing and delayed cell spreading. Cells deficient of Fmn1-IV also showed reduced efficiency of focal adhesion formation. Additionally, we generated an enhanced green fluorescence protein (EGFP)-fused Fmn1-IV knock-in mouse to monitor the endogenous subcellular localization of Fmn1-IV. Its localization was found within the cytoplasm and along microtubules, yet it was largely excluded from adherens junctions. Conclusions/Significance: It was determined that Fmn1-IV, as an actin nucleator, contributes to protrusion of the cell’s leading edge and focal adhesion formation, thus contributing to cell motility

Evaluation of a standard provision versus an autonomy promotive exercise referral programme: rationale and study design

Background The National Institute of Clinical Excellence in the UK has recommended that the effectiveness of ongoing exercise referral schemes to promote physical activity should be examined in research trials. Recent empirical evidence in health care and physical activity promotion contexts provides a foundation for testing the utility of a Self Determination Theory (SDT) -based exercise referral consultation. Methods/Design Design: An exploratory cluster randomised controlled trial comparing standard provision exercise on prescription with a Self Determination Theory-based (SDT) exercise on prescription intervention. Participants: 347 people referred to the Birmingham Exercise on Prescription scheme between November 2007 and July 2008. The 13 exercise on prescription sites in Birmingham were randomised to current practice (n=7) or to the SDT-based intervention (n=6). Outcomes measured at 3 and 6-months: Minutes of moderate or vigorous physical activity per week assessed using the 7-day Physical Activity Recall; physical health: blood pressure and weight; health status measured using the Dartmouth CO-OP charts; anxiety and depression measured by the Hospital Anxiety and Depression Scale and vitality measured by the subjective vitality score; motivation and processes of change: perceptions of autonomy support from the advisor, satisfaction of the needs for competence, autonomy, and relatedness via physical activity, and motivational regulations for exercise. Discussion This trial will determine whether an exercise referral programme based on Self Determination Theory increases physical activity and other health outcomes compared to a standard programme and will test the underlying SDT-based process model (perceived autonomy support, need satisfaction, motivation regulations, outcomes) via structural equation modelling. Trial registration The trial is registered as Current Controlled trials ISRCTN07682833

Chemotactic response and adaptation dynamics in Escherichia coli

Adaptation of the chemotaxis sensory pathway of the bacterium Escherichia coli is integral for detecting chemicals over a wide range of background concentrations, ultimately allowing cells to swim towards sources of attractant and away from repellents. Its biochemical mechanism based on methylation and demethylation of chemoreceptors has long been known. Despite the importance of adaptation for cell memory and behavior, the dynamics of adaptation are difficult to reconcile with current models of precise adaptation. Here, we follow time courses of signaling in response to concentration step changes of attractant using in vivo fluorescence resonance energy transfer measurements. Specifically, we use a condensed representation of adaptation time courses for efficient evaluation of different adaptation models. To quantitatively explain the data, we finally develop a dynamic model for signaling and adaptation based on the attractant flow in the experiment, signaling by cooperative receptor complexes, and multiple layers of feedback regulation for adaptation. We experimentally confirm the predicted effects of changing the enzyme-expression level and bypassing the negative feedback for demethylation. Our data analysis suggests significant imprecision in adaptation for large additions. Furthermore, our model predicts highly regulated, ultrafast adaptation in response to removal of attractant, which may be useful for fast reorientation of the cell and noise reduction in adaptation.Comment: accepted for publication in PLoS Computational Biology; manuscript (19 pages, 5 figures) and supplementary information; added additional clarification on alternative adaptation models in supplementary informatio

Nogo-B regulates migration and contraction of airway smooth muscle cells by decreasing ARPC 2/3 and increasing MYL-9 expression

<p>Abstract</p> <p>Background</p> <p>Abnormal proliferation, apoptosis, migration and contraction of airway smooth muscle (ASM) cells in airway remodeling in asthma are basically excessive repair responses to a network of inflammatory mediators such as PDGF, but the mechanisms of such responses remain unclear. Nogo-B, a member of the reticulum family 4(RTN4), is known to play a key role in arteriogenesis and tissue repair. Further studies are needed to elucidate the role of Nogo-B in airway smooth muscle abnormalities.</p> <p>Methods</p> <p>A mouse model of chronic asthma was established by repeated OVA inhalation and subjected to Nogo-B expression analysis using immunohistochemistry and Western Blotting. Then, primary human bronchial smooth muscle cells (HBSMCs) were cultured <it>in vitro </it>and a siRNA interference was performed to knockdown the expression of Nogo-B in the cells. The effects of Nogo-B inhibition on PDGF-induced HBSMCs proliferation, migration and contraction were evaluated. Finally, a proteomic analysis was conducted to unveil the underlying mechanisms responsible for the function of Nogo-B.</p> <p>Results</p> <p>Total Nogo-B expression was approximately 3.08-fold lower in chronic asthmatic mice compared to naïve mice, which was obvious in the smooth muscle layer of the airways. Interference of Nogo-B expression by siRNA resulted nearly 96% reduction in mRNA in cultured HBSMCs. In addition, knockdown of Nogo-B using specific siRNA significantly decreased PDGF-induced migration of HBSMCs by 2.3-fold, and increased the cellular contraction by 16% compared to negative controls, but had limited effects on PDGF-induced proliferation. Furthermore, using proteomic analysis, we demonstrate that the expression of actin related protein 2/3 complex subunit 5 (ARPC 2/3) decreased and, myosin regulatory light chain 9 isoform a (MYL-9) increased after Nogo-B knockdown.</p> <p>Conclusions</p> <p>These data define a novel role for Nogo-B in airway remodeling in chronic asthma. Endogenous Nogo-B, which may exert its effects through ARPC 2/3 and MYL-9, is necessary for the migration and contraction of airway smooth muscle cells.</p

Depression, Anxiety and Glucose Metabolism in the General Dutch Population: The New Hoorn Study

BACKGROUND: There is a well recognized association between depression and diabetes. However, there is little empirical data about the prevalence of depressive symptoms and anxiety among different groups of glucose metabolism in population based samples. The aim of this study was to determine whether the prevalence of increased levels of depression and anxiety is different between patients with type 2 diabetes and subjects with impaired glucose metabolism (IGM) and normal glucose metabolism (NGM). METHODOLOGY/PRINCIPAL FINDINGS: Cross-sectional data from a population-based cohort study of 2667 residents, 1261 men and 1406 women aged 40-65 years from the Hoorn region, the Netherlands. Depressive symptoms and anxiety were measured using the Centre for Epidemiologic Studies Depression Scale (CES-D, score >or=16) and the Hospital Anxiety and Depression Scale--Anxiety Subscale (HADS-A, score >or=8), respectively. Glucose metabolism status was determined by oral glucose tolerance test. In the total study population the prevalence of depressive symptoms and anxiety for the NGM, IGM and type 2 diabetes were 12.5, 12.2 and 21.0% (P = 0.004) and 15.0, 15.3 and 19.9% (p = 0.216), respectively. In men, the prevalence of depressive symptoms was 7.7, 9.5 and 19.6% (p<0.001), and in women 16.4, 15.8 and 22.6 (p = 0.318), for participants with NGM, IGM and type 2 diabetes, respectively. Anxiety was not associated with glucose metabolism when stratified for sex. Intergroup differences (NGM vs. IGM and IGM vs. type 2 diabetes) revealed that higher prevalences of depressive symptoms are mainly manifested in participants with type 2 diabetes, and not in participants with IGM. CONCLUSIONS: Depressive symptoms, but not anxiety are associated with glucose metabolism. This association is mainly determined by a higher prevalence of depressive symptoms in participants with type 2 diabetes and not in participants with IGM

Problematic mobile phone use and smartphone addiction across generations: the roles of psychopathological symptoms and smartphone use

Contemporary technological advances have led to a significant increase in using mobile technologies. Recent research has pointed to potential problems as a consequence of mobile overuse, including addiction, financial problems, dangerous use (i.e. whilst driving) and prohibited use (i.e. use in forbidden areas). The aim of this study is to extend previous findings regarding the predictive power of psychopathological symptoms (depression, anxiety and stress), mobile phone use (i.e. calls, SMS, time spent on the phone, as well as the engagement in specific smartphone activities) across Generations X and Y on problematic mobile phone use in a sample of 273 adults. Findings revealed prohibited use and dependence were predicted by calls/day, time on the phone and using social media. Only for dependent mobile phone use (rather than prohibited), stress appeared as significant. Using social media and anxiety significantly predicted belonging to Generation Y, with calls per day predicted belonging to Generation X. This finding suggests Generation Y are more likely to use asynchronous social media-based communication, whereas Generation X engage more in synchronous communication. The findings have implications for prevention and awareness-raising efforts of possibly problematic mobile phone use for educators, parents and individuals, particularly including dependence and prohibited use

Combined Boyden-Flow Cytometry Assay Improves Quantification and Provides Phenotypification of Leukocyte Chemotaxis

Chemotaxis has been studied by classical methods that measure chemotactic and random motility responses in vitro, but these methods do not evaluate the total number and phenotype of migrating leukocytes simultaneously. Our objective was to develop and validate a novel assay, combined Boyden-flow cytometry chemotaxis assay (CBFCA), for simultaneous quantification and phenotypification of migrating leukocytes. CBFCA exhibited several important advantages in comparison to the classic Boyden chemotaxis assay (CBCA): 1) improved precision (intra-assay coefficients of variation (CVs): CBFCA-4.7 and 4.8% vs. CBCA-30.1 and 17.3%; inter-observer CVs: CBFCA-3.6% vs. CBCA 30.1%); 2) increased recovery of cells, which increased assay to provide increased sensitivity; 3) high specificity for determining the phenotype of migrating/attracted leukocytes; and 4) reduced performance time (CBFCA 120 min vs. CBCA 265 min). Other advantages of CBFCA are: 5) robustness, 6) linearity, 7) eliminated requirement for albumin and, importantly, 8) enabled recovery of migrating leukocytes for subsequent studies. This latter feature is of great benefit in the study of migrating leukocyte subsets. We conclude that the CBFCA is a novel and improved technique for experiments focused on understanding leukocyte trafficking during the inflammatory response

Locomotion Guidance by Extracellular Matrix Is Adaptive and Can be Restored by a Transient Change in Ca2+ Level

Navigation of cell locomotion by gradients of soluble factors can be desensitized if the concentration of the chemo-attractant stays unchanged. It remains obscure if the guidance by immobilized extracellular matrix (ECM) as the substrate is also adaptive and if so, how can the desensitized ECM guidance be resensitized. When first interacting with a substrate containing micron-scale fibronectin (FBN) trails, highly motile fish keratocytes selectively adhere and migrate along the FBN paths. However, such guided motion become adaptive after about 10 min and the cells start to migrate out of the ECM trails. We found that a burst increase of intracellular calcium created by an uncaging technique immediately halts the undirected migration by disrupting the ECM-cytoskeleton coupling, as evidenced by the appearance of retrograde F-actin flow. When the motility later resumes, the activated integrin receptors render the cell selectively binding to the FBN path and reinitiates signaling events, including tyrosine phosphorylation of paxillin, that couple retrograde F-actin flow to the substrate. Thus, the calcium-resensitized cell can undergo a period of ECM-navigated movement, which later becomes desensitized. Our results also suggest that endogenous calcium transients as occur during spontaneous calcium oscillations may exert a cycling resensitization-desensitization control over cell's sensing of substrate guiding cues

Murine Missing in Metastasis (MIM) Mediates Cell Polarity and Regulates the Motility Response to Growth Factors

Missing in metastasis (MIM) is a member of the inverse BAR-domain protein family, and in vitro studies have implied MIM plays a role in deforming membrane curvature into filopodia-like protrusions and cell dynamics. Yet, the physiological role of the endogenous MIM in mammalian cells remains undefined.We have examined mouse embryonic fibroblasts (MEFs) derived from mice in which the MIM locus was targeted by a gene trapping vector. MIM(-/-) MEFs showed a less polarized architecture characterized by smooth edges and fewer cell protrusions as compared to wild type cells, although the formation of filopodia-like microprotrusions appeared to be normal. Immunofluorescent staining further revealed that MIM(-/-) cells were partially impaired in the assembly of stress fibers and focal adhesions but were enriched with transverse actin filaments at the periphery. Poor assembly of stress fibers was apparently correlated with attenuation of the activity of Rho GTPases and partially relieved upon overexpressing of Myc-RhoA(Q63L), a constitutively activated RhoA mutant. MIM(-/-) cells were also spread less effectively than wild type cells during attachment to dishes and substratum. Upon treatment with PDGF MIM(-/-) cells developed more prominent dorsal ruffles along with increased Rac1 activity. Compared to wild type cells, MIM(-/-) cells had a slower motility in the presence of a low percentage of serum-containing medium but migrated normally upon adding growth factors such as 10% serum, PDGF or EGF. MIM(-/-) cells were also partially impaired in the internalization of transferrin, fluorescent dyes, foreign DNAs and PDGF receptor alpha. On the other hand, the level of tyrosine phosphorylation of PDGF receptors was more elevated in MIM depleted cells than wild type cells upon PDGF treatment.Our data suggests that endogenous MIM protein regulates globally the cell architecture and endocytosis that ultimately influence a variety of cellular behaviors, including cell polarity, motility, receptor signaling and membrane ruffling