Emerging role of the calcium-activated, small conductance, SK3 K ⁺ channel in distal tubule function: Regulation by TRPV4

The Ca2+-activated, maxi-K (BK) K+ channel, with low Ca2+-binding affinity, is expressed in the distal tubule of the nephron and contributes to flow-dependent K+ secretion. In the present study we demonstrate that the Ca2+-activated, SK3 (KCa2.3) K + channel, with high Ca2+-binding affinity, is also expressed in the mouse kidney (RT-PCR, immunoblots). Immunohistochemical evaluations using tubule specific markers demonstrate significant expression of SK3 in the distal tubule and the entire collecting duct system, including the connecting tubule (CNT) and cortical collecting duct (CCD). In CNT and CCD, main sites for K+ secretion, the highest levels of expression were along the apical (luminal) cell membranes, including for both principal cells (PCs) and intercalated cells (ICs), posturing the channel for Ca2+- dependent K+ secretion. Fluorescent assessment of cell membrane potential in native, split-opened CCD, demonstrated that selective activation of the Ca2+-permeable TRPV4 channel, thereby inducing Ca2+ influx and elevating intracellular Ca2+ levels, activated both the SK3 channel and the BK channel leading to hyperpolarization of the cell membrane. The hyperpolarization response was decreased to a similar extent by either inhibition of SK3 channel with the selective SK antagonist, apamin, or by inhibition of the BK channel with the selective antagonist, iberiotoxin (IbTX). Addition of both inhibitors produced a further depolarization, indicating cooperative effects of the two channels on Vm. It is concluded that SK3 is functionally expressed in the distal nephron and collecting ducts where induction of TRPV4-mediated Ca2+ influx, leading to elevated intracellular Ca2+ levels, activates this high Ca2+- affinity K+ channel. Further, with sites of expression localized to the apical cell membrane, especially in the CNT and CCD, SK3 is poised to be a key pathway for Ca2+-dependent regulation of membrane potential and K+ secretion. © 2014 Berrout et al

A Kinome RNAi Screen Identified AMPK as Promoting Poxvirus Entry through the Control of Actin Dynamics

Poxviruses include medically important human pathogens, yet little is known about the specific cellular factors essential for their replication. To identify genes essential for poxvirus infection, we used high-throughput RNA interference to screen the Drosophila kinome for factors required for vaccinia infection. We identified seven genes including the three subunits of AMPK as promoting vaccinia infection. AMPK not only facilitated infection in insect cells, but also in mammalian cells. Moreover, we found that AMPK is required for macropinocytosis, a major endocytic entry pathway for vaccinia. Furthermore, we show that AMPK contributes to other virus-independent actin-dependent processes including lamellipodia formation and wound healing, independent of the known AMPK activators LKB1 and CaMKK. Therefore, AMPK plays a highly conserved role in poxvirus infection and actin dynamics independent of its role as an energy regulator

Metabolic Regulation of Invadopodia and Invasion by Acetyl-CoA Carboxylase 1 and De novo Lipogenesis

Invadopodia are membrane protrusions that facilitate matrix degradation and cellular invasion. Although lipids have been implicated in several aspects of invadopodia formation, the contributions of de novo fatty acid synthesis and lipogenesis have not been defined. Inhibition of acetyl-CoA carboxylase 1 (ACC1), the committed step of fatty acid synthesis, reduced invadopodia formation in Src-transformed 3T3 (3T3-Src) cells, and also decreased the ability to degrade gelatin. Inhibition of fatty acid synthesis through AMP-activated kinase (AMPK) activation and ACC phosphorylation also decreased invadopodia incidence. The addition of exogenous 16∶0 and 18∶1 fatty acid, products of de novo fatty acid synthesis, restored invadopodia and gelatin degradation to cells with decreased ACC1 activity. Pharmacological inhibition of ACC also altered the phospholipid profile of 3T3-Src cells, with the majority of changes occurring in the phosphatidylcholine (PC) species. Exogenous supplementation with the most abundant PC species, 34∶1 PC, restored invadopodia incidence, the ability to degrade gelatin and the ability to invade through matrigel to cells deficient in ACC1 activity. On the other hand, 30∶0 PC did not restore invadopodia and 36∶2 PC only restored invadopodia incidence and gelatin degradation, but not cellular invasion through matrigel. Pharmacological inhibition of ACC also reduced the ability of MDA-MB-231 breast, Snb19 glioblastoma, and PC-3 prostate cancer cells to invade through matrigel. Invasion of PC-3 cells through matrigel was also restored by 34∶1 PC supplementation. Collectively, the data elucidate the novel metabolic regulation of invadopodia and the invasive process by de novo fatty acid synthesis and lipogenesis

Pan-cancer analysis of whole genomes

Cancer is driven by genetic change, and the advent of massively parallel sequencing has enabled systematic documentation of this variation at the whole-genome scale(1-3). Here we report the integrative analysis of 2,658 whole-cancer genomes and their matching normal tissues across 38 tumour types from the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA). We describe the generation of the PCAWG resource, facilitated by international data sharing using compute clouds. On average, cancer genomes contained 4-5 driver mutations when combining coding and non-coding genomic elements; however, in around 5% of cases no drivers were identified, suggesting that cancer driver discovery is not yet complete. Chromothripsis, in which many clustered structural variants arise in a single catastrophic event, is frequently an early event in tumour evolution; in acral melanoma, for example, these events precede most somatic point mutations and affect several cancer-associated genes simultaneously. Cancers with abnormal telomere maintenance often originate from tissues with low replicative activity and show several mechanisms of preventing telomere attrition to critical levels. Common and rare germline variants affect patterns of somatic mutation, including point mutations, structural variants and somatic retrotransposition. A collection of papers from the PCAWG Consortium describes non-coding mutations that drive cancer beyond those in the TERT promoter(4); identifies new signatures of mutational processes that cause base substitutions, small insertions and deletions and structural variation(5,6); analyses timings and patterns of tumour evolution(7); describes the diverse transcriptional consequences of somatic mutation on splicing, expression levels, fusion genes and promoter activity(8,9); and evaluates a range of more-specialized features of cancer genomes(8,10-18).Peer reviewe

A multi-element psychosocial intervention for early psychosis (GET UP PIANO TRIAL) conducted in a catchment area of 10 million inhabitants: study protocol for a pragmatic cluster randomized controlled trial

Multi-element interventions for first-episode psychosis (FEP) are promising, but have mostly been conducted in non-epidemiologically representative samples, thereby raising the risk of underestimating the complexities involved in treating FEP in 'real-world' services

Alterations in cell maturity and serum survival factors may modulate neutrophil numbers in sickle cell disease

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Leukocytes are known to exacerbate inflammatory and vaso-occlusive processes in sickle cell disease (SCD). The aim of this study was to determine whether alterations in neutrophil maturity and/or cell-death modulating factors in the circulation contribute to the increased leukocyte counts and leukocyte survival observed in SCD. The maturity of circulating neutrophils from healthy control individuals (CON), SCD and SCD patients on hydroxyurea therapy (SCDHU) was determined immunophenotypically. Serum factors affecting neutrophil apoptosis (determined by annexin V-binding) were analyzed by culturing control neutrophils (CON neutrophils) with pooled serum from CON, SCD and SCDHU individuals. Immunophenotypic characterization of neutrophils suggested a slight, but significant, increase in the circulation of immature neutrophils in SCD. While SCD neutrophils cultured in the presence of CON serum presented delayed apoptosis, unexpectedly, the culture of CON neutrophils with SCD serum significantly augmented apoptosis and caspase-9 activity. Inhibition of the activity of serum interleukin-8, a neutrophil-apoptosis-inhibiting cytokine, significantly increased SCD serum-induced CON neutrophil apoptosis, indicating that SCD serum may have both apoptotic and antiapoptotic properties. The decreased maturity of SCD neutrophils observed is suggestive of an accelerated immigration of leukocytes from the bone marrow to the circulating pool that may contribute to an increase in cell survival, subject to modulation by a complex balance of both anti- and proapoptotic factors contained in the circulation of SCD individuals.2361112391246Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

Improvements in motor tasks through the use of smartphone technology for individuals with Duchenne muscular dystrophy

Camila Miliani Capelini,1 Talita Dias da Silva,2 James Tonks,3–5 Suzanna Watson,6 Mayra Priscila Boscolo Alvarez,1 Lilian Del Ciello de Menezes,1 Francis Meire Favero,2 Fátima Aparecida Caromano,1 Thais Massetti,1 Carlos Bandeira de Mello Monteiro1 1Graduate Program in Rehabilitation Sciences, Faculty of Medicine, University of São Paulo, 2Department of Medicine, Paulista School of Medicine, Federal University of São Paulo, São Paulo, SP, Brazil; 3University of Exeter Medical School, Exeter, 4University of Lincoln, Lincoln, 5Haven Clinical Psychology Practice, Cornwall, 6The Cambridge Centre for Paediatric Neuropsychologicial Rehabilitation, Cambridge, UK Background: In individuals severely affected with Duchenne muscular dystrophy (DMD), virtual reality has recently been used as a tool to enhance community interaction. Smartphones offer the exciting potential to improve communication, access, and participation, and present the unique opportunity to directly deliver functionality to people with disabilities.Objective: To verify whether individuals with DMD improve their motor performance when undertaking a visual motor task using a smartphone game.Patients and methods: Fifty individuals with DMD and 50 healthy, typically developing (TD) controls, aged 10–34 years participated in the study. The functional characterization of the sample was determined through Vignos, Egen Klassifikation, and the Motor Function Measure scales. To complete the task, individuals moved a virtual ball around a virtual maze and the time in seconds was measured after every attempt in order to analyze improvement of performance after the practice trials. Motor performance (time to finish each maze) was measured in phases of acquisition, short-term retention, and transfer.Results: Use of the smartphone maze game promoted improvement in performance during acquisition in both groups, which remained in the retention phase. At the transfer phases, with alternative maze tasks, the performance in DMD group was similar to the performance of TD group, with the exception of the transfer to the contralateral hand (nondominant). However, the group with DMD demonstrated longer movement time at all stages of learning, compared with the TD group.Conclusion: The practice of a visual motor task delivered via smartphone game promoted an improvement in performance with similar patterns of learning in both groups. Performance can be influenced by task difficulty, and for people with DMD, motor deficits are responsible for the lower speed of execution. This study indicates that individuals with DMD showed improved performance in a short-term motor learning protocol using a smartphone. We advocate that this technology could be used to promote function in this population. Keywords: motor skills, physical therapy, cell phone, virtual reality exposure therapy, transfer of learnin

Tidal inlets in the Anthropocene: geomorphology and benthic habitats of the Chioggia inlet, Venice Lagoon (Italy)

Within a multidisciplinary approach, we mapped with unprecedented detail the seafloor morphology, sediment distribution and benthic habitats of a tidal inlet which has been highly impacted by human activity. We identified an unusual habitat for lagoon environment connected to rip-rap used for jetties and hard structures and we estimated that the new pattern of flow around these hard structures caused the erosion of 430 000 m(3) of sediment in eight years