Calcium Signaling in Live Cells on Elastic Gels under Mechanical Vibration at Subcellular Levels 英文参考文献.docVIP

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Calcium Signaling in Live Cells on Elastic Gels under Mechanical Vibration at Subcellular Levels 英文参考文献.doc

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Calcium Signaling in Live Cells on Elastic Gels under Mechanical Vibration at Subcellular Levels 英文参考文献

CalciumSignalinginLiveCellsonElasticGelsunder MechanicalVibrationatSubcellularLevels WagnerShinNishitani1,4,TaherA.Saif2*,YingxiaoWang1,3* 1DepartmentofBioengineeringandBeckmanInstituteforAdvancedScienceandTechnology,UniversityofIllinois,Urbana-Champaign,Urbana,Illinois,UnitedStatesof America, 2Department of Mechanical Science and Engineering, University of Illinois, Urbana-Champaign, Urbana, Illinois, United States of America, 3Integrative and MolecularPhysiology,CenterforBiophysicsandComputationalBiology,InstituteforGenomicBiology,UniversityofIllinois,Urbana-Champaign,Urbana,Illinois,United StatesofAmerica,4TheCapesFoundation,MinistryofEducationofBrazil,Bras?′lia,DistritoFederal,Brazil Abstract A new device was designed to generate a localized mechanical vibration of flexible gels where human umbilical vein endothelial cells (HUVECs) were cultured to mechanically stimulate these cells at subcellular locations. A Fluorescence Resonance Energy Transfer (FRET)-based calcium biosensor (an improved Cameleon) was used to monitor the spatiotemporal distribution of intracellular calcium concentrations in the cells upon this mechanical stimulation. A clear increaseinintracellularcalciumconcentrationsoverthewholecellbody(global)canbeobservedinthemajorityofcells undermechanicalstimulation.ThechelationofextracellularcalciumwithEGTAortheblockageofstretch-activatedcalcium channelsontheplasmamembranewithstreptomycinorgadoliniumchloridesignificantlyinhibitedthecalciumresponses upon mechanical stimulation. Thapsigargin, an endoplasmic reticulum (ER) calcium pump inhibitor, or U73122, a phospholipaseC(PLC)inhibitor,resultedinmainlylocalcalciumresponsesoccurringatregionsclosetothestimulationsite. ThedisruptionofactinfilamentswithcytochalasinDorinhibitionofactomyosincontractilitywithML-7alsoinhibitedthe global calcium responses. Therefore, the global calcium response in HUVEC depends on the influx of calcium through membranestretch-activatedchannels,followedbythereleaseofinositol