1. NANOYOUTeachersTrainingKitinNanotechnologiesChapter1MedicineandHealthcareMODULE2ApplicationofNanotechnologiesWrittenbyLuisaFilipponiandDuncanSutherlandInterdisciplinaryNanoscienceCentreAarhusUniversity,DenmarkJanuary2010CreativeCommonsAttributionShareAlike3.0unlessindicatedintextorfigurecaptions.

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NANOYOUTeachersTrainingKitModule2Chapter1ContentsDiagnosis.........................................................................................................................................5Biosensors.................................................................................................................................................................. 6 Cantileverbiosensor.............................................................................................................................................. 7 Plasmonicbiosensors............................................................................................................................................. 8 Artificialnosebiosensor....................................................................................................................................... 01Microarrays............................................................................................................................................................. 1 1Nanobarcodes.......................................................................................................................................................... 2 1LabOnAChip ......................................................................................................................................................... 2 .1Imaging .........................................................................................................................................13 .Diagnosticimaging.................................................................................................................................................. 3 1Insitudiagnosticdevices......................................................................................................................................... 51Therapy.........................................................................................................................................16Drugdevelopmentandtargeteddrugdelivery ....................................................................................................... 6 .1 Drugdesignandscreening.................................................................................................................................. 6 1 Targeteddrugdelivery........................................................................................................................................ 71 Currentandfuturenanodrugcarriers................................................................................................................ 1 2Externallyactivatedtherapiesthatusenanoparticles............................................................................................ 22Theranostic.............................................................................................................................................................. 22Page2of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 3. NANOYOUTeachersTrainingKitModule2Chapter1Regenerativemedicine.................................................................................................................23Tissueandbiomaterialengineering ........................................................................................................................ 3. 2 Nanoengineeringboneregeneration .................................................................................................................. 5. 2 Nanoengineeringneuronregeneration............................................................................................................... 62Neuroprosthetics..................................................................................................................................................... 82 Neuronalstimulation,monitoringandpainmanagement.................................................................................. 03 Noninvasivebrainmachineinterfaces................................................................................................................. 13This document has been created in the context of the NANOYOU project (WP 4 Task 4.1). All information isprovidedasisandnoguaranteeorwarrantyisgiventhattheinformationisfitforanyparticularpurpose.Theuser thereof uses the information at its sole risk and liability. The document reflects solely the views of itsauthors. The European Commission is not liable for any use that may be made of the information contained Page3of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 4. NANOYOUTeachersTrainingKitModule2Chapter1Chapter1: ApplicationsofNanotechnologiesMedicine&HealthcareTheapplicationofnanotechnologiestothemedicalsectorisreferredtoasNanomedicine.Specifically,this area of application uses nanometre scale materials and nanoenabled techniques to diagnosis,monitor, treat and prevent diseases. These include cardiovascular diseases, cancer, musculoskeletaland inflammatory conditions, neurodegenerative and psychiatric diseases, diabetes and infectiousdiseases (bacterial and viral infections, such as HIV), and more. The potential contribution ofnanotechnologiesinthemedicalsectorisextremelybroadandincludesnewdiagnostictools;imagingagents and methods; drug delivery systems and pharmaceuticals; therapies; implants and tissueengineeredconstructs.Why nanotechnologies? Nanomaterials are defined as materials in the nanoscale regime, which innanomedicineoftengoesbeyondthe100nmlinemarkanduptoabout500nm.Theseareexactlythesizerangeofbiomolecules(e.g.,proteins,enzymes,DNAetc.)andmolecularcomplexessuchastheionpump.Thosenaturalnanomaterialsaretheconstituentsoflargerhierarchicalstructureswhichregulatethefunctionofthecell.Bacteriaandvirusesarelarger(fewmicrometers),buttheirfunctions(includingtoxicitytohealthycells)derivefromtheinteractionsbetweenthebiomoleculesthatcomposethemandthe surrounding media (including surrounding cells). Basically nanotechnologies allow engineeringmaterials(suchasdrugdeliverysystems,diseaseimagingprobes,oreventissueengineeredconstructs)thathavedimensionsonthescaleofbiomolecules,whichinturnisthescalethatregulatesthefunctionsofcells.Nanotechnologieshavethepotentialtoimprovethewholecareprocessthatstartsforapatientonce a disease is suspected, from diagnosis, to therapy and followup monitoring. The aim is thedevelopmentofnewmaterialsandmethodstodetectandtreatdiseasesinatargeted,precise,effectiveand lasting way, with the ultimate goal of making medical practice safer, less intrusive and morepersonalised. The timescale from invention of a medical device or drug to release for clinical use isextremely long. In a few cases (such as drug delivery devices) nanotechnology is already in use for Page4of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 5. NANOYOUTeachersTrainingKitModule2Chapter1improvingpatientcare,butin mostoftheareasdiscussedbelowtheapplicationsarestillsomeyearsfrombeingproducts.DiagnosisDiagnosisofasuspecteddiseaseisoneofthemostcriticalstepsinhealthcareandmedicine.Wewantthediagnosistobefast,butalsoreliable,specific,accurate,andminimizetherisksoffalsepositives.Nanomedicinehasthepotentialtogreatlyimprovetheentirediagnosticprocess.Insteadofcollectingabloodsampleinavialandsendthistoaspecialisedlaboratoryfortesting(whichcantakedays)doctorswill be able to use miniaturized in vitro diagnostic devices in their office. These are small but highlyintegrated devices capable of carrying out quickly many tests at the same time using very smallquantitiesofsampletoperformtheanalysis.Someminiaturisedinvitrodiagnosticdevicesalreadyexistsuchasthebreathalyzersthatpolicecarriesforalcoholscreeningortheportableglucosetestdevicesused by diabetics. These devices can measure ions, small molecules, proteins, or can test for specificDNAsequencesthatarediagnosticforaparticulardiseaseormedicalcondition.Inthelastyearstherehas been a trend to make these devices even smaller, able to perform hundreds of tests at the sametime and be easier to use. Nanotechnologies have an important role in this development:nanomaterials, such as nanoparticles or nanotubes, can be integrated in the device. Scientist canengineernanomaterialstobeveryspecific,sotheirusewillmakethedevice evenmoreaccurateandable to carry our even more tests simultaneously. Nanomaterials have the characteristic of exhibitingsomepeculiarquantumeffectswhichcanbeusedforamplifyingthesignalarisingfromthedetection.Thereforetheuseofnanomaterialsinminiaturizedinvitrodiagnosticdeviceswillallowimprovingboththe specificity of the analysis, the throughoutput of the analysis (number of tests that can be donesimultaneously)anditsreadout.Inthefuture,thesetypeofdeviceswillallowtoperformpointofcarediagnostics,whichmeanstoperformadiagnostictestanywhereandnotonlyinthedoctorsofficeorinahospital.Thenatureofthesampletobetestedwillprobablychange,andbecomesalivaratherthanblood,whichismuchmoreconvenientandsafertohandle.Thiswillallowfortestinglargenumberofpatients, for instance in the event of an epidemic, or for testing large numbers of diseases or manyparametersforonespecificdisease,whichisneededforthediagnosisofcomplexmedicalconditions. Page5of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 6. NANOYOUTeachersTrainingKitModule2Chapter1Miniaturized diagnostic devices include biosensors, microarrays and labonachip (LOC) devices, alsocalled miniaturized total analysis systems (TAS). The first two are based on a parallel processingtechnique,whereasLOCdevicesarebasedonaserialprocessingtechnique.BiosensorsGenerallyspeaking,asensorisadevicecapableofrecognizingaspecificchemicalspeciesandsignallingthe presence, activity or concentration of that species in solution through some chemical change. Atransducer converts the chemical signal (such as a catalytic activity of a specific biomolecule) into aquantifiablesignal(suchasachangeincolourorintensity)withadefinedsensitivity.Whenthesensingisbasedonbiomolecularrecognitionitiscalledabiosensor.Therearevarioustypesofbiosensors,suchas antibody/antigen based, nucleic acid based and enzyme based. Also, depending on the techniqueusedinsignaltransduction,biosensorsareclassifiedasopticaldetectionbiosensors(asintheexampleabove),electrochemicalbiosensors,masssensitivebiosensorsandthermalbiosensors.Therearenumerousnanoparticlesthatcanbeusedasbiosensorscomponents.Theseworkasprobesrecognizing an analyte or differentiating between analytes of interest. In such applications somebiological molecular species are attached to the surface of the nanoparticles to recognize, through akeyandlockmechanism,thetargetofinterest.Theprobesthensignalthepresenceofthetargetbyachangeincolour,massorotherphysicalchange.Nanoparticlesthatareusedaselementsforbiosensorsinclude quantum dots, metallic nanoparticles, silica nanoparticles, magnetic beads, and fullerenes,whicharehollowcagesofcarbon,shapedlikesoccerballs.Other biosensors use nanostructured particles as nanosieves through which charged molecules aretransportedinanelectricfield.Inthiscaseparticleswithengineerednanoporesareused.Carbonnanotubesandnanowiresarealsoemployedforsensing.Thelattercanbefabricatedoutofasemiconductormaterialandtheirsizetunedtohaveaspecificconductingproperty.This,togetherwiththe ability to bind specific analyte on their surface, yields a direct, labelfree electrical readout. Thesenanowires biosensors allow detecting a wide range of chemical and biological species, including lowconcentrationofproteinandvirusesandtheirapplicationspansfromthemedicaltotheenvironmentalsector.Figure1&2illustratesasiliconnanowiresbiosensorbasedonbiorecognition. Page6of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 7. NANOYOUTeachersTrainingKitModule2Chapter1 Figure1.Biorecognitiononasiliconnanowire biosensor.Thesurfaceofthenanowireismodified withavidinmolecules(purplestars)whichcan selectivelybindastreptavidinfunctionalisedmolecule ornanoparticle.(Imagecredit:L.Filipponi,iNANO, AarhusUniversity.CreativeCommonsShareAlike3.0)Figure2.Thisscanningelectronmicrographdepictsthefunctionalpartofananobiosensor containing silicon nanowires. (Image credit: P Mohanty,Boston University, NISE Network, www.nisenet.org, licensed under NISEnetworktermsandconditions).Nanoscalebiosensorshavethepotentialtogreatly aidin thediagnosisofdiseasesand monitoring oftherapies. A large number of approaches have been developed in a recent years while relatively fewhavebeensofarconvertedintoclinicaldiagnostictools,thereforetheirwideapplicationinpatientcareisforeseeninthenext510years.CantileverbiosensorA cantilever biosensor is a biosensor made of numerous arms(calledcantilevers)whicharetensofmicrometreslongbutverythin(few micrometres). These devices are fabricated throughlithography and etching. The surface of the cantilever isfunctionalized with a nanometer thick layer of coating whichensures anchoring the probe material (which can be a DNA strandoraprotein,forexample).EachcantileverisdifferentandcanprobeFigure3.Schematicdiagramofacantileverbasedbiosensor.Theyellowmoleculesbindspecificallytotheredmolecules on the right hand cantilever and are detected by the bending of the cantilever. (Image credit: M.Lorenzen,iNANO,UniversityofAarhus.CopyrightM.Lorentzen. Page7of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 8. NANOYOUTeachersTrainingKitModule2Chapter1foradifferenttarget,asschematisedinFigure3.Inthistypeofsensor,theadsorptionoftheanalytetothespecifictargetsonacantilevercausesasurfacestressandbendsthecantilever.Themostcommonread out is optical, where the angular perturbation of a laser beam is measured upon bending of thecantilever. Although common, this method suffers from the limitation that measures are difficult inopaqueliquids,suchasblood,becauseoftheabsorptionofthelaserlight.Analternativetothismethodis the piezoresistive readout, where a piezoresistor is integrated in the cantilever. Upon detection oftheanalyte,thestressappliedtotheresisterchanges,whichisreflectedinachangeofitsresistance,which in turn is measured as an electrical signal. This approach offers the advantage of allowing thedetection in opaque media, the possibility of miniaturizing the sensor and incorporating it in portabledevicesforpointofusesensing.PlasmonicbiosensorsThe optical properties of noble metal nanoparticles have received significant research attention inrecent years for their potential as components in many applications, including chemical/biochemicalsensors.TheopticalpropertiesofnoblemetalnanoparticlesaredominatedbyaneffectcalledLocalizedSurfacePlasmonResonance(LSPR)whichwasdescribedinChapter4ofModule1Fundamentalnanoeffects.OneoftheconsequencesoftheLSPReffectinmetalnanoparticlesisthattheyhaveverystrongvisible absorption due to the resonant coherent oscillation of the plasmons. As a result, colloids ofmetalnanoparticlessuchasgoldorsilvercandisplaycolourswhicharenotfoundintheirbulkform,likered,purpleororange,dependingonthenanoparticlesshape,sizeandsurroundingmedia.TheenergyofLSPRsissensitivetothedielectricfunctionofthematerialandthesurroundingsandtotheshapeandsizeofthenanoparticle.Thismeansthatifaligand,suchaprotein,attachestothesurfaceofthemetalnanoparticle,itsLSPRenergychanges. Similarlythe LSPReffectissensitivetoothervariationssuchasthedistancebetweenthenanoparticles,whichcanbechangedbythepresenceofsurfactantsorions.The fact that the LSPR depends on the dielectric environment means that the refractive index can beused as the sensing parameter: changes in the local dielectric environment, induced by the sensingprocess,areusedfordetectingthebindingofmoleculesintheparticlenanoenvironment.Inaplasmonicbiosensorsthenanoparticlescanbedispersedinamedia(inwhichcasethebiosensorisacolloidalplasmonicbiosensor)orsupportedonasurface(surfaceplasmonicbiosensors).Bothtypesofsensors exploit the fact that the sensing event changes the LSPR of the metal nanoparticles, but usedifferentreadoutreportstrategies: Page8of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 9. NANOYOUTeachersTrainingKitModule2Chapter1 In a colloidal plasmonic biosensor (for instance made of goldnanoparticles) the sensing event results in a change of aggregationamong the nanoparticles that form the colloid (Figure 4), which candetermineacolourchangeofthecolloid.Absorptionspectroscopyisused to quantify the biosensing event. In the case of gold colloid,which is normally red, the sensing event can result in the colloid Figure4.Schematicrepresentationbecoming blue. Thus metal colloids can be used as plasmonic ofacolloidalplasmonicbiosensor.colorimetric biosensors. In nanomedicine this effect is used forinstanceforgeneticscreeningwherescientistlookforaspecificgenesequenceinasamplewhichcanbeindicativeforaspecificdisease.Howisthisdone:First,thesequenceofbasesinthetargetDNAisidentified.ThentwosetsofgoldparticlesarepreparedonehasDNAattachedthatbindstooneendofthe target DNA, and the second set carries DNA that binds to the other end. The nanoparticles aredispersedinwater.WhenthetargetDNAisadded,itbindsbothtypesofnanoparticletogether,linkingthem together to form an aggregate. The formation of this aggregate causes a shift in the lightscattering spectrum from the solution, that is, a colour change in the solution that can easily bedetected.TheexampleisillustratedinFigure5. The EXPERIMENT C in the Experiment Module deals with synthesizing and testingagoldcolloidalplasmonicnano sensor. In the experiment students use electrolytes to seethecolourchangeduetothechangeinaggregation ofthenanoparticlesasthesaltisaddedtothecolloid. Figure 5. . A plasmonic colloidal nanosensors. (Imagecredit:reprintedwithpermissionfromJinetal.,Journalof American Chemical Society (2003), 125 (6), 1643 .Copyright2003AmericanChemicalSociety). Page9of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 10. NANOYOUTeachersTrainingKitModule2Chapter1Inasurfaceplasmonicsensor,metalnanoparticlesareimmobilizedonasurfaceasillustratedinFigure6(a).Themetalnanoparticlesareattachedtothesurfacebymeansofchemicallinkersorpreparedbynanolithography(b),andarethanmodifiedwiththesensormoiety(c).Theanalyte(thetarget)attachesfromsolutionspecificallyontotherecognitionfunctionadsorbedontotheparticles(d),causingachangeintherefractiveindexaroundtheparticleresultinginanLSPRshift.TheLSPRshiftismeasuredthroughatechniquecalledextinctionspectroscopy(e).Figure 6. Schematic representation of the preparation and response of LSPR biosensors based on refractiveindex changes. (Image credit: Reprinted from: Borja Seplveda et al., LSPRbased Nanobiosensors, NanoToday(2009),4(3),244251,withpermissionfromElsevier).ArtificialnosebiosensorAn artificial nose biosensor is a device that mimics the ability of some mammals, like dogs, to detectexplosiveanddrugsthoughttheirolfactorysystem.Recentresearchhasshownthatthisfinecapacityindogsmayalsobeusedtodetectmoleculeswhich,ifpresent,areearlyindicatorsofdifferentdiseaseslike cancer. Numerous research programs are running globally to create an artificial nose, one is theEuropean project called BOND. The application of such type of biosensor ranges from medicine (earlydiseasedetection),security(explosivedetection)toalsothefoodindustry,tomeasureifthefoodhasgoneoff.Thistypeofbiosensorisanelectrochemicalsensorthatmimicsthenaturalmammalolfactorysystem. The nosebiosensor, in the same manner as our nose, is composed of 3 main parts, abiomolecule receptor, an electrode, and a transducer. When the detector finds the target, a chemicalreaction occurs between one or some of the biomolecules receptor for odour. The chemical reactionbetweenthereceptorandthesubstancewesmell,emitsachemicalsignal.TheelectrodetranslatesthisPage10of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 11. NANOYOUTeachersTrainingKitModule2Chapter1chemical signal into electrical signals, and transports them to the brain, or, in the case of the nosebiosensortheyaretransportedtoatransducer.Inmammalstheequivalentoftheelectrodeswouldbetheneurons.Thetransducer(thebraininmammals)receivestheelectricalsignalandtranslatesitintoanalyticalinformation.Inthenosebiosensorthetransducerwouldgiveinformationonascreen. NANOYOUVIRTUALGAME:Studentscanbecome nanoscientistsandfabricateandtestanosebiosensorthroughtheNANOYOUVirtualGame.Detailsarefoundatwww.nanoyou.eu/virtuallab.htmlNANOYOUDILEMMA:Diagnosticnanosensors(cantileverbased,plasmonic,nanowiresbasedetc)willallowfortheearlydetectionofvariousdiseases,likecancer,attheveryonsetofthesymptoms,beforethediseaseisperceivedbythepatient.Earlydetectionmeanshigherchanceofsuccessfullytreatandovercomethedisease.Ontheotherhandsomeworrythatthiswillgivedoctorsaccesstoalargeamountofpersonalinformation.Thequestionis:whereisthisinformationgoingtobestored,andwhowillhaveaccesstoit?Also,whatifthosedevicesareusednotasadiagnostictoolbutasameantoassessapersonsmedicalconditionbyotherentities,suchasinsurancecompanies,orjobagencies?Thedilemmais:Shouldnanosensorsbeusedtodiagnosemedicalconditionsintheearlystageswhentherearestillnodefiniterestrictionsinplacetoprotectpatientsprivacy?ThisdilemmaispartoftheNANOYOURolePlayCardGame(seewww.nanoyou.eu/en/decide)MicroarraysThese devices are used for diagnostics purposes such as DNA analysis (DNA microarray), proteindetection (protein microarrays) as well as whole cell analysis. Microarrays are platforms made ofhundreds of detection sites, that have micronsize dimension, and allow the specific detection of a(bio)chemicalwithinamixtureorthesimultaneousdetectionofmany(bio)chemicals.Thedetectionisrelated to the chemical functionality on the micronsized spots in the array and it leads to a singlechemical yes/no reaction per spot. Microarrays are used as screening tools, not only for diagnosticpurposes but also for screening new drugs. Nanotechnology can impact microarray technology bycreating densely packed, smaller, nanosized arrays (nanoarrays) that could allow faster screening oflarger number of (bio)chemicals. Problems associated with the handling of ultrasmall quantities ofliquid exist, though. Therefore nanotechnologies offer the most promising advantages in the sampledetectiononthearray.Theconventionalmethodusedfordetectingtheyes/noreactionateachspotis Page11of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 12. NANOYOUTeachersTrainingKitModule2Chapter1fluorescence.Thistechniqueusesfluorescentprobesmadeoforganicmoleculesattachedtothespeciesto be detected (e.g., a protein, or a fragment of DNA): when reaction occurs, this is attached to thedetection spot, which becomes fluorescent in a colour corresponding to the emission of thefluorescent probe. Fluorescent staining suffers from some disadvantages, mainly fast bleaching of thefluorescentmolecules(thatis,lossofbrightnessofthecolourintimeduringimaging);limitednumberofdyemoleculesthathavedistinctcoloursandthatcanbecanbesimultaneouslyimaged;andlimitedsensitivity. Nanoparticles in the form of quantum dots (QD) can be used as an alternative toconventionalorganicdyes,beingthesemorestable,sensitive,andmonochromatic.Asubstantial10foldenhancementinsensitivitycomparedtocommonfluorescentmarkershasbeenaccomplishedthroughthe use of gold and silver particles of uniform dimensions in the range of 40 nm to 120 nm. Signalamplification is also obtained using metal nanoparticles labels, such as DNAmodified goldnanoparticles. These nanosized probes have molecules attached to their surface that ensure theselectively of the detection, while the nanoproperties of the probe is responsible for enhancing thesignal. The overall effect is an improvement of the sensitivity and selectively of the microarraytechnology.NanobarcodesTheuniquepropertiesofnanoparticles,suchastherelationshipbetweenparticlessizeandcolour,canbealsoemployedtocreatemultiplexeddetectionsystemsintheformnanobarcodes,forinstanceusingQDtocreatedifferentcolourbasedcodes.Alternatively,fragmentsofDNAonnanospherescanbeusedto create a biobarcode, for instance for protein detection. A biobarcode was used to detect smalllevelsofthe cancermarkerprostatespecificantigen(PSA) inserum. Theresultsshowed anincreasedsensitivitytothePSAproteincomparedtoconventionalproteinassaysdemonstratingthepotentialofsuchapproachestodetectcancersatanearlierstage.LabOnAChipThese devices are miniaturized integrated laboratories that allow the separation and analysis ofbiological samples (e.g., blood) in a single device. They are made of microfluidic systems, includingmicropumps and microvalves, integrated with microelectronic components. The device can alsointegrateoneormoresensors.Aswithmicroarraytechnology,theimpactofnanotechnologiesinthisareaisfurtherminiaturizationofthesedevices,althoughthehandingofultrasmallvolumesofsampleswould pose a problem. Presently, nanotechnologies are making an impact in improving specific Page12of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 13. NANOYOUTeachersTrainingKitModule2Chapter1components and functions of labonachip devices. For instance, analysis is commonly done bydielectrophoresis,wherenonuniformalternatingelectricalfieldsareusedtoseparateandguidesmallobjects through field gradients. This manipulation requires high electrical field strengths that can beobtainedusingnanosizedelectrodes.Anotherexampleisnanoporebasedseparationsystemsthatcanbeintegratedinthemembranesusedinlabonachipdevices.Forinstance,nanoporemembranesareproposedforDNAsequencing.ImagingThesecondstepinthediagnosisofadiseaseinvolvesinvivoimaging,whichsearchesforthesymptomsofthediseasewithinthelivetissuesuspectedofbeinginfectedwithouttheneedtoperformsurgery.Nanotechnologiesarehavingaveryimportantimpactinthisarea,particularlybydevelopingmolecularimagingagents.Latestimprovementsintheareaofimagingdealwiththecapabilityoftrackingchangesatthecellularandmolecularlevelthroughtheanalysisofsomespecificbiologicalmarkers(atechniquecalled targeted molecular imaging or nanoimaging). A biomarker is an indicator of a biologicalprocessorstate,suchasadisease,ortheresponsetoatherapeuticintervention.Thiscanbeanalteredgene, or change in protein production, or even a physical feature of a cell. The aim is to detectbiomarkers of disease and diagnose illnesses before or at the onset of the first symptoms, this waymakinginvivoimagingatoolfortheearlydetectionofadisease.Effectiveearlydetectioniscrucialforplanning a therapy with less severe and costly therapeutic demands, especially in those diseases, likecancers,wheretimingisvitalforthesuccessofthetreatment.Biomarkerscouldbeusedalsoasearlyindicators of the success of a treatment, therefore reducing treatment time and cost. Targetedmolecularimagingisimportantnotonlyfordiagnosticpurposes,andformonitoringtheprogressofatherapy,butalsoforresearchincontrolleddrugrelease,inassessingthedistributionofadrugwithinthepatientsbody,andfortheearlydetectionofunexpectedandpotentiallytoxicdrugaccumulations.Theabilitytotracethedistributionofadrugleadstothepossibilityofactivatingitonlywhenandwhereneeded,thusreducingpotentialdrugtoxicity.DiagnosticimagingTechniques such as Xray, computer tomography (CT), ultrasound (US), magnetic resonance imaging(MRI)andnuclearmedicine(NM)arewellestablishedimagingtechniques,widelyusedbothinmedicineandbiochemicalresearch.Originally,imagingtechniquescouldonlydetectchangesintheappearanceof Page13of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 14. NANOYOUTeachersTrainingKitModule2Chapter1a tissue when the symptoms of the disease were relatively advanced. Later, targeting and contrastagents were introduced to mark the disease site at the tissue level, increasing imaging specificity andresolution. It is in this specific area that nanotechnologies are making their highest contribution bydevelopingbettercontrastagentsfornearlyallimagingtechniques.Thephysiochemicalcharacteristicsofthenanoparticles(particlesize,surfacecharge,surfacecoatingandstability)allowtheredirectionandtheconcentrationofthemarkeratthesiteofinterest.Anexampleofnanoparticlesusedinresearchforimagingareperfluorocarbonnanoparticlesemployedascontrastagentsfornuclearimaging,magneticresonanceimagingandultrasound,withapplicationtotheimagingofbloodclots,angiogenesis,cancermetastases and other pathogenic changes in blood vessels. Gadolium complexes have beenincorporatedintoemulsionnanoparticlesforthemolecularimagingofthrombusresultinginadramaticenhancementofthesignalcomparedtousualMRIcontrastagents.Fullerenesarealsousedinmagneticresonance imaging research, filled with small molecules that act like contrastenhancement agents.MetalsandsiliconnanoparticlesareusedalsotoenhanceMRI.Siliconparticlesfabricatedintodifferentshapesandcoatedwithconductivelayerscanhaveenhancedmagneticresonanceinteractionswithanimagingfield.Inxrayimagingtoenhancethesignalanagentmustdeliveradetectablenumberofheavyatomsintotargetedtissuewithouttoxiceffects.Nanoparticlesofheavymetalshavethehighestdensityofsurfaceatoms but they must be inert and stable. Nanoparticles of inert metals like silver and gold are tooexpensive and would make the technique not costeffective. A solution was proposed by GeneralElectricintheformofnanoparticlesmadeofheavymetalcompoundsencapsulatedingoldshells.Theaddedadvantageisthatorganiccompoundswithsulphide(SH)groups(thiols)canbeeasilyattachedto the gold surface through the thiol end (forming an SAu bond). The thiol molecule can befunctionalized at the other end with groups that act as receptors for specific binding of antigens,antibodies or even target compounds on the surface of the cell. By targeting receptors unique to acertaintypeofcancercell,goldnanoparticlescanenhanceanxrayimageofasuspectedcancertissuewithmanyordersofmagnitudes.Goldnanoshellsareapromisingmaterialfortheopticalimagingofcancer.Opticaltechnologiescouldprovide high resolution, noninvasive functional imaging of tissue at competitive costs. However,presently these technologies are limited by the inherently weak optical signals which come from theendogenous chromophores and the little spectral differences of normal and diseased tissue. Gold Page14of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 15. NANOYOUTeachersTrainingKitModule2Chapter1nanoshellsaremadeofadielectriccore(silica)coveredbyathinmetallic(gold)shield.Goldnanoshellspossess physical properties similar to gold colloid (described before), in particular, a strong opticalabsorptionduetothecollectiveelectronicresponseofthemetaltolight(theLSPReffect).Bychangingtherelative dimensionsofthecoreandshell,theopticalresonanceofthenanoparticlescanpreciselyand systematically varied over a broad region ranging from the nearUV to the midinfrared (Figure7).This range includes the nearinfrared (NIR) wavelength region where tissue transmissivity is higher.Researchersareusingthesegoldnanoshellscellsascontrastagentsforopticalcoherencetomography(OCT)1ofcancercells.Aswewilldiscusslater,goldnanoshellsarealsocapableoftreatingcancelcellsthroughoverheatingofthecells.Thiswillbediscussedinthenextsession.Figure 7. (Left): SEM image of gold nanoshells each beingabout120nm;(right):Opticalimageofthesame nanoshells after having being dispersed in water and dried on a microscope slide. The colours are due to selective scattering of light by the nanoparticles. (Images credit: G.Koeing, University of Wisconsin Madison, NISE Network, www.nisenet.org, licensedInsitudiagnosticdevicesIn recent years insitu diagnostic devices have been developed, such as wireless capsule endoscopycameras.Thesedevicescanbeswallowedbythepatientandallowtocloselymonitorandlocatethesiteof bleeding and other intestinal problems. Currently many of these devices, such as the CamPillproducedandsoldbyGivenImaging.Ltd.canonlyimagetheproblem.Inthefuturethesedevicescouldalso incorporate sensors for the detection of specific chemicals, pH, bacteria, viruses etc. Micro andnanotechnologies now allow the creation of extremely small sensors; therefore research is beingconductedforintegratingthoseintoswallowablecapsules.Thiswillwidentheirapplicabilityandutility.Inthefuture,thesecouldalsobedrugloadedfortargeteddrugdelivery.1 Optical coherent tomography (OCT) is a stateoftheart imaging technique which produces high resolution (typically 1015 m), realtime,crosssectionalimagesthroughbiologicaltissues.Themethodisoftendescribedasanopticalanaloguetoultrasound. Page15of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 16. NANOYOUTeachersTrainingKitModule2Chapter1TherapyThesamedisease,suchascancer,canexpressitselfinmanydifferentforms;forinstancethereareatleast14differenttypesofbreastcancer.Thus,inanidealworld,atherapyshouldbespecific,inordertoremoveonlythebadcells,andeffective,bothintermsofactionandtime.Atherapynormallyinvolvesapharmaceuticalroute(drugs)totreatthediseasefromtheinsideofthebody, or, when a pharmaceutical therapy is not possible or not effective, other routes to fight thediseasefromtheoutsideofthebody,suchasradiativetherapies.Incertaincircumstances,surgeryisrequired,andanorgansubstituteisinsertedinthebodyintheformofanimplantoradonatedorgan.Inallthoseapproaches,whichareoftenusedincombination,theaimisalwaysthesame:toeliminateselectivelythesourceofthediseaseinalonglastingway.Nanotechnologiesaremakingatremendousimpact in this field, with new drugs and new type of treatments under development, some of whichhavealreadyprovenclinicallyeffectiveandhaveenteredthemarket.DrugdevelopmentandtargeteddrugdeliveryAdvancement in the field of pharmacology deal with two main concepts: development of newbiologicallyactivedrugs(drugdiscovery)anddevelopmentofnewdrugdeliverysystemsabletoreachthespecificsiteofthedisease.Drugdeliverysystems(DDS)arenotanewconcept:researchinthisfieldstartedinthemid1960sandresultedinthetypeofdrugsweusetoday,thatis,medicineswheretheactiveingredientisencapsulatedandreleasedinsidethebodybygradualdissolution,osmoticeffects,orothermechanisms.DDSareintheformofpillsthatwefrequentlytakeandthatcanreleasegraduallytheiractivecomponentintime(slowreleasedrugs)ordissolvebasedonsomephysiologicalconditions(e.g.,acidityoftheenvironment).DDSalsoexistintheformofimplants,inserts,orotherdrugreleasingsystems.DrugdesignandscreeningThe structure of biological macromolecules defines a three dimensional nanoenvironment thatmediatesspecificfunctionsinthecell.Thedesignofnewdrugsrequiresaverydetailedunderstandingof this nanoenvironment. Therefore gaining insight into the structure of macromolecules on thenanoscale through electron microscopy, nuclear magnetic resonance spectroscopy (NMR) and xraycrystallography is of fundamental importance for understanding biological processes and for thedevelopmentofnewmedicines. Page16of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 17. NANOYOUTeachersTrainingKitModule2Chapter1Oneofthebottlenecksindrugdiscoveryisthenecessityofscreeningthousandsofcandidatedrugsfortheirefficacyinfightingtargetedmacromoleculesinadiseasestate.Microand,now,nanotechnologieshaveenabledthedevelopmentofmicroarrayplatformandnewdetectionmethods(includinglabelfree)to investigate the effects of candidate drugs against disease macromolecules with unprecedentedspeed.TargeteddrugdeliveryPharmaceuticaldrugsdevelopedusingconventionalsyntheticalroutesarelimitedbyproblemssuchaslow efficacy, low solubility in water and lack of selectivity. In addition, physiological barriers oftenpreventthedrugfromreachingandactingatthetargetsiteaphenomenoncalleddrugresistance.Thelow solubility and limited bioavailability of conventional drugs is responsible for their limitedeffectiveness:thebodyoftenclearsawaythedrugbeforeitsactionhasfullyoccurred.Theefficacyofdrugsisalsodependentonthedoseused,butdosedependentsideeffectsoftenlimittheiracceptableusage.Thelackofselectivityisespeciallydetrimentalforinstanceincancertherapies,sinceanticancerdrugs,usuallyusedinlargevolumeofdistribution,aretoxictobothnormalandcancercells.Arecognisedneedexiststoimprovedrugcomposition,delivery,releaseandaction,andthustodevelopnewdrugsthatcanactatthespecificsiteofthedisease,maximizingthedrugstherapeuticactionwhileminimizingsideeffects.Fordrugstobeabletodoso,thedeliverysystemsneedtobeminiaturizedinsize to become much smaller than the target, and specific in composition to elicit a certain response.With the use of nanotechnologies, targeted drugs (in terms of composition and delivery system) arebecomingareality.Inthefuturethiscouldleadtotargetedtherapiesandpersonalizedmedicine.Theaimistodesignanddeliverdrugsinsuchawaythattheycanrecognizethebad`cellsatamolecularlevel and penetrate the cell membrane and act inside the infected cell. This is often crucial for theefficacyofadrug,sinceitisacrossthecellmembraneandinsidethecellthatmostvirusreplicationandotherdiseaseconditionstakeplace.Thisway,thetreatmentwillbedeliveredwhereisneededandwillbe specific, eliminating the problem of the drug killing healthy cells. An example of this approach issiRNAdrugdelivery.Targeted drugs and targeted DDS could allow the creation of drug formulations with optimal loading,thereforedeliveringtothebodyonlythenecessaryamountofthedrugsandreducingsideeffectsforthepatients.TogetherwiththepossibilityofhavingnanoDDSthatarebiodegradableinsidethebody, Page17of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 18. NANOYOUTeachersTrainingKitModule2Chapter1this will help to reduce drug toxicity. Drug safety can be further enhanced by the possibility ofintroducinginsidethedrugformulationalabelthatchangescolourwhenthedrugreachesitsexpiringdateorisnolongerfunctioning.Thiswillallowtheimprovementofdrugshelflifeandbettermonitoringofdrugsafety.siRNAdrugdeliveryRNAinterferenceisanatural,fundamentalmechanismingeneregulationthatoccursbothinplantsandin animals, humans included. Genes carry the genetic material of an individual, the DNA, and arecontainedinthenucleusofacell.Whengenesareexpress(thatis,activated)thegeneticinformationiscopiedfromDNAtomessengermolecules,calledmessengerRNA(mRNA),whichthenorchestratetheformationofproteinsoutsidethenucleusofthecell.In1998AndrewFireandCraigMellodiscoveredthat double stranded RNA (dsRNA) can interfere and break down the mRNA for a specific gene, thusstoppingtheproductionofaspecificprotein.Thegeneinthereforesilenced`andtheproductionoftheproteinisturnedoff.FireandMellofoundthatthisRNAinterferencemechanismisspecificandcanbeobtainedwithfewmoleculesofdsRNA,andthattheeffectofdsRNAcouldspreadfromcelltocellandfromtissuetotissue,andevenpassedontooffspring.ThediscoveryawardedthescientiststheNobelPrize in medicine in 2006. Now researchers know that RNA interference plays an important role inswitching off genes during an organisms development and to control cellular functions. But thediscoveryofRNAinterferencenotonlyallowsscientiststobetterunderstandthefundamentalsofgeneregulation;italsoopensnewpossibilitiesforgeneticengineeringinbiologicalandmedicalresearch.Inthe laboratory scientists can now tailor RNA moleculessilencing RNAsthat activate the breakdown ofendogenous mRNAs (that is, RNA that belongs to that specific cell). When silencing RNA (siRNA)moleculesenterthecelltheyactivateRNAinterference,andendogenousmRNAmoleculesthatbindtotheaddedsiRNAaredestroyed.ResearchersarenowhopingtouseRNAinterferencetotreatdiseaseslikeviralinfections,cardiovasculardiseases,cancerandmetabolicdisorders.Sofar,manyexperimentswithRNAinterferencehaveyieldedpromisingresults,butinordertomaximizethetherapeuticefficacyofthetechniquesomefundamentaldifficultieshavefirsttobeovercome.TheseincludelowstabilityofsiRNA in biological fluids and low specificity of action due to gene offtarget effects caused by thesimilarityinbehaviourofsyntheticsiRNAwithnaturalmicroRNAproducedbythecell.Therefore,thereis the need to develop delivery methods capable of overcoming the extracellular and intracellular Page18of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 19. NANOYOUTeachersTrainingKitModule2Chapter1barriersandgetthesiRNAmoleculesintotherighttypeofcell(targeteddelivery)whilstmaintainingthestabilityofsiRNA.ResearchersatiNANO(AarhusUniversity)andotherinstitutionworldwidearedevelopingnanocarriersfor the targeted delivery of siRNA. For example, they are studying a novel chitosanbased siRNAnanoparticledeliverysystemforRNAinterferenceinvitroandinvivo.Chitosanisanaturallyoccurringcationic polysaccharide that has been widely used in drug delivery systems. It contains positivelycharged amine groups that can interact with the negatively charged backbone of siRNA and formpolyplexes in the form of nanoparticles about 200 nm in size. The protonated amine groups allowtransport across cellular membranes and subsequent endocytosis into cells. It was shown thatchitosan/siRNA nanoparticle delivery system silences genes both in vivo and in vitro. Moreover, thisdelivery system has been shown to be biocompatible, nontoxic and biodegradable. Anotherrequirement in targeted siRNA delivery is its capability of the carrier to reach a specific cellularcompartmentandreleasethecargo(thesiRNA)insidethatcell.Syntheticvectorsbasedonpolycationssuch as polyLlysine have been widely used buthave several drawbacks such as high cellulartoxicity,sequestrationinsubcellularcompartmentsand lack of intracellular targeting. In contrast,bioresponsivecopolypeptidescontainingreducible disulfide bridges that respond to intracellularFigure 8. Live cellular uptake of chitosan/siRNA acidity conditions have proven advantageous innanoparticles into NIH 3T3 cells. Fluorescencemicroscopy was used to visualize cellular uptake and delivering nucleic acids into cells. These systemstranslocation of Cy5labeled siRNA within Chitosan exploit the redox potential gradient existingnanoparticles after 4 hours of reaction. Images show between the extracellular and the intracellularfluorescentoverlayofsiRNA(redCy5labeled)andnuclei environment so that the disulfide bridges are(blueHoechstlabeled)adjacenttophasecontrastimage broken (and the cargo released) only in one (scale bar, 10 m). (Image credit: reprinted by permission from Macmillan Publishers: K.A. Howard et compartmentofthecell(inthiscase,thenucleus). al.,MolecularTherapy(2006),14(4),476484.Copyright For these reasons research is being conducted in 2006.) developingnanocarriersrichinhistidinegroups. Page19of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 20. NANOYOUTeachersTrainingKitModule2Chapter1StimuliactivateddrugdeliveryInthisareaofresearchtheideaistoincorporateintothedeliverysystemsomespecificpropertiessothatthedrugcanbeactivatedonlyuponreachingthetarget,andtheactivecomponentreleasedatacontrolled rate. This is called stimuliactivated drug delivery. Controlled activation could be linked tosome environment properties, such as pH, or lockandkey` molecular recognition mechanisms. Oneexampleisstimuliactivatedgenedelivery.In gene therapy one of the biggest challenges is the targeted delivery of the nuclei acid load to thetarget (e.g., plasmidDNAorsiRNA)either tosilence(RNAsilencing)ortoactivatetheexpressionofaprotein as a way to treat a number of diseases. In the section before we discussed how nanocarrierdelivery systems formed by electrostatic interactions between cation polymers and DNA or RNA havebeen developed to overcome extracellular and intracellular barriers to maximize the delivery of thenucleicacidsinthecell.Onewaytocontrolthespatialandtemporalactivityofnucleicacidsistousepolymers that change properties in response to stimuli such as temperature and redox potentialgradients.ThisapproachtotargeteddeliveryisinvestigatedatiNANOandisschematicallyillustratedinFigure 9. The idea is to utilize ananocarrierthatpassivelyaccumulatesinthe diseased tissues (e.g., tumours),followedbystimuliinducedactivationatthe required site (inside or outside thecell). In the case of thermoresponsivesystems, the application of heat inpreciselocationsofthetissuecaninducethe deposition of the nanocarrier in theextracellulartargetregion.Researchersarestudyingforinstancetheuse of a thermoresponsive polymer toFigure9.Aschematicrepresentationofananocarrierthermoform a polyplex with plasmid DNA and activatedforgenetherapy.(Imagecredit:Howardetal.,Smalluse AFM to visualize the resulting(2007), Volume 3, Issue 1, pp. 5457. Copyright WileyVCHnanoparticles. They found that theyGmbH&Co.KGaA.Reproducedwithpermission.could change the size of the polyplex Page20of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 21. NANOYOUTeachersTrainingKitModule2Chapter1nanoparticles from around 50nm to more then 200nm by heating the particles. The AFM imagesrevealedthatsmallerparticlesmergedintolargerparticlesduringthetemperaturetreatment.Sincetheabilityofpolyplexestocrossvasculatureendothelialbarriersandenter/exittissuesdependsonparticlesize, a thermostimulus could be used to control the migration inside tissues. The idea is to apply athermostimuliinthediseasedtissuesotoinduceasizeincreaseofthenanoparticlesandpreventthesefromreentryintothebloodstream(seeFigure8).Thisgeneralapproachcouldbeusedfornanocarrierscontainingdrugsorimagingagentsfortherapeuticanddiagnosticapplications.CurrentandfuturenanodrugcarriersNanosized drug carriers that are currently under development include either materials that selfassemble,orconjugatedmulticomponentsystems,forinstanceadruglinkedtoaproteinandapolymer(called polymerdrug conjugate). Numerous nanosystems are now investigated, and include micelles,nanoemulsions, nanotubes, nanofibres, liposomes, dendrimers, polymer therapeutics, nanoparticles,nanocapsules,nanospheresandhydrogels.Someofthesenanosizeddrugcarriersareestablishedinthefieldofdrugdelivery,suchasliposomes,othershavemadetheirwaytothemarketinrecentyears,suchaspolymerproteinconjugates(polymerpharmaceutics).Manyarenowusedfortreatingsomeformsofcancer, hepatitis, and leukaemia. An example is an anticancer drug called DOXIL (from SequusPharmaceuticals).Figure10.(left)Schematicrepresentationof a polymerdrug conjugate, and of atargeted polymerdrug conjugate (right).(Image credit: Reprinted by permissionfrom Macmillan Publishers: Duncan R.,Nature Reviews Cancer (2006), 6(9), pp.688701.Copyright2006.The future of nanoDDS enabled by nanotechnologies could be miniaturised implantable chips loadedwithdifferentdrugsthatcanbereleaseduponexternalstimuli.Thiscouldfreepatients,likediabetics,fromhavingtoadministerdrugsrepeatedlyduringtheday.Researchinthisareaisveryactivebutstillrequireyearsforitscommercialrealization. Page21of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 22. NANOYOUTeachersTrainingKitModule2Chapter1ExternallyactivatedtherapiesthatusenanoparticlesOneofthedistinguishingpropertiesofnanosizeddrugcarriersistheirabilitytoaccumulatepassivelyincanceroussolidtumourtissueduetoaneffectcalledenhancedpermeabilityandretention`(EPR).Thispassivemechanismhasbeenattributedtotheleaky`natureoftumourvessels.ThebloodvesselsthatsupplytumourswithnutrientshavetinygapsinthemthatallownanoDDS(60400nminsize)togetinthetumourregionandaccumulatein.Thisfurtherenhancesthetargetedapproachtotreatinginfectedcells. Moreover, this allows the accumulation of therapeutic agents inside the tumour region andactivatingitusinganexternalsource.Basedonthisconceptsomenewanticancertherapieshavebeendeveloped and have entered advanced clinical trial stages. In these therapies, nanoparticles aredelivered to the tumour site where they accumulate. An external source is then used to specificallyactivatethenanoparticlesandoverheatthetumourregion(thetherapyiscalledhyperthermia).ThankstotheEPReffect,thenanoparticlesaccumulateonlyinthetumourregionsothetreatmentisextremelylocalisedandhealthytissuesarenotaffected.Overheatingthetumoursitecanbedoneforinstancebyactivating magnetic nanoparticles with an alternating magnetic field, where they start vibrating andgenerate heat. This is the principle based on which a new anticancer therapy has been developed,calledMagForce,whichhasenteredPhaseIIclinicaltrialin2007forthetreatmentofprostatecancer.Anotherapproachusesgoldnanoshells(describedbefore)designedtoabsorblightinthenearinfrared(NIR) region. This is the region where light penetration through tissue is optimal (8001300 nm). ThenanoshellsabsorbNIRlight,deliveredwithalaser,convertinglightintoheat.Inanimalmodelstudies,the nanoshell treatment has shown to induce complete resorption of a tumour in 10 days and allanimals remained healthy and tumourfree for more then 3 months after treatment. These examplesshowtheinnovativeapproachtotumourtreatmentenabledbynanoparticles.TheranosticOneofthemostexcitingopportunitiesthatnanotechnologies havebroughttothetherapeuticfieldisthe possibility of integrating the diagnosis, therapy and followup of a disease. This is referred to astheranostic, and could be enabled by nanoparticles incorporated inside a drug that can change somepropertysuchascolouroncethedrughasreachedthetarget(forinstance,quantumdots).Drugscouldtherefore have a feedback action. Together with a slow, targeted release system, the nanoparticlescouldgraduallychangecolourduringthedrugaction,thereforeinformingdoctorsoftheprogressofatherapy.Thisapproachiscalledfind,fightandfollow`andcouldbecomearealitythankstotheparallel Page22of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 23. NANOYOUTeachersTrainingKitModule2Chapter1progressofthefieldofmolecularimaging.Onevisionisthat,oneday,theentireprocessofdiagnosis,preimaging, therapy and postimaging of a specific disease will be integrated. An example oftheranosticistheuseofgoldnanoshellsforimagingandtreatingcancercellsatthesametime.RegenerativemedicineAttimes,theonlywaytotreatadiseaseistheremovaloftheinfectedorganortissue.Thelosscanalsoderivefromaninjuryoracongenitalcondition(e.g.,visionorhearingimpairment).Tocompensateforthe lost or impaired body function, an artificial construct is implanted in the body. Depending on thetype,siteandextentoftheloss,thisconstructcanbeintheformofanengineeredtissueoranimplant.TissueandbiomaterialengineeringTissueengineeringdealswiththefabricationofartificialscaffoldstosupportthegrowthofdonorcells,which differentiate and grow into a tissue that mimics the lost, natural one. This tissueengineeredconstructisthenimplantedinthepatientand,intime,resorbedbythebodyandfullyintegratedbythehost tissue. Current applications of tissue engineered constructs include engineering of the skin,cartilage and bone for autologous implantation (i.e., implantation of a tissue regenerated by seedingcellsofthepatient).Thescaffoldthatsupportscellgrowthisthecoreofthistechnique.Inthebody,cellsaresupportedintheir growth and function by a natural scaffold, called the extra cellular matrix (ECM). This is a verycomplexandintricatewebofnanofibresthatprovidethemechanicalarchitectureforcellulargrowth.Moreover,theECMisfilledwithsmallmolecules(e.g.,growthfactors)andproteinsthatdirectmanycellprocesses, such as adhesion, migration, growth, differentiation, secretion and gene expression. Thethreedimensionalspatialorganisationofthesecuesiscriticalforcontrollingtheentirecelllifecycle.Ultimately,thiscomplexnano,threedimensionalarchitectureguidescellstoformtissuesascomplexasbone,liver,kidneyandheart.Thebiggestchallengeinregenerativemedicineistheartificialreplicationofthisperfectnanoscaffold.Theabilitytoengineermaterialstohaveasimilarlevelofcomplexityisnowbecomingarealitythankstonanotechnology.Microfabricationtechniquesderivedfromthesemiconductorindustry(suchasphotolithographyorionbeam lithography) have long been used for the fabrication of microstructures to support and control Page23of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 24. NANOYOUTeachersTrainingKitModule2Chapter1cellulargrowth.Forinstance,oneofthepioneeringworksinthisfieldwaspublishedinthelate1970s8.In recent years new nanotechnology techniques have enabled studies at higher and higher resolutionrevealingthenanoscaledetailoftheECM.AnalyticaltoolsliketheAFMandnanofabricationtoolshaveallowed scientists to fabricate scaffolds with nanoscale features. A great deal of research is nowdedicated to engineering scaffolds with tailored material composition and properties, includingnanotopographyandcontrolledalignment,tostudyhowthesecansupportanddirectcellulargrowth.TheaimisthefabricationofscaffoldsthatmostcloselyresemblenaturalECM.Researchershavenowaccesstotechniquestoproducemacroscalestructureswithnanometredetails.Conventionalpolymerchemistrycombinedwithnewnanofabricationmethodsarenowusedtomanufactureawiderangeofstructures, such as nanofibres of different and well defined diameters and surface properties;nanofibrousandporousscaffolds;nanowires,nanotubes,nanospheresandnanocomposites.Close to the field of tissue engineering, and in many cases an integral part of it, is biomaterialengineering.Materialsusedinregenerativemedicinearecalledbiomaterialsinthesenseofbeingabletotriggerandsupportabiologicalresponse.Oneofthedistinguishingfeaturesofnanotechnologiesisitsabilitytocreatenew,functionalmaterials.Thiscanbeexploitedinthefabricationofnewbiomaterialsthathavebettermechanicalpropertiestoincreasetheimplantstabilityandreducefatiguefailure,forinstancefor orthopaedicimplants,andmaterials thathaveenhancedelectricalproperties,neededforinstance in neural prostheses. Nanotechnologies can also be used for fabricating implants made ofmaterialsthataremoreresorbable,toincreasefunctionalityanddurability.Forinstance,nanocoatingsarestudiedtobetterintegratesyntheticimplantswiththebiologicaltissue,inordertopreventtissueinflammationandtheonsetofrejection.Nanotechnologies are also employed for the fabrication of biomaterials that are responsive to theenvironment (for instance, responsive to the pH or to the presence of specific biomolecules), for thisreasoncalledsmartbiomaterials`.Moreover,researchisconductedtoincludenanoscalepatternsinthebiomaterial,tosimulatethenaturalcuesandmimicmolecularsignallingmechanisms,inordertotriggerdesired biological events, like cell adhesion, differentiation and spreading. This could enable thefabricationofdynamicimplantsthatarenotlimitedtoreplacingalostorganbuttrulyrestoringtheloss.Finally,nanosizedsensorscouldbeinsertedinsidethebiomaterial(forinstance,nanowirebiosensors)functionalisedwithreceptorsthatcanmonitorthepresenceofsmallorganicmolecules,proteins,cells Page24of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 25. NANOYOUTeachersTrainingKitModule2Chapter1(e.g., cancer cells) and viruses. This could be used to collect information on the implant status andactivity.Thisfeedbackinformationcouldbeusedtomaximizetheimplantefficacyandsafety.Tissue and biomaterial engineering have application in basically all aspects of regenerative medicine,i.e., neuroprosthetics and neuron regeneration (e.g., spinal cord repair), bone restoration, audit andvisionrestoration,motorrestorationetc.NanoengineeringboneregenerationBonesandteetharematerialthathavetobearcomplexloadsofmovingbodies,provideaprotectivecage for vital organs, anchor tendons and muscles, and act as joints and levers. This functionalcomplexity is reflected in a structural complexity. Bones have a complex hierarchical structure at thenano,micronandmacrolevel,whichdeterminesitsamazingproperties.The classic way to promote the regrowth of bone after injury is toprovide a scaffold into which bone forming cells can migrate and growattached (fused) to the scaffold. In the past numerous scaffolds havebeen used, mainly coated with hydroxylapatite which is a natural bonecomponent.Morerecently,researchhasfocusedontryingtomimicthenanostructureofthescaffold.Thisinvolves Figure11.Thisscanningelectronmicroscopeimageshowsaworking on the topography of the scaffoldhydrogelscaffoldgrownforstudyingbraintissue(surface nanoroughness) but also atengineeringandnerveregeneration.Theimageis100manchoring specific biomolecules to the wide(Imagecredit:DNisbet,MonashUniversity,NISEnanosurfaceofthescaffold.Theideaisto network,www.nisenet.org,licensedunderNISEnetworkmimic the natural fine organization oftermsandconditions)natural bone which is a combination ofnanotopographyand(bio)chemistry.Anovelapproachapproachisthedevelopmentofartificialbone,whichmeansusingmacromoleculesthat selfassemble into large structures that mimic the natural structure of bone. This is a bottomupapproachtoboneengineeringwhichleadstomaterialswithnanoscalelevelcontrol.Forinstancesomeresearchers have developed a bone scaffold by biomimetic synthesis of nanohydrohylapatite andcollagen.Collagenisthemostplentifulproteininthehumanbody.Itisfoundinmosthumantissues Page25of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 26. NANOYOUTeachersTrainingKitModule2Chapter1including bone, cartilage, the heart, eye skin and gives these tissues their structural strength. Thesebiomaterialsassembleinto3Dmineralizedfibrilsthatmimickeyfeaturesofhumanbone.Thismaterialshowssomesimilaritieswithnaturalboneintermsofhierarchicalmicroandnanostructure,andthreedimensionalporosity.Cellsgrowninvitrooverthisscaffoldgrewandproliferatedwell.Theadvantageofthis approach is that the building blocks are biomimetic macromolecules: once assembled, the finalmacromaterialcanintegratewithnaturaltissues,openingthewaytonewclinicalapproachestoboneregeneration.When these synthetic nanofibresformthey makea gelwhich couldbeused asasortofglueinbonefractures or to create a scaffold for other tissues to regenerate on to. As a result of its chemicalstructure the nanofibre gel would encourage attachment of natural bone cells which would help topatch up the fracture. The gel could also be used to improve implants or hip and other jointreplacements.NANOYOUDILEMMA:Theexampleofartificialboneisone of theNANOYOUdilemmapartoftheNANOYOURole Play Card Game (see www.nanoyou.eu/en/decide).The dilemma arises from the fact that if suchtechnology is being developed to heal bones, the natural progression could be that it might be used tostrengthenotherwisehealthybonestomakethemalmostunbreakable.Thissolutionmightnotbeavailableforsomepeople(duetocostoraccessibilityrestriction),soitcouldbecomeahumanenhancementtoolonlyforfew.Thedilemmais:Isitacceptabletouseprocessesdevelopedformedicaltreatmenttoenhancethehumanbody?NanoengineeringneuronregenerationThelossofneuronfunctionsisoneofthemostdramaticmedicalconditionsintermsofconsequencesforthepatient:itcaninterferewithbasicfunctions(likemovement)andcognitivecapabilities.Therearenumerous neurodegenerative diseases (Parkinsons, Alzeimer etc.) that are connected with aneurologicaldamage(neurodegenerativediseases).Neuronfunctionlosscanalsoderivefromasevere(spinalcordinjury)orminoraccident(peripheralneurondamage).Thelossorimpairmentofapersonsneurological function can have detrimental effects his/her life. The research in this field therefore isPage26of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 27. NANOYOUTeachersTrainingKitModule2Chapter1massive, and covers a very wide range of disciplines and subdisciplines. There are basically to mainapproachestoneuronregeneration:tissueengineeringandneuronprosthetics.Untilrecentlythesetwoapproacheswerefairlyseparated,mainlyduetothetypeofmaterialsemployedinthetwoapproaches:softbiomaterialsinthefirstcase(biopolymers,proteins,peptidesetc.),andmicrochiptypematerialsinthesecondcase(semiconductors,metalsetc).Withtheadventofnanotechnologiesthesetwotypesofmaterialsarestartingtobeintegratednotjustintermofphysicalattachment(e.g.,proteincoatings)butintermoffunction:forinstanceinnanotechnologiesbiomoleculesareusedasnanoscalemotors,orasenergyharvestingmaterials.Thereforeinthefuturetheapproachtoneuronregenerationwillbeinthe form of hybrid nanoscale devices. Below is a short review on neuron tissue engineering;neuroprostheticsarediscussedinthenextsession.NeurontissueengineeringTheuseofscaffoldstoencourageneuronregrowthafterinjuryisanestablished method. At first simple biocompatible polymers wereused; nowadays it is recognized the need to engineer the scaffold atthe nanoscale level in two ways: physical, by inserting nanoscalepaths to encourage directional growth and biochemical, by addingcues in the form of neuron growth factors and other essentialbiomolecules to encourage regrowth. These two elements must beengineeredsothattheircoordinatedactionresultsinneuronregrowth.Inthelastfewyearstheresearchinthisareahasbeen impressive and nanotechnologies have been theenabling tool. For instance researchers from the StuppLaboratory (Northwester University, Figure12.AnengineerednanomaterialthatsupportsspecificUSA) have fabricated a nanogel ofcellulargrowthandcanpromotedesiredneurobiologicalelongated micelles arranged in aeffects.Thematerialisformedofsmallbioactivemoleculesnanofibrematrixanddemonstratedthatthatresemblepartsofnaturalproteinswhichspontaneouslythis can support the directional growthassembleinnanofibres(A)makingamacrogel(B).Thegelof neurons. The aim of this and othersupportsandsteersthegrowthofstemcells(C).(Imagecredit:worksistoengineernanoscaffoldsthatReprintedbypermissionfromMacmillanPublishersLtd:G.A. Silva,NatureReviews(2006),7(1),6574.Copyright2006. Page27of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 28. NANOYOUTeachersTrainingKitModule2Chapter1can support the regrowth of neurons for healing patients affected of neurodegenerative disease orsevereneuronallosses,likeinthecaseofspinalcordinjury.NeuroprostheticsAneuronprosthesisisadeviceimplantedtorestorealostoralteredneuronfunction.Therearetwomainkindsofneuronprosthesis:motorandsensory.Muchprogressinthelastdecadehasbeenenabledbyminiaturization.Nanotechnologiesofferopportunitiestocontinuethisminiaturizationtrendbutalsoto introduce new features, like electrodes that actively interface with the nerves, smaller and morepowerful sensors, actuators, and control systems throughout the prosthesis to render it more naturalandeffective.A motor neuroprosthetic device takes the signals from the brain or motor pathway and covert thatinformationintocontrolofanactuatordevicetoexecutethepatientintentions.Examplesareartificiallimbsorhands.Thetaskisextremelycomplicated,sincethemotorneuroprostheticmustbeintegratedalsowiththeownersnervoussystem.Thereforeprosthesishaveasophisticateddistributednetworkofcontrol, actuation and feedback. Limb and hand prosthesis also need to be mechanically similar tonaturallimbs,otherwisethetaskoflearningtousethemwillbedifficult.Thereforetobeeffectivethedevicemustbedesignedwithnanoactuatorsandnanosensorsfullyintegratedwiththecontrolsystemfrom design to implementation, including the very special ergonomic interface between patient anddevice.What can nanotechnologies do? Progress in current motor neuroprosthetic devices has already beenmadepossiblebynanotechnologies,whichareenablingmorenaturalprosthesisbyproviding: Smaller and more affordable sensors, processor elements, and the wiring and interconnectorsneededtonetworktheminadistributedcontrolsystem; Smaller,morepowerful,efficientandresponsiveactuatorsthatmovesmoothly,resemblingnaturalmovement.Thisispossiblebecausetheseactuatorsarebasedonmolecularforces. Engineering materials that match the strength/weight ratios, elasticity/rigidity, and mechanicalenergystoragecharacteristicsofkeycomponentsofnaturallimbs. Page28of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 29. NANOYOUTeachersTrainingKitModule2Chapter1A key contribution of nanoscience is in the form of novel materials (e.g., carbon nanotubes) that canbecome elements of new sensors, computing elements and even artificial muscles. Nanoscalemagnetometers,accelerometers,pressuresensorsandgyroscopicdeviceswillbeabletomorepreciselydetect even minute movements and angle changes. These nanomaterials will support the design ofinternal movement devices to render the prosthetic movement more natural, and to ensure theaccuratetransmittanceofcontrolandfeedbackinformationbetweendeviceandpatient.Theimpactofnanotechnologies will not be in the form of miniaturized robots but rather assemblies of cooperatinginterconnected networks of computing, communicating, sensing, actuating, and so on, to make up aprosthesisthatresemblesasmuchaspossiblethenaturallostlimb.Motorneuroprostheticscommonlyintegratesensoryneuroprostheticsaswell,sincethereconstructionof motor functions needs to be associated with the reconstruction of hapticsthe sense of touch (ormorespecificallythesenseofpressureandforcefeedbackfromthebodytothebrain).Haptics,alongwithvision,hearing,andbalanceisanessentialcomponentofthestreamoffeedbackinformationthatthe nervous system sends to the brain. Socalled smart materials (or dynamic materials, meaningmaterials that change their shape or function due to an external stimulation) and nanosensors areexpectedtoprovidemanyoptionstoimplementsensoryneuroprosthetics.Research in the area of motor and sensory prosthetics is very active and is not only directed to thefabrication of human motor prosthesis but also robotics for computer assistedsurgery, deepmarineinvestigation,astronomyetc.Inadditiontoneuroprostheticsthatrestoremotionfunctions,visionandauditoryneuroprostheticsarealsoextremelyimportant.Bothtechnologieshavemadeenormousprogressinthelastdecadeandmuchhasbeenenabledbymicroandnanotechnologies.Onesuchexampleiscochlearimplants,whichareatype of electronic neuroprosthesis implanted in the middle ear, where they stimulate the ossicleselectromechanically, rather than acoustically, through either electromagnetic or piezoelectrictransducers. Nanotechnologies have enhanced microelectronics, batteries and micromechanicaltransducersincochlearimplantdevices.Problemsinthosedevicesstillexist,inparticularrefinementofthe signal processing (for instance to recognize voice pitch in music), improvement in their longtermbiocompatibility (build up of biofilms and plaques) and prevention of bacterial fungal infections.Nanomaterialsareexpectedtohaveanimportantimpactinalltheseareas. Page29of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 30. NANOYOUTeachersTrainingKitModule2Chapter1Neuronalstimulation,monitoringandpainmanagementThecardiacpacemakerisoneofthebestknownandmostwidelyusedneuroprosthetic.Othertypesofelectronic stimulators are cardiac defibrillators, cochlear implants, bonegrowth stimulators, neuralstimulators for the deep brain for controlling tremors in Parkinsons disease, neuronal stimulators forspinalcordrestoration,sacralandothernervestimulators.Thewaynanotechnologiesisimpactingthoseelectronic stimulators is through improved battery technologies, biocompatible materials and surfacetreatments for enclosures and leads, electrode miniaturization and efficiency improvements, andsmallersizedintegratedcircuitsforcontrolandpowerwithincreasedspeedandprocessingcapabilities.Electrical stimulation of neural tissue by surgically implanted neuroelectronic devices is already anestablished modern therapy. Integrated microand nanoscale devices allow applying many moreelectrodestothetargetsitewithfineresolutionandinacoordinated,dynamicway.Recordingofneuralactivity is also implemented by using nanoscale electrodes, since a much larger number of recordingsitesispossible.Thoseimplantedprobesmustberesistanttochallengingenvironments,thusthenanoscale surface engineering of these probes is essential. In addition to protection of the probesnanotechnologies is contributing in developing improved active interfaces between neurons andelectrostimulationdevices.Anexampleofanelectronicstimulatorisadeepbrainstimulator.Thisdevicewillallowtreatingpatientswith severe Parkinsons disease a disorder of the central nervous system that often impairs thesufferersmotorskills,speech,andotherfunctions.Ithasbeenfoundthattheuncontrollabletremorofpatientscanberemovedifafinenanosizeelectrodeisinsertedintothebrainthatdeliversacontinuouselectricalstimulussentthroughthebrain.Thispreciseelectricalstimulushastheeffectofremovingthetremors.Reductionofthesizeandpowerrequirementswiththeintegrationofmicroelectronicdevicesmakesitfeasible in many cases to energize an implanted device by RF electromagnetic transmission of power,this way eliminating wires and batteries. This is already the case for implanted pacemakers.Improvementsinenergystoragethoughtnanoengineeredenergymaterials,suchassupercapacitorsandconductivepolymers,coupledwithlowpowerrequirementsfornanoengineeredelectronics,willallowgreatimprovementsintermsofsizesandcapabilitiesofsuchdevices.Thesedevicesmakeitpossibletoperformelectricalstimulationinselectedpointsofnervous,sensoryandneuromuscularsystems.Such Page30of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 31. NANOYOUTeachersTrainingKitModule2Chapter1improvementsmightallowonedayusingimplantedelectricalstimulationforboneandtissuegrafts,andtostimulatefunctionintheendocrinesystemandotherorgans.NANOYOU DILEMMA: The example of deep brain stimulation is one of the NANOYOU dilemma part of theNANOYOU Role Play Card Game (see www.nanoyou.eu/en/decide).The dilemma arises from considering adeepbrainstimulatordesignedtotreatParkinsondisease.Howeverthisdevicecouldbeusedtotreatothermedicalandhealthconditionssuchasintractableepilepsy,aswellasmoodandeatingdisorders.Couldthisnanotechnology be used to increase the capability of the brain in areas for which it was not originallydeveloped?Anexampleofthismightbestudentsenhancingtheirconcentrationduringexams.Thedilemmais:Isitethicallyacceptabletousetechnologiesdevelopedforspecificmedicaltreatmentsforothersscopeslikeimprovinghumancapabilities?NoninvasivebrainmachineinterfacesThecontrolofphysicalobjectsbythepowerofthoughtalonehasalwayscapturedtheimaginationofhumans.Untilrecentlythispossibilitywasonlyconsideredpossibleinsciencefictionbooks.Nowwiththe aid of new technologies, and thanks to centuries of studies on neuron activity in the brain, thecontrol of machines and computers by the brain is becoming a reality. Systems are being developedwhere the patterns of neuronal firing in the brain are translated into electronic controls to supportcommunication,mobilityandindependenceofparalyzedpeople.Thisispossiblebecausethefiringsofneuronsandthetravelofioncurrentsalongaxonmembranesgeneratemagneticfields.Thismagneticfieldisproportionaltothevelocityofthechangeandthechangeinmagneticfieldisproportionaltotheacceleration or deceleration of the change. Therefore magnetism can become a noninvasivecommunicationtoolwithnerveswithoutimplantedelectrodesandpainfultranscutaneousshocks.The technique is called magnetic monitoring and it requires extremely sensitive magnetometersbecausethemagneticfieldsproducedbybrainactivityareverysmall.Todaymagnetoencephalography(MEG)canmapbrainactivityonaonemillimetregridorless.ThefirstgenerationofMEGequipmentwasverybulky,requiringshieldedrooms,highpowerconsumption,cryogeniccoolingofdetectorsandsignificant processing time. The technique has so far been limited to research labs or extremelyspecialisedmedicalinvestigations.Nanofabricationisenablingreductioninsizeofmostcomponentsof Page31of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 32. NANOYOUTeachersTrainingKitModule2Chapter1MEG equipments (sensors, magnets etc.) and new concepts are being developed; in prototypes thisprogress has already lead to 1000fold improvements in sensitivity and reductions in size and powerrequirementsbyfactorsof10to100.Magnetism can also be used to induce electrical currents in the neuron cell membrane like thoseinduced by implanted electrodes, but without physical contact. Magnetic stimulation is indeed a newmedicaltechniquewhichrequiresstrongmagneticfieldsthatmustvaryorpulseinordertogenerateanelectricfield.Theimpactsofnanotechnologiesinthiscaseare:nanofabricationofcompoundsandalloysthat produce better hightemperature conducting materials. This will allow reducing the size of thedevice and the cryogenic environment needed for the performance of superconducting magnets.Nanoparticlethinfilmsarealsobeingdevelopedasshieldingmaterials.Numerous new nanoscale magnetometer designs are being developed, one of which is the opticalatomicmagnetometer.Thisinstrumentisbasedontheinteractionoflaserlightwithatomsorientedinamagneticfieldinagasphase.Theinstrumentmeasuresthechangeinalignmentwhenatomswithamagneticspinmomentinteractwithabeamofalaser.Intheabsenceofamagneticfieldtheatomswillalign with the electric and magnetic field of the laser crossing the atoms. Any perturbation by amagnetic field will disorient the alignment with the beam, reducing the amount of light transmittedthoughtthegas.AprototypeofsuchasystemwasdevelopedbyNIST(U.S)containingabout100billionatomsofrubidiumgasinavialthesizeofagrainofrice.Thechangeofspinwaseasilydetectableandscalable to much smaller sizes. The NIST prototype was able to detect the heartbeat of a rat.Researchers predict that with the small size and high performance such sensors could lead tomagnetocardiogramsthatprovidesimilarinformationtoanelectrocardiogram(ECG)withoutrequiringelectrodes on the patients body, even from outside clothing. This technique could become a realisticalternativetoMRIandPETimaging,withoutinjectionofcontrastenhancementagentsortracers.Whatisreallyexcitingisthatevenwiththelaserandheatingcomponents,thisnewdeviceuserelativelylowpower and can be extremely small compared to any current magnetic stimulation device. It could beonedaypossiblethatsensorswillbeusedtomakeportableMEGhelmetsforbrainmachineinterfaces. Page32of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433 33. NANOYOUTeachersTrainingKitModule2Chapter1Although much progress has been made, and much research is underway, there are some majorobstaclesto overcomebeforebrainmachineinterfacesbecomeareality.Wirelesssignal transmissionfrom brain implants is still futuristic, along with wearable magnetic brainmachine interfaces. Anotherobstacleistheoptimizationofthemicroelectrodesthatrecordneuronalactivity,whichtendtodegradein time due to biofilm formation. Risk of infection is also a major challenge. Better engineering ofinterfacesusingnanoengineeredmaterialsisneededtoimprovebiocompatibility,durability,andallowlowerstimulationpotentials.ELSATOPIC:Neuroprostheticsbringalong anumberofELSAissues.Normallythesetypesofdevicesareonlyconsidered when other pharmacologic and neurosurgical options have been exhausted. Bioengineers andmedicalengineersclaimthattheirroleshouldbetocompensateforabodysdeficit(asresultofanaccident,oradisease),notatreplacinganyremainingfunction.Itshouldnotleadtoenhancementofhumancapabilities.Nevertheless nanotechnologies are making those developments more feasible and affordable, obligingresearchers in the field, as well as regulators, ethicists and sociologist to reflect on the social, medical andethical consequences of these devices. The same applies to the concept of cognitive prosthesis, a systemdevelopedtosupportandaugmentthecognitiveabilitiesofitsuser.Ofcoursesuchdevicescouldbeextremelyuseful to people with impaired communication. However the concept does raise the possibility ofenhancement of cognitive capabilities for other uses. The idea of course also raises many social, ethical andregulatory questions. For instance one is the accessibility to these types of devices. Are these going to beaffordableforeverybody,coveredbythesocialhealthsystems,oravailableonlytowealthypeople?Isitethicaltohaveatechnologythatcanaugmentapersonsmentalability,butonlyifshe/hecanaffordit?Howisthistechnology going to be regulated in those circumstances where the mental/cognitive ability of numerousindividualsisassessed(jobinterviews,competitionsetc.)?Page33of33TheresearchleadingtotheseresultshasreceivedfundingfromtheEuropeanCommunitysSeventhFrameworkProgramme (FP7/20072013)undergrantagreementn233433