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Microspectroscopieetimagerievibrationnelles:desoutilsphotoniquesd’intérêt
enbiologieetdansledomainebiomédical
ValérieUntereiner
CHU DE REIMS
BioSpecT(BioSpectroscopieTranslationnelle),EA7506Plate-formed’ImagerieCellulaireetTissulaire(PICT)
UniversitédeReimsChampagne-Ardenne
1
Wavenumber(∝Frequency∝Energy)(cm-1)
Absorbance(u
.a.)
Interestofvibrationalbiospectroscopy
2
CH2:asymetricstretching
CH2:symetricstretching
CH3:umbrellabend
CH2:scissorbend
CH2:rocking
Nondestructivelight/matterinteraction
Label-free
Micrometerresolution
Vibrationalmicro-spectroscopies
Ramanscattering
Sourcelaser
Infraredabsorption
SourceIR
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Insituanalysisofthechemicalbonds,withoutalteringtheintegrityofthesample
Highmolecularandstructuralspecificity
IRspectrumofabiologicalsample
1656cm-1 1669cm-1 1665cm-1
Secondarystructureofproteins
Interestofvibrationalbiospectroscopy
Ourcoreexpertise:vibrationalspectroscopyforbiologicalapplicationsandbiomedicaltranslation
Vibrationalspectrum=Molecularsignatureofthesampleinaspecificphysiologicalorpathophysiologicalstate
Exvivoandinvivosamples
Cell,tissue,biofluid
Live,frozenorfixed
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Acquisition Dataprocessing
Detectionandcharacterizationofphysiologicalor
pathologicalconditions
Identificationofspectroscopicbiomarkers
Discriminantvibrationsindependentofthebiologicalvariabilityusingmultivariatestatisticaldataprocessing
Ramanspectralanalysisonlivercells
Infraredspectralanalysisoncolontissue
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Vibrationalspectroscopyforbiologicalapplicationsandbiomedicaltranslation
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Ramanimagingofasingleliversinusoidalendothelialcell(LSEC)
SzafraniecEetal.,2019
Ø Colon-lastpartofthedigestivesystem
Ø Cecum,colon,rectumandanalcanal-largeintestine
ColonanatomyandhistologyHEstainedsections(non-tumoral)
(a)Transverse(b)Longitudinal
Source:BlueHistology,Univ.ofWesternAustralia
(a)
(b)
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(Source:cancerinformaiton.org.uk)
HEstained
ColorectalcancerØ Lossoffunctionalcontrolinthenormalcellsliningthecolonorrectum
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Candidatemethod:infraredspectralimaging
Ø Screeningmethods:Fecaloccultbloodtest(FOBT),colonoscopyandsigmoidoscopy,etc
Currentscreeninganddiagnosticmethods
Presentneed:Newerclassificationmethodsprovidingbiomolecularinformationfromcellsandtissuescomplementarytohistopathology
Diagnosisconfirmation:HISTOPATHOLOGY
Automatedapproachesforrapiddiagnosis
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10
X
σ
Y
hyperspectralimageDatacube(X!Y!σ)!
Spectralimaging
Tensofthousandsofspectra/image
Pixelsize:1–10µm
Uncolouredtissuesection,
possiblyparaffin-embedded
Millimeter
Millim
eter
HPSstainedtissue(5Xmagnification)
K-meansclusteredIRimage
Non
-tum
oraltissue
Tu
moraltissue
K-meansclusteringresultsoftheIRspectralimagesusingrandompseudo-colors
Crypts(innerpart)Crypts(outerpart)LaminapropriaSubmucosa
TumorTumorassociatedstroma
Others-notattributedtoany
histologicalclass
Histologicalattribution
Annotationbyanexpertpathologist
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Libraryofspectralmarkers
Spectrafromdifferentcolonichistologicalclasses
1 Tumorepithelium
2 Crypt-outerpart
3 Crypt-innerpart
4 Muscularispropria
5 Tumorassociatedstroma
6 Laminapropria
7 Submucosa
8 Bloodvessel
Confusionmatrix
1080-1300cm-1
Constructionandapplicationofthepredictionmodelbasedonlineardiscriminantanalysis
Internalvalidation
(1/3rdspectra)
Trainingset(2/3rdspectra)
Predictionmodel
Testset(identificationofunknownsamples)
(N=71samples)
posteriorprobability0.5
(N=9samples)
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EXTERNALVALIDATIONIdentificationofunknowncolonictissuesbyspectralhistopathology
(No.ofspectratested:3620287)
HPSstainedcolontissue(5Xmagnification)
IR-LDApredictedimage
Nontumoral
Finaldiagnosis
Adenocarcinoma
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Individualsatriskofthedisease Diseaseatan
earlystage
Overtdisease
Reponsetotreatment
Personalizedtreatment
Recurrence
Spectroscopicbiomarkers Diagnosis
Prognosis
Predictionofthetherapeuticresponse
Monitoring
ScreeningScreening
Translationaldevelopmentofvibrationalspectroscopy
Monitoring
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EquipeBioSpecT:BioSpectroscopieTranslationnelle
Plate-formeIBiSAd’ImagerieCellulaireetTissulaire(PICT)