Microscopie non-linéaire pour l'imagerie des cordes vocales

Les cordes vocales sont deux replis de tissus loges dans le larynx et essentiels a la phonation. Malgre l'apparente simplicite de leur principe de fonctionnement, leur structure interne est complexe. Chaque corde est composee de dierentes couches de tissus aux proprietes mecaniques variables. La technique de reference pour les etudier, l'histologie, presente l'inconv enient d'^etre tres invasive. La microscopie non-lineaire est une technique d'imagerie optique qui permet d'imager des echantillons en profondeur de maniere non-invasive. De plus, elle ore des modes de contraste qui sont interessants pour visualiser des proteines breuses, l'elastine et le collagene, en partie responsables des proprietes mecaniques des tissus epitheliaux. Ce projet de recherche s'interesse a evaluer la capacite de la microscopie non-lineaire a fournir des performances similaires a l'histologie dans le cadre de l'imagerie des cordes vocales. L'objectif principal est de developper un microscope non-lineaire pour evaluer la capacite de ce type d'imagerie a observer des cordes vocales. La demarche choisie est de faire dans un premier temps des images de coupes nes de cordes vocales pour les comparer a une technique de reference, l'histologie. Dans un second temps, des tests visant a transcrire les resultats obtenus dans le cadre de l'imagerie in vivo ont ete eectues. Un systeme d'imagerie non-lineaire multimodal a ete concu speciquement pour permettre les experiences realisees pendant ce projet. Il a en particulier ete equipe pour prendre des images a grand champ de coupes de cordes vocales. Pour ce faire, il a fallu de creer un programme de contr^ole du microscope, et un autre de traitement des images obtenues. Apres avoir ete assemble, le systeme a ete teste pour verier que ses performances s'approchent des limites theoriques denies dans la litterature. Des coupes nes de cordes vocales ont ete obtenues aupres de l'equipe du Pr. Christopher J. Hartnick du Massachusetts Eye and Ear Inrmary, Harvard Medical School. Les specialistes de cette equipe ont analyse les images histologiques pour extraire de l'information structurelle sur les cordes vocales. Une correlation a ete obtenue entre ces informations et les analyses faites sur les images non-lineaires des m^emes echantillons. Une premiere etape pour evaluer la possibilite de transcrire ces resultats vers l'imagerie in vivo a ete realisee durant ce projet. Un larynx de porc a ete obtenu et les cordes vocales en ont ete extraites pour faire des images. Cette experience a montre qu'il est possible de localiser les dierentes macrostructures des cordes vocales avec la microscopie non-lineaire.

Abstract

The vocal cords are two folds of epithelial tissues located in the larynx and are involved in production of the human voice. Despite their apparent simplicity, their internal structure is complex. Each fold can be divided into several layers with dierent mechanical properties. The gold standard for studying their structure { histology { has the inconvenience of being very invasive. Non-linear microscopy is an optical imaging technique which allows images to be taken in depth within samples in a non invasive manner. It also oers intrinsic contrasts, allowing the identication of certain brous proteins { elastin and collagen { which are responsible for the mechanical properties of epithelious tissues. The main goal of this research project was to assess nonlinear microscopy's performances for vocal fold imaging. The study has been broken down in two separate tasks. The rst one was to evaluate the nonlinear modalities contrast against histology. For that purpose, we chose to rst take images of thin samples and compare them to the corresponding histological slides. The second task was to make tests to transcribe the results obtained to in vivo imaging. A custom-built nonlinear imaging system was used for these experiments. It was developped to allow acquisition of wide-eld images. A C++ based software was developped to control the microscope and allow treatment and visualization of the images. After being built, the system was further tested to check its performances in comparison with the theoretical limit as described in the literature. Thin slices of vocal folds were obtained from the team of Pr Christopher J. Hartnick from Massachusetts Eye and Ear Inrmary, Harvard Medical School. Specialists from his team analysed the histological samples to extract structural data from the vocal folds. A good correlation was measured between histological and nonlinear data. A rst step in evaluating the possibility for translating these results towards in vivo imaging was performed during this project. A swine's larynx was obtained, and vocal folds were extracted for imaging purposes. This experiment showed that it is indeed possible to localize various macrostructures of the tissues with nonlinear microscopy.