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Présenté Pour L'obtention Du Diplôme De MAGISTER En N° d’ordre :19/2009-M/SN REPUBLIQUE ALGERIENNE DEMOCRATIQUE ET POPULAIRE MINISTERE DE L’ENSEIGNEMENT SUPERIEUR ET DE LA RECHERCHE SCIENTIFIQUE UNIVERSITE DES SCIENCES ET DE LA TECHNOLOGIE HOUARI-BOUMEDIENE FACULTE DES SCIENCES BIOLOGIQUES MEMOIRE Présenté pour l’obtention du diplôme de MAGISTER En : Sciences de la Nature Spécialité : Biotechnologie et Santé Par : Melle GUESTINI FOUZIA THEME Soutenu publiquement le: 09/12/2009, devant le jury composé de: Mme, Laraba-Djebari F. Professeur (USTHB) Présidente Mme, Hammoudi-Triki D. Maitre de Conférences (USTHB) Directrice de Thèse Mme, Touil-Boukoffa C. Professeur (USTHB ) Examinatrice Mme, Oussedik-Oumehdi H. Maitre de Conférences (USTHB) Examinatrice Mme, Hamouli Z. Maitre de Conférences (USTHB) Examinatrice Remerciements Mes premiers remerciements vont à ma directrice de thèse, le Dr. Hammoudi-Triki Djelila, pour m'avoir offert l'opportunité de réaliser ce projet, pour ses conseils précieux, ses orientations scientifiques, ainsi que son optimisme affiché malgré toutes les contraintes, pour son encadrement tant apprécié assuré pendant l’expérimentation et la rédaction de la thèse, ses encouragements et ses conseils judicieux. J'ai beaucoup appris, notamment grâce à la grande liberté qu’elle m'a accordée quant à l'accomplissement de mon projet. Je voudrais vivement remercier les membres du jury pour la correction de ma thèse et le jugement de mon travail, et particulièrement le Pr. Laraba-Djebari Fatima, présidente du jury pour l’accès accordé à certains équipements au sein du Laboratoire de l’Institut Pasteur. J’adresse aussi mes remerciements au Professeur Touil-Boukouffa sa rigueur et son esprit scientifique seront toujours ma référence, je la remercie également pour les cours prodiguées durant mon année théorique et pour m’avoir encore plus fait aimer la recherche. Je souhaite sincèrement remercier Mme Oussedik et Mme Hammouli pour avoir examiné et jugé mon travail, leurs remarques ne seront que constructives et me permettront de mieux entrevoir mes lacunes. Tous mes remerciements également à toute l’équipe du laboratoire Contrôle Qualité de l’Institut Pasteur d’Alger. D’abord à Mme Touabti, je tiens à lui exprimer toute ma reconnaissance pour la confiance qu’elle m’a témoignée en m’accueillant dans son laboratoire, à Mr. Segouani, sa disponibilité, ses conseils et ses compétences scientifiques m’ont permis de mener à bien mes travaux, d’enrichir mes connaissances et de me former à des techniques nouvelles. Je remercie chaleureusement Ait-Said Lamia, pour son soutien moral et ses conseils dans les moments difficiles, j’ai trouvé en elle la loyauté d’une amie et la sincérité d’une sœur, qu’elle trouve dans ces remerciements une promesse d’une amitié sincère. Je tiens également à exprimer ma reconnaissance aux autres membres du laboratoire pour leur gentillesse et pour m’avoir intégrer dans leur équipe, je garderai de très bons souvenirs de cette année passée parmi vous Ahlem, Nacira, Mme Boukechabia, Maya, Khadidja, Lila, Chahinez, Fouzia… et sans oublier Sid Ahmed, Ali, Mustafa, Karim… pour toute l’aide fournie au niveau de l’animalerie. A Mr Kézar pour les nombreuses fois ou il m’a attendu après son service. Je n’oublierai évidemment pas de remercier tous mes collègues de travail en particulier Nadjia pour les coups de main, à Hayet Rafa pour ses conseils aux moments de doutes, pour les informations cruciales et sa sincérité, Ipticem, Hadjer, Hayet et Amina pour l’aide qu’elle m’a apporté quant à la préparation des TD. Je voudrais sincèrement et fièrement remercier mes parents, ma tendre et bien aimée mère, pour sa confiance, son soutien et son ouverture d'esprit, je la remercie aussi pour ses encouragements et pour l’AMOUR qu’elle me porte, mon père pour ces encouragements depuis l’enfance à poursuivre mes études. A mes sœurs : Amina et Nesrine, pour leur amour et leur compréhension, leur aide et surtout leur patience, je déplore mes absences, j’espère que dieu me donnera l’occasion de me rattraper. Bien sure, je remercie chaleureusement Mohamed Abdelaziz, tous les plus tendres mots n’exprimeront pas toute ma gratitude, pour tous les services rendus, tes encouragements, ton soutien et ton aide tous le long de cette période mélancolique que nous avons traversé. Un GRAND merci à mes meilleures amies Maya, Selma, Nesrine, Nadjwa et Fifi pour avoir toujours été là pour moi, surtout aux moments où j’avais le plus besoin d’elles. Merci pour les agréables moments passés avec vous après de longues semaines de travail interminables. Merci aussi à mes nouveaux amis (es) du groupe du CCE, en particulier Samira, pour les fous rires partagés, leur soutien et l’agréable sortie organisée en montagne. Je remercie cordialement mon cousin Lotfi, pour avoir été présent dans les moments difficiles, pour son aide précieuse, son soutien moral et matériel. Que dieu le garde. Je remercie également Abderrahim, pour son aide précieuse, son soutien moral et pour m’avoir consacré une place dans son bureau. En outre je remercie chaleureusement toutes les personnes qui ont été sincères avec moi. Que dieu les garde, et emplisse leur vie de bonheur. Je tiens à achever mes remerciements en rendant un hommage à un grand homme de foie et de science le Pr. AYAD qui nous a quitté prématurément. Il gardera à jamais une grande place dans nos cœurs. Je voudrais également rendre hommage à Tata Zoulikha, une femme dont la gentillesse, la modestie et la chaleur humaine réchauffai le cœur de plus d’un, que dieu ai leurs âmes. Et Merci aux oubliés… SOMMAIRE Sommaire LISTE DES ABREVIATIONS INTRODUCTION…………………………………………………………………………….1 CHAPITRE I. Généralités Envenimation scorpionique 1. Les scorpions………………………………………………………………………………..3 2. Les espèces dangereuses…………………………………………………………………….4 2.1. Espèces de l'Ancien Monde……………………………………………………………….4 2.2. Espèces du Nouveau Monde………………………………………………………………4 3. Les venins de scorpion………………………………………………………………………5 4. Les toxines du venin de scorpion……………………………………………………………5 4.1. Les toxines longues actives sur les canaux sodium……………………………………….6 4.2. Les toxines courtes actives sur les canaux potassium……………………………………..7 4.3. Les toxines actives sur les canaux calcium………………………………………………..8 4.4. Les toxines actives sur les canaux chlore………………………………………………….8 4.5. Les toxines de scorpion sans ponts disulfures…………………………………………….9 5. Les toxines du venin de scorpion Androctonus australis hector……………………………9 6. Rôles des toxines de scorpion dans l’ingénierie…………………………………………...10 6.1. Les toxines utilisées dans la lutte contre les insectes…………………………………….10 6.2. Effets antibactérien et antifongique du venin de scorpion……………………………….11 6.3. Activité anti-tumoral du venin de scorpion……………………………………………...12 6.4. Utilisation des constituants du venin de scorpion dans l’immunothérapie……………....12 6.5. Autres activités…………………………………………………………………………...13 Stress oxydant 7. Le stress oxydant…………………………………………………………………………...14 SOMMAIRE 7.1. Les espèces réactives de l’oxygène………………………………………………………15 7.2. Conséquences du Stress oxydant………………………………………………………...16 7.2.1. Oxydation des protéines et protéolyse…………………………………………………17 7.2.2. Attaque radicalaire de l’ADN………………………………………………………….21 7.2.3. Peroxydation lipidique…………………………………………………………………22 8. Les systèmes antioxydants…………………………………………………………………23 8.1. Système de défense Non-enzymatique…………………………………………………..23 8.1.1. Le glutathion…………………………………………………………………………...23 8.1.2. Autres molécules anti-oxydantes………………………………………………………25 8.2. Système de détoxification enzymatique………………………………………………….26 8.2.1. Fonction de détoxification des glutathion S-transférases……………………………...26 8.2.2. Les glutathion peroxydases…………………………………………………………….26 8.2.3. La glutathion réductase………………………………………………………………...27 8.2.4. La thioredoxine réductase ……………………………………………………………..27 8.2.5. Les superoxyde dismutases……………………………………………………………27 8.2.6. Les catalases……………………………………………………………………………28 CHAPITRE II. Matériel et Méthodes I. Matériel…………………………………………………………………..…………………31 1. Matériel biologique………………………………………………………………………...31 2. Choix des modèles d’études………………………………………………………………..31 3. Réactifs et produits chimiques……………………………………………………………..32 II. Méthodes…………………………………………………………………………………..32 1. Etude de l’effet du venin d’Androctonus australis hector sur la viabilité cellulaire………32 1.1. Activation des cellules spléniques avec le venin………………………………………...33 a. Préparation des cellules spléniques………………………………………………………...33 SOMMAIRE b. Mise en culture des cellules spléniques……………………………………………………33 c. Mesure de la viabilité des cellules spléniques……………………………………………...33 1.1.1. Détermination de la cytotoxicité du venin sur les cellules de rate de souris envenimées ex vivo………………………………………………………………………………………...34 1.1.2. Effet du temps d’incubation sur la cytotoxicité du venin d’Androctonus australis hector sur les splénocytes in vitro…………………………………………………………………....34 1.1.3. Effet de la concentration du venin d’Androctonus australis hector sur la viabilité des splénocytes in vitro …..……………………………………………………………...............35 1.1.4. Etude de l’intégrité membranaire par la mesure de l’activité de la lactate déshydrogénase………………………………………………………………………………35 1.2. Activation des cellules Vero avec le venin………………………………………………36 a. Entretien des cellules Vero…………………………………………………………………36 b. Evaluation de la cytotoxicité du venin de scorpion sur la lignée Vero…………………….36 2. Etude de l’effet du venin d’Androctonus australis hector sur le métabolisme cellulaire….37 2.1. Quantification des cytokines pro- et anti-inflammatoires………………………………..37
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