L'œsophage et la trachée proviennent du diverticule endodermique du tube de l'intestin antérieur au cours de l'embryogenèse. Des événements cellulaires et moléculaires bien régulés et organisés entraînent la séparation du tube de l'intestin antérieur en œsophage et trachée. Cette séparation est encore mal connue et la perturbation de ce processus se traduit par une anomalie congénitale sévère telle qu'une l’atrésie de l'œsophage avec ou sans fistule trachéo-œsophagienne (AO/FTO). L'AO/FTO est l'une des malformations congénitales gastro-intestinales les plus courantes affectant 1 naissance sur 3000. Cette malformation nécessite une intervention chirurgicale urgente à la naissance et est fréquemment associée à une morbidité à long terme. Les mécanismes sous-jacents au développement embryonnaire de l'AO/FTO sont mal compris. Les modèles animaux ont été largement utilisés pour comprendre les maladies humaines depuis des décennies et ont considérablement contribué à la compréhension du développement de l'œsophage. Cependant, des différences structurelles et morphologiques clés existent entre l'œsophage humain et animal, ce qui nécessite un modèle plus fiable pour comprendre le développement trachée-œsophagien. Les cellules souches pluripotentes induites par l'homme ont été un outil précieux pour comprendre l'organogenèse en imitant le développement et en déchiffrant les mécanismes qui conduisent à des maladies congénitales et acquises. Cette thèse se concentre donc sur l'utilisation de cellules souches pluripotentes induites (IPS) par des patients pour déchiffrer les mécanismes de signalisation impliqués dans le développement de l'œsophage et les maladies congénitales telles que l’OA/FTO. Il étudie également l'une des maladies œsophagiennes acquises possibles, comme l'œsophage de Barrett. Nous avons orienté la différenciation des IPS saines et dérivées de patients vers différents stades de développement, tels que l'endoderme définitif, l'intestin antérieur, l'épithélium œsophagien et trachéal. De plus, l'épithélium œsophagien a été développé davantage dans un environnement tridimensionnel sans matrice pour générer des organoïdes œsophagiens matures. À chaque étape de la progression du développement, des analyses d'immunofluorescence, de qPCR et de séquençage d'ARN ont été effectuées. Nos résultats suggèrent que l'expression des marqueurs endodermiques CXCR4, SOX17, et GATA4 était similaire dans les cellules différenciées des patients et des cellules saines. Cependant, au stade de l'intestin antérieur, nous avons observé une diminution significative de l'expression des gènes et des protéines du facteur transcriptionnel clé SOX2 dans les cellules dérivées du patient. De plus, en utilisant le séquençage d'ARN à molécule unique, nous avons observé que les gènes critiques GSTM1, et RAB37 impliqués dans la morphogenèse cellulaire et associés à l’OA/FTO étaient dérégulés au stade de l'intestin antérieur dans les cellules dérivées du patient. Nous avons également observé une augmentation significative de l'expression du facteur de transcription NKX2.1 habituellement exprimé uniquement dans les cellules trachéales, dans l'épithélium oesophagien dérivé du patient. NKX2.1 est maintenue dans les organoïdes oesophagiens matures même après 2 mois. Ensuite, nous voulions valider l'utilisation potentielle de nos organoïdes dérivés des IPS pour modéliser les maladies acquises de l'œsophage telles que l'œsophage de Barrett. Nous avons induit une métaplasie ou transformation épithéliale avec surexpression de BMP4 dans des organoïdes de l'œsophage sains et dérivés du patient sur une période d'un mois. Nos résultats préliminaires montrent que les organoïdes de l'œsophage dérivés des patients exprimaient des niveaux d'ARNm plus élevés de MUC5AC, un marqueur épithélial cylindrique par rapport au groupe sain. Cela suggère une plus grande sensibilité de l'organoïde de l'œsophage dérivé du patient aux changements epitheliales métaplasiques. En conclusion, nous avons développé les premiers organoïdes œsophagiens tridimensionnels matures sans matrice différenciés des patients OA/FTO et identifié une signature moléculaire unique dans les cellules dérivées du patient au cours de la différenciation dirigée de l'œsophage. De plus, sur la base des résultats préliminaires, nous avons pu confirmer l'incidence plus élevée de l'œsophage de Barrett chez les patients OA/FTO par rapport au groupe sain. Notre travail met donc en évidence l'importance de l'utilisation des IPS dérivées des patients pour modéliser les maladies œsophagiennes congénitales et acquises afin de fournir de nouvelles informations sur le développement des organes au cours de l'embryogenèse. / The esophagus and trachea originate from the endodermal diverticulum of the anterior foregut tube during embryogenesis. Well-regulated and organized cellular and molecular events result in the compartmentalization of the anterior foregut tube into the esophagus and trachea. This compartmentalization is still poorly understood and disruption in this process results in a severe congenital anomaly such as esophageal atresia with or without tracheoesophageal fistula (EA/TEF). EA/TEF is one of the most common gastrointestinal congenital defects affecting 1 in 3,000 births. This malformation requires urgent surgery at birth and is frequently associated with long-term morbidity. The mechanisms underlying the embryonic development of EA/TEF are poorly understood. Animal models have been widely used to understand human diseases for decades and have significantly contributed to the understanding of esophageal development. However, key structural and morphological differences exist between human and animal esophagus, thus necessitating a more reliable model to understand trachea-esophageal development. Human induced pluripotent stem cells (iPSC) have been a valuable tool to understand organogenesis by mimicking development and deciphering mechanisms that lead to congenital and acquired diseases. This thesis therefore focuses on the use of patient-derived induced pluripotent stem cells to decipher signaling mechanisms involved in esophageal development and congenital diseases such as EA/TEF. It also focuses on one of the possible acquired esophageal diseases, namely, Barrett’s esophagus. We directed the differentiation of healthy and patient-derived iPSCs toward different developmental stages, such as definitive endoderm, anterior foregut, esophageal and tracheal epithelium. Furthermore, the esophageal epithelium was matured further in a matrix free 3-dimensional environment to generate mature esophageal organoids. At each stage of development progression, immunofluorescence, qPCR, and RNA sequencing analysis were performed. Our findings suggest that the expression of endodermal markers CXCR4, SOX17, and GATA4, were similar in both patient and healthy differentiated cells. However, at the anterior foregut stage, we observed a significant decrease in the gene and protein expression of key transcription factor SOX2 in patient-derived cells. Furthermore, using nanopore RNA sequencing, we observed that critical genes GSTM1, and RAB7 involved in cellular morphogenesis and associated with EA/TEF to be dysregulated at the anterior foregut stage in patient-derived cells. We also observed a significant increase in the expression of transcription factor NKX2.1, usually expressed only in tracheal cells, in the patient-derived esophageal epithelium. NKX2.1 expression was maintained in matured esophageal organoids even after 2 months. Next, we wanted to validate the potential use of our PSC-derived organoids to model acquired esophagus diseases such as Barrett’s esophagus (BE). We induced epithelial metaplasia with BMP4 overexpression in healthy and patient-derived esophagus organoids over a 1-month period. Our preliminary results show that patient-derived esophagus organoids expressed higher mRNA levels of MUC5AC, an epithelial columnar marker compared with the healthy group. This suggests a higher susceptibility of patient-derived esophagus organoid to metaplastic changes. In conclusion, we developed the first matrix free mature 3-dimensional esophageal organoids differentiated from EA/TEF patient-derived and identified a unique molecular signature in patient derived cells during directed esophagus differentiation. Furthermore, based on the preliminary results, we could confirm the higher incidence of Barrett’s esophagus in EA/TEF patients compared with the healthy group. Our work therefore highlights the significance of using patient-derived iPSCs to model congenital and acquired esophageal diseases to yield new insights on organ development during embryogenesis. It lays the foundation for a personalized medical approach to other diseases and the ones affecting the whole gastrointestinal system in both children and adults.

Nowadays, natural polymeric materials extracted from plants are the new alternatives for synthetic chemicals in water and wastewater treatment. The aim of this study is to evaluate the ability of Cassia fistula seed gum (CFG) as a coagulant aid with PAC in the treatment of textile wastewater. Jartest experiments were carried out to identify the optimal parameters of coagulation-flocculation for removing color and COD in synthesis wastewater containing Methyl blue and RB21 dyes, including pH, settling time, PAC dose, the optimal CFG dosage in comparing with the cationic polymer. After that, actual textile wastewater was treated by using PAC, PAC plus cationic polymer, and PAC plus CFG for evaluating the role of CFG. CFG supplementation has assisted the process effects at nearly 98% color, 85% COD for RB21 and 90% color, 70% COD for MB at the best dose of CFG 0.15 mL and 0.1 mL, respectively. The optimized parameters for the coagulation of real textile wastewater using PAC were pH = 6 and dose = 0.6 mL can removal 66% of color. By adding CFG to PAC, the efficient of treatment was increased about 70% even at the lower dosage of PAC and CFG (0.5 mL for each reagent). The yield of combining PAC and polymer was a little bit lower than PAC and CFG, for instant 68% color was decreased at the same condition. These achievements demonstrated a workable substitute of natural products such as Cassis fistula seed gum for synthetic chemical products in coagulation-flocculation process. / Hiện nay các loại vật liệu sinh học chiết xuất từ thực vật đang được nghiên cứu ứng dụng trong xử lý nước và nước thải thay cho các chất hóa học. Mục tiêu của nghiên cứu này là đánh giá hiệu quả của việc sử dụng gum được chiết xuất từ hạt cây Muồng Hoàng Yến (MHY) làm chất trợ keo tụ trong xử lý nước thải dệt nhuộm. Thí nghiệm Jartest được tiến hành nhằm xác định các điều kiện tối ưu cho quá trình xử lý nước thải tổng hợp chứa thuốc nhuộm Methyle Blue (MB) và RB21 bao gồm pH, thời gian lắng, liều PAC, liều gum MHY và liều polymer. Sau đó tiến hành xử lý nước thải thật với các điều kiện thích hợp đã xác định nhằm đánh giá vai trò của gum MHY. Gum MHY làm tăng hiệu quả của quá trình xử lý, đạt gần 98% đối với độ màu, 85% COD đối với RB21, 90% độ màu và 70% COD đối với MB với liều lượng tương ứng là 0,15 mL và 0,1 mL. Các thông số tối ưu cho quá trình xử lý trên mẫu nước thải thật là pH = 6, liều PAC = 0.6 mL có thể làm giảm 66% độ màu. Bổ sung gum MHY làm chất trợ keo tụ giúp gia tăng hiệu quả xử lý màu lên 70% dù với liều lượng rất thấp là 0,5 mL. Hiệu suất xử lý khi sử dụng kết hợp PAC và polymer thấp hơn trong trường hợp sử dụng PAC và gum MHY, cụ thể khoảng 68% độ màu được xử lý ở cùng một điều kiện. Những kết quả này cho thấy tiềm năng của việc sử dụng các vật liệu gum tự nhiên nhằm thay thế cho các hợp chất hóa học trong các quá trình keo tụ tạo bông để xử lý nước thải.

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