Design of an in vitro brain-like model for examining cytophagy of balamuthia mandrillaris in a three-dimensional manner
5
Issued Date
2022
Copyright Date
2022
Resource Type
Language
eng
File Type
application/pdf
No. of Pages/File Size
xii, 92 leaves
Access Rights
open access
Rights
ผลงานนี้เป็นลิขสิทธิ์ของมหาวิทยาลัยมหิดล ขอสงวนไว้สำหรับเพื่อการศึกษาเท่านั้น ต้องอ้างอิงแหล่งที่มา ห้ามดัดแปลงเนื้อหา และห้ามนำไปใช้เพื่อการค้า
Rights Holder(s)
Mahidol University
Bibliographic Citation
Thesis (M.Sc. (Biodesign in Medicine))--Mahidol University, 2022)
Suggested Citation
Worakamol Pengsart Design of an in vitro brain-like model for examining cytophagy of balamuthia mandrillaris in a three-dimensional manner. Thesis (M.Sc. (Biodesign in Medicine))--Mahidol University, 2022). Retrieved from: https://repository.li.mahidol.ac.th/handle/123456789/113936
Title
Design of an in vitro brain-like model for examining cytophagy of balamuthia mandrillaris in a three-dimensional manner
Author(s)
Advisor(s)
Abstract
Infection of Balamuthia mandrillaris, a free-living amoeba in soil and water, causes a fatal brain inflammation known as granulomatous amoebic encephalitis. Regardless of host immune status, more than a 95% mortality rate has been documented in all ages. The current treatment regimen relies on various combinations of antimicrobial and antifungal drugs; however, none of them possesses a radical cure, leading to death. Therefore, understanding the mechanism underlying pathogenicity is essential for identifying drug targets. Studies of pathogenesis and assessment of amoebicidal drugs rely on a culture of human cancer cells in a monolayered manner. Given a two-dimensional (2D) cell-cell interaction, the conventional culture models are not physiologically relevant to the human tissue. Therefore, translating data from the in vitro 2D models fails to recapitulate the complexity of human tissue. Spheroid, a cellular aggregate in a 3D form, reportedly provides a drug dose similar to that in the human body, implying more relevance to human tissue. This research aimed to demonstrate the use of 3D imaging and the spheroid for examining a process by which B. mandrillaris trophozoites ingest human cells, known as cytophagy. Human neuroblastoma SH-SY5Y cells were subjected to the spheroid formation by hanging-drop technique. A clinical isolate of pathogenic B. mandrillaris trophozoites was co-cultured with the 2D culture and the SH-SY5Y spheroid. The cytophagy was examined using a fluorescence-based and label-free microscope in a 3D manner. Host cell viability and apoptosis were examined. Differential expression of neuronal lineage markers in 2D and 3D cultures was performed by real-time PCR. B. mandrillaris co-cultured with SH-SY5Y monolayer cells exhibited a trogocytosis-like feeding mechanism, resulting in multiple protein-containing granules inside the trophozoite cytoplasm. However, preincubation of trogocytosis inhibitor failed to suppress the ingestion. Higher expression of neuronal maturation gene was shown in SH-SY5Y spheroid. Moreover, 3D live imaging revealed trophozoite transformation during neurospheroid invasion. In conclusion, spheroid culture can potentially demonstrate cell-parasite interaction in a 3D manner, which is more physiological relevance to the human brain than conventional monolayer culture. IMPLICATION OF THESIS: The significance of this study is to gain knowledge of the B. mandrillaris cytophagy mechanism. Results of the research can demonstrate the utility of 3D imaging and the spheroid for disease modeling, a platform necessary for identifying a therapeutic target in the context of the host-parasite interaction.
Degree Name
Master of Science
Degree Level
Master's degree
Degree Department
Faculty of Medicine Siriraj Hospital
Degree Discipline
Biodesign in Medicine
Degree Grantor(s)
Mahidol University