Investigating the Role of Fucosylation on the Stemness of Human CD34+ Mobilized Peripheral Blood Progenitor Cells
Jana MS Thesis - Investigating the Role of Fucosylation on the Stemness of Human CD34+ Progenitor Cells.pdf
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Committee membersOrlando, Valerio
KAUST DepartmentBiological and Environmental Science and Engineering (BESE) Division
Embargo End Date2024-02-05
Permanent link to this recordhttp://hdl.handle.net/10754/687488
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Access RestrictionsAt the time of archiving, the student author of this thesis opted to temporarily restrict access to it. The full text of this thesis will become available to the public after the expiration of the embargo on 2024-02-05.
AbstractIt has been well-established that the process of stem cell homing is initially mediated by E-Selectin, a cell adhesion molecule constitutively expressed on the bone marrow vasculature. The ligand for E-selectin is a carbohydrate modification known as sialyl-Lewis X (sLex) found mainly on proteins, and it has been shown that ex vivo fucosylation of stem cells, including hematopoietic stem cells (HSCs) enhances these ligands, resulting in more efficient delivery of stem cells to their home in the bone marrow. However, the exact biological effects that fucosylation has on HSC function has not been extensively studied. In vivo mouse experiments from our lab where short-term CD34+ hematopoietic stem cells were fucosylated improved their delivery to the bone marrow but also exhibited improved longevity and apparent stemness as assessed by secondary transplantation. Therefore, to investigate the role fucosylation has on this phenotype and to uncover whether E-Selectin binding is also required alongside it to trigger molecular changes in hematopoietic stem cells, we set up in vitro cultures with CD34+ cells from GCSF-mobilized human peripheral blood (mPB-CD34+) that had been either left untreated or treated with fucosyltransferase VI (FUT6) in the presence and absence of recombinant E-selectin protein as well as the fucosylation inhibitor 2-fluorofucose (2-FF). We then performed characterization assays to assess cell cycle, signaling, differentiation, and viability using flow cytometry, western blotting, Giemsa staining, and a variety of viability assays. We found that fucosylation enhances the effects of E-Selectin binding, activating stem cell proliferation, triggering the PI3K/AKT/NFkB, P38, and EGFR pathways, induces a transient increase in pre-apoptotic cells, and may alter cell differentiation. These results uncover the role of fucosylation in hematopoietic stem cells and highlights the PI3K/AKT/NFkB pathway as a signaling route mediated by E-selectin to influence stem cell longevity.
CitationMalki, J. (2023). Investigating the Role of Fucosylation on the Stemness of Human CD34+ Mobilized Peripheral Blood Progenitor Cells [KAUST Research Repository]. https://doi.org/10.25781/KAUST-524UJ