title: Metabolic glycoengineering of mesenchymal stromal cells with N-propanoylmannosamine.
abstract: There is an increasing interest in the modification of cell surface glycosylation to improve the properties of therapeutic cells. For example, glycosylation affects the biodistribution of mesenchymal stromal cells (MSCs). Metabolic glycoengineering is an efficient way to modify the cell surface. The mammalian biosynthetic machinery tolerates the unnatural sialic acid precursor, N-propanoylmannosamine (ManNProp), and incorporates it into cell surface glycoconjugates. We show here by mass spectrometric analysis of cell surface N-glycans that about half of N-acetylneuraminic acid was replaced by N-propanoylneuraminic acid in the N-glycans of human umbilical cord blood-derived MSCs supplemented with ManNProp. In addition, the N-glycan profile was altered. ManNProp-supplemented cells had more multiply fucosylated N-glycan species than control cells. The fucosylated epitopes were shown in tandem mass spectrometric analysis to be Lewis x or blood group H epitopes, but not sialyl Lewis x (sLex). The amounts of tri- and tetra-antennary and polylactosamine-containing N-glycans also increased in ManNProp supplementation. In accordance with previous studies of other cell types, increased expression of the sLex epitope in ManNProp-supplemented MSCs was demonstrated by flow cytometry. In light of the N-glycan analysis, the sLex epitope in these cells is likely to be carried by O-glycans or glycolipids. sLex has been shown to target MSCs to bone marrow, which may be desirable in therapeutic applications. The present results represent the first structural analysis of an N-glycome of ManNProp-supplemented cells and demonstrate the feasibility of modifying cell surface glycosylation of therapeutic cells by this type of metabolic glycoengineering.
authors.affiliation: Finnish Red Cross Blood Service, 00310 Helsinki, Finland. suvi.natunen@veripalvelu.fi
authors.forename: Suvi
authors.initials: S
authors.lastname: Natunen
authors.forename: Milla
authors.initials: M
authors.lastname: Lampinen
authors.forename: Heli
authors.initials: H
authors.lastname: Suila
authors.forename: Ilja
authors.initials: I
authors.lastname: Ritamo
authors.forename: Virve
authors.initials: V
authors.lastname: Pitkänen
authors.forename: Alison V
authors.initials: AV
authors.lastname: Nairn
authors.forename: Jarkko
authors.initials: J
authors.lastname: Räbinä
authors.forename: Saara
authors.initials: S
authors.lastname: Laitinen
authors.forename: Kelley W
authors.initials: KW
authors.lastname: Moremen
authors.forename: Werner
authors.initials: W
authors.lastname: Reutter
authors.forename: Leena
authors.initials: L
authors.lastname: Valmu
mesh_terms: D054794:Glycomics; D006031:Glycosylation; D006595:Hexosamines / Q000378:metabolism*; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000378:metabolism*; D019158:N-Acetylneuraminic Acid / Q000378:metabolism; D009438:Neuraminic Acids / Q000378:metabolism; D009844:Oligosaccharides / Q000235:genetics / Q000378:metabolism; D000080506:Sialyl Lewis X Antigen
chemical_list: D006595:Hexosamines; C425503:N-propanoylneuraminic acid; D009438:Neuraminic Acids; D009844:Oligosaccharides; D000080506:Sialyl Lewis X Antigen; C076271:N-propanoylmannosamine; D019158:N-Acetylneuraminic Acid
keywords: N-propanoylmannosamine; glycoengineering; mesenchymal stromal cell; sialic acid; umbilical cord blood