title: O-GlcNAcylation Negatively Regulates Cardiomyogenic Fate in Adult Mouse Cardiac Mesenchymal Stromal Cells.
abstract: In both preclinical and clinical studies, cell transplantation of several cell types is used to promote repair of damaged organs and tissues. Nevertheless, despite the widespread use of such strategies, there remains little understanding of how the efficacy of cell therapy is regulated. We showed previously that augmentation of a unique, metabolically derived stress signal (i.e., O-GlcNAc) improves survival of cardiac mesenchymal stromal cells; however, it is not known whether enhancing O-GlcNAcylation affects lineage commitment or other aspects of cell competency. In this study, we assessed the role of O-GlcNAc in differentiation of cardiac mesenchymal stromal cells. Exposure of these cells to routine differentiation protocols in culture increased markers of the cardiomyogenic lineage such as Nkx2.5 and connexin 40, and augmented the abundance of transcripts associated with endothelial and fibroblast cell fates. Differentiation significantly decreased the abundance of O-GlcNAcylated proteins. To determine if O-GlcNAc is involved in stromal cell differentiation, O-GlcNAcylation was increased pharmacologically during the differentiation protocol. Although elevated O-GlcNAc levels did not significantly affect fibroblast and endothelial marker expression, acquisition of cardiomyocyte markers was limited. In addition, increasing O-GlcNAcylation further elevated smooth muscle actin expression. In addition to lineage commitment, we also evaluated proliferation and migration, and found that increasing O-GlcNAcylation did not significantly affect either; however, we found that O-GlcNAc transferase--the protein responsible for adding O-GlcNAc to proteins--is at least partially required for maintaining cellular proliferative and migratory capacities. We conclude that O-GlcNAcylation contributes significantly to cardiac mesenchymal stromal cell lineage and function. O-GlcNAcylation and pathological conditions that may affect O-GlcNAc levels (such as diabetes) should be considered carefully in the context of cardiac cell therapy.
authors.affiliation: Institute of Molecular Cardiology; Diabetes and Obesity Center, Department of Medicine, Division of Cardiovascular Medicine, University of Louisville, Louisville, Kentucky, United States of America.
authors.forename: Ayesha
authors.initials: A
authors.lastname: Zafir
authors.affiliation: Institute of Molecular Cardiology; Diabetes and Obesity Center, Department of Medicine, Division of Cardiovascular Medicine, University of Louisville, Louisville, Kentucky, United States of America.
authors.forename: James A
authors.initials: JA
authors.lastname: Bradley
authors.affiliation: Institute of Molecular Cardiology; Diabetes and Obesity Center, Department of Medicine, Division of Cardiovascular Medicine, University of Louisville, Louisville, Kentucky, United States of America.
authors.forename: Bethany W
authors.initials: BW
authors.lastname: Long
authors.affiliation: Institute of Molecular Cardiology; Diabetes and Obesity Center, Department of Medicine, Division of Cardiovascular Medicine, University of Louisville, Louisville, Kentucky, United States of America.
authors.forename: Senthilkumar
authors.initials: S
authors.lastname: Muthusamy
authors.affiliation: Institute of Molecular Cardiology; Diabetes and Obesity Center, Department of Medicine, Division of Cardiovascular Medicine, University of Louisville, Louisville, Kentucky, United States of America.
authors.forename: Qianhong
authors.initials: Q
authors.lastname: Li
authors.affiliation: Institute of Molecular Cardiology; Diabetes and Obesity Center, Department of Medicine, Division of Cardiovascular Medicine, University of Louisville, Louisville, Kentucky, United States of America.
authors.forename: Bradford G
authors.initials: BG
authors.lastname: Hill
authors.affiliation: Institute of Molecular Cardiology; Diabetes and Obesity Center, Department of Medicine, Division of Cardiovascular Medicine, University of Louisville, Louisville, Kentucky, United States of America.
authors.forename: Marcin
authors.initials: M
authors.lastname: Wysoczynski
authors.affiliation: Institute of Molecular Cardiology; Diabetes and Obesity Center, Department of Medicine, Division of Cardiovascular Medicine, University of Louisville, Louisville, Kentucky, United States of America.
authors.forename: Sumanth D
authors.initials: SD
authors.lastname: Prabhu
authors.affiliation: Institute of Molecular Cardiology; Diabetes and Obesity Center, Department of Medicine, Division of Cardiovascular Medicine, University of Louisville, Louisville, Kentucky, United States of America.
authors.forename: Aruni
authors.initials: A
authors.lastname: Bhatnagar
authors.affiliation: Institute of Molecular Cardiology; Diabetes and Obesity Center, Department of Medicine, Division of Cardiovascular Medicine, University of Louisville, Louisville, Kentucky, United States of America.
authors.forename: Roberto
authors.initials: R
authors.lastname: Bolli
authors.affiliation: Institute of Molecular Cardiology; Diabetes and Obesity Center, Department of Medicine, Division of Cardiovascular Medicine, University of Louisville, Louisville, Kentucky, United States of America.
authors.forename: Steven P
authors.initials: SP
authors.lastname: Jones
mesh_terms: D000117:Acetylglucosamine / Q000378:metabolism; D000818:Animals; D002454:Cell Differentiation; D019070:Cell Lineage; D002465:Cell Movement; D049109:Cell Proliferation; D017630:Connexins / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology; D015971:Gene Expression Regulation, Enzymologic; D006031:Glycosylation; D049452:Green Fluorescent Proteins / Q000378:metabolism; D000072576:Homeobox Protein Nkx-2.5; D018398:Homeodomain Proteins / Q000378:metabolism; D018942:Macrolides / Q000737:chemistry; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D051379:Mice; D008810:Mice, Inbred C57BL; D009206:Myocardium / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000166:cytology*; D017351:N-Acetylglucosaminyltransferases / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D015398:Signal Transduction; D014157:Transcription Factors / Q000378:metabolism; D000097003:Gap Junction alpha-5 Protein
chemical_list: D000117:Acetylglucosamine; D017630:Connexins; D049452:Green Fluorescent Proteins; D000072576:Homeobox Protein Nkx-2.5; D018398:Homeodomain Proteins; D018942:Macrolides; D017351:N-Acetylglucosaminyltransferases; D014157:Transcription Factors; D000097003:Gap Junction alpha-5 Protein; C484469:Nkx2-5 protein, mouse; C040929:bafilomycin A1; C449880:O-GlcNAc transferase