title: Ectopic expression of delta FBJ murine osteosarcoma viral oncogene homolog B mediates transdifferentiation of adipose-like spheroids into osteo-like microtissues.
abstract: Differentiation and transdifferentiation strategies have a large role in the manipulation of cells in replacing dysfunctional cells and tissues. We developed adipose-like microtissues using gravity-enforced self-assembly of monodispersed human primary preadipocytes to determine their transdifferentiation capacity to form bone-like tissues. Using lentivirus-derived particles to induce ectopic bone morphogenetic protein (BMP)-2 and delta FBJ murine osteosarcoma viral oncogene homolog B (DeltaFosB) gene expression, we demonstrated a time-dependent induction of osteoblast-specific genes and properties such as calcium deposits, bone-like extracellular matrix (ECM), and matrix mineralization. DeltaFosB was able to trigger partial Pref-1-mediated de-differentiation of adipocytes, which also retained their adipocytic cell phenotype. Osteoblast-specific structures could be co-localized in the ECM of lipid-containing cells analyzed using immunofluorescence and transmission electron microscopy when BMP-2 and DeltaFosB were co-expressed, suggesting that differentiated adipocytes are able to transdifferentiate into osteoblasts via a transient hybrid adipocyte-preadipocyte-osteoblast cell phenotype. Microtissues transgenic for BMP-2 and DeltaFosB expression were able to reproduce bone matrix, which occurs to a lesser extent in conventional two-dimensional (2D) cultures but is known to play a decisive role in the development and function of bone in vivo. This demonstrates that ECM-inclusive studies are essential for future characterization assays. Therefore, 3D cultures provide a superior ex vivo system for the improved characterization of phenotypical and functional alterations resulting from interventions directed toward differentiation processes. Precise control of transdifferentiation of adipocytes into osteoblasts in a 3D culture mimicking in vivo tissue conditions as closely as possible will foster important advances in regenerative medicine and tissue engineering.
authors.affiliation: Institute for Chemical and Bioengineering, ETH Zurich, Zurich, Switzerland.
authors.forename: Carlota
authors.initials: C
authors.lastname: Diaz Sanchez-Bustamante
authors.forename: Jens Maria
authors.initials: JM
authors.lastname: Kelm
authors.forename: Marcus
authors.initials: M
authors.lastname: Egermann
authors.forename: Valentin
authors.initials: V
authors.lastname: Djonov
authors.forename: Martin
authors.initials: M
authors.lastname: Fussenegger
mesh_terms: D017667:Adipocytes / Q000166:cytology* / Q000648:ultrastructure; D000818:Animals; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000378:metabolism; D002113:Calcification, Physiologic; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins; D054338:Cell Transdifferentiation; D005455:Fluorescent Antibody Technique; D020869:Gene Expression Profiling; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism; D050356:Lipid Metabolism; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D018613:Microscopy, Confocal; D010006:Osteoblasts / Q000166:cytology*; D016760:Proto-Oncogene Proteins c-fos / Q000378:metabolism*; D018874:Spheroids, Cellular / Q000166:cytology* / Q000648:ultrastructure; D013997:Time Factors; D016212:Transforming Growth Factor beta / Q000378:metabolism
chemical_list: C525718:BMP2 protein, human; C525719:Bmp2 protein, mouse; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins; D002135:Calcium-Binding Proteins; C512941:DLK1 protein, human; C064269:FOSB protein, human; C483819:Fosb protein, mouse; D036341:Intercellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D016760:Proto-Oncogene Proteins c-fos; D016212:Transforming Growth Factor beta; D002118:Calcium