title: Preliminary evidence for the dedifferentiation of RAW 264.7 cells into mesenchymal progenitor-like cells by a purine analog.
abstract: Dedifferentiation of cells to multipotential cells is of interest since they have a potential regenerative capacity. Our purpose was to de- and redifferentiate murine RAW 264.7 cells, a committed macrophage cell line of hematopoietic origin, into mesenchymal-like cells such as osteoblasts. RAW 264.7 cells in culture were treated with 5 μM reversine, a purine analog that was shown to dedifferentiate myoblasts in osteoblasts. Treatment with reversine resulted in a significant increase in the expression of the STRO-1 antigen, a marker of mesenchymal stem/progenitor cells: from 0.6%±0.5% cells in untreated RAW cells to 19.0%±8.6% in treated cells, but there was no increase in the expression of SH-2 (CD105), an earlier marker of mesenchymal stem cells. The effects of reversine were significantly curtailed by 67% when cultures were pretreated with the c-Jun N-terminal kinase pathway blocker SP600125. These STRO-1+ cells retained a multipotential status and were capable of redifferentiating into cells with osteogenic and lipogenic characteristics under inductive conditions. We showed that STRO-1+ cells in an osteogenic medium significantly increased expression of the osteoblast marker osteocalcin, and formed mineralized nodules. When seeded on a demineralized scaffold of human bone in vitro, these cells deposited a calcium matrix. Under adipogenic conditions, expression of the adipocyte marker peroxisome proliferator-activated receptor gamma 2 on STRO-1+ cells was elevated, and cultures stained positive with Oil red O. Our results demonstrated that treating a committed hematopoietic cell line with a purine analog can alter cell development and result in cellular reverse transformation into stage-limited multipotential cells. These cells could subsequently be redifferentiated into cells with characteristics of the mesenchymal lineage, such as those of an osteoblast and/or adipocyte, under inductive conditions.
authors.affiliation: Faculty of Dentistry, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Guowei
authors.initials: G
authors.lastname: Qu
authors.forename: Herbert P
authors.initials: HP
authors.lastname: von Schroeder
mesh_terms: D050156:Adipogenesis / Q000187:drug effects; D000818:Animals; D000873:Anthracenes / Q000494:pharmacology; D000954:Antigens, Surface / Q000378:metabolism; D001842:Bone and Bones / Q000000981:diagnostic imaging / Q000187:drug effects; D002232:Carbocyanines / Q000378:metabolism; D054337:Cell Dedifferentiation / Q000187:drug effects*; D002460:Cell Line; D000071063:Endoglin; D005434:Flow Cytometry; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D048031:JNK Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D008264:Macrophages / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D008856:Microscopy, Fluorescence; D009025:Morpholines / Q000494:pharmacology; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011687:Purines / Q000494:pharmacology*; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013194:Staining and Labeling; D054457:Tissue Scaffolds; D055114:X-Ray Microtomography
chemical_list: D000873:Anthracenes; D000954:Antigens, Surface; D002232:Carbocyanines; D000071063:Endoglin; C502004:Eng protein, mouse; D047908:Intracellular Signaling Peptides and Proteins; D009025:Morpholines; D047428:Protein Kinase Inhibitors; D011687:Purines; C525590:STRO-1 antigen, mouse; C085321:cyanine dye 5; C432165:pyrazolanthrone; D048031:JNK Mitogen-Activated Protein Kinases; C484369:2-(4-morpholinoanilino)-6-cyclohexylaminopurine