title: Establishment of Immortalized BMP2/4 Double Knock-Out Osteoblastic Cells Is Essential for Study of Osteoblast Growth, Differentiation, and Osteogenesis.
abstract: Bone morphogenetic proteins 2 and 4 (BMP2/4) are essential for osteoblast differentiation and osteogenesis. Generation of a BMP2/4 dual knock-out ((ko/ko)) osteoblastic cell line is a valuable asset for studying effects of BMP2/4 on skeletal development. In this study, our goal was to create immortalized mouse deleted BMP2/4 osteoblasts by infecting adenoviruses with Cre recombinase and green fluorescent protein genes into immortalized murine floxed BMP2/4 osteoblasts. Transduced BMP2/4(ko/ko) cells were verified by green immunofluorescence and PCR. BMP2/4(ko/ko) osteoblasts exhibited small size, slow cell proliferation rate and cell growth was arrested in G1 and G2 phases. Expression of bone-relate genes was reduced in the BMP2/4(ko/ko) cells, resulting in delay of cell differentiation and mineralization. Importantly, extracellular matrix remodeling was impaired in the BMP2/4(ko/ko) osteoblasts as reflected by decreased Mmp-2 and Mmp-9 expressions. Cell differentiation and mineralization were rescued by exogenous BMP2 and/or BMP4. Therefore, we for the first time described establishment of an immortalized deleted BMP2/4 osteoblast line useful for study of mechanisms in regulating osteoblast lineages.
authors.affiliation: Department of Developmental Dentistry, The University of Texas Health Science Center at San Antonio, Texas.|Department of Pediatric Dentistry, School of Stomatology, The Fourth Military Medical University, Xi-an, China.
authors.forename: Li-An
authors.initials: LA
authors.lastname: Wu
authors.affiliation: Department of Developmental Dentistry, The University of Texas Health Science Center at San Antonio, Texas.|Department of Anatomy, Histology & Embryology, Basic Medical College, Fujian Medical University, Fuzhou, China.
authors.forename: Feng
authors.initials: F
authors.lastname: Wang
authors.affiliation: Department of Developmental Dentistry, The University of Texas Health Science Center at San Antonio, Texas.
authors.forename: Kevin J
authors.initials: KJ
authors.lastname: Donly
authors.affiliation: Department of Developmental Dentistry, The University of Texas Health Science Center at San Antonio, Texas.
authors.forename: Andrew
authors.initials: A
authors.lastname: Baker
authors.affiliation: Department of Developmental Dentistry, The University of Texas Health Science Center at San Antonio, Texas.
authors.forename: Chunyan
authors.initials: C
authors.lastname: Wan
authors.affiliation: Department of Developmental Dentistry, The University of Texas Health Science Center at San Antonio, Texas.
authors.forename: Daoshu
authors.initials: D
authors.lastname: Luo
authors.affiliation: Department of Oral/Maxillofacial Surgery, University of Alabama at Birmingham, School of Dentistry, Birmingham, Alabama.
authors.forename: Mary
authors.initials: M
authors.lastname: MacDougall
authors.affiliation: Department of Developmental Dentistry, The University of Texas Health Science Center at San Antonio, Texas.
authors.forename: Shuo
authors.initials: S
authors.lastname: Chen
mesh_terms: D000818:Animals; D055396:Bone Morphogenetic Protein 2 / Q000172:deficiency* / Q000235:genetics; D055415:Bone Morphogenetic Protein 4 / Q000172:deficiency* / Q000235:genetics; D002454:Cell Differentiation; D002460:Cell Line; D049109:Cell Proliferation; D048429:Cell Size; D005109:Extracellular Matrix / Q000378:metabolism; D059585:G1 Phase Cell Cycle Checkpoints; D059565:G2 Phase Cell Cycle Checkpoints; D005786:Gene Expression Regulation; D055785:Gene Knockdown Techniques; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D019426:Integrases / Q000235:genetics / Q000378:metabolism; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D051379:Mice; D010006:Osteoblasts; D010012:Osteogenesis; D010641:Phenotype; D013997:Time Factors; D014161:Transduction, Genetic
chemical_list: C525719:Bmp2 protein, mouse; C525810:Bmp4 protein, mouse; D055396:Bone Morphogenetic Protein 2; D055415:Bone Morphogenetic Protein 4; D049452:Green Fluorescent Proteins; C045073:Cre recombinase; D019426:Integrases; D020778:Matrix Metalloproteinase 2; C522362:Mmp2 protein, mouse; D020780:Matrix Metalloproteinase 9; C501830:Mmp9 protein, mouse