New insights into BMP-7 mediated osteoblastic differentiation of primary human mesenchymal stem cells.
Bone Morphogenetic Proteins (BMPs) are members of the TGF-beta superfamily of growth factors. Several BMPs exhibit osteoinductive bioactivities, and are critical for bone formation in both developing and mature skeletal systems. BMP-7 (OP-1) is currently used clinically in revision of posterolateral spine fusions and long bone non-unions. The current study characterizes BMP-7 induced gene expression during early osteoblastic differentiation of human mesenchymal stem cells (hMSC). Primary hMSC were treated with BMP-7 for 24 or 120 h and gene expression across the entire human genome was evaluated using Affymetrix HG-U133 Plus 2.0 Arrays. 955 probe sets representing 655 genes and 95 ESTs were identified as differentially expressed and were organized into three major expression profiles (Profiles A, B and C) by hierarchical clustering. Genes from each profile were classified according to biochemical pathway analyses. Profile A, representing genes upregulated by BMP-7, revealed strong enrichment for established osteogenic marker genes, as well as several genes with undefined roles in osteoblast function, including MFI2, HAS3, ADAMTS9, HEY1, DIO2 and FGFR3. A functional screen using siRNA suggested roles for MFI2, HEY1 and DIO2 in osteoblastic differentiation of hMSC. Profile B contained genes transiently downregulated by BMP-7, including numerous genes associated with cell cycle regulation. Follow-up studies confirmed that BMP-7 attenuates cell cycle progression and cell proliferation during early osteoblastic differentiation. Profile C, comprised of genes continuously downregulated by BMP-7, exhibited strong enrichment for genes associated with chemokine/cytokine activity. Inhibitory effects of BMP-7 on cytokine secretion were verified by analysis of enriched culture media. Potent downregulation of CHI3L1, a potential biomarker for numerous joint diseases, was also observed in Profile C. A focused evaluation of BMP, GDF and BMP inhibitor expression elucidated feedback loops modulating BMP-7 bioactivity. BMP-7 was found to induce BMP-2 and downregulate GDF5 expression. Transient knockdown of BMP-2 using siRNA demonstrated that osteoinductive properties associated with BMP-7 are independent of endogenous BMP-2 expression. Noggin was identified as the predominant inhibitor induced by BMP-7 treatment. Overall, this study provides new insight into key bioactivities characterizing early BMP-7 mediated osteoblastic differentiation.
Lavery
Karen
K
Stryker Biotech, Hopkinton, MA 01748, USA.
Hawley
Sara
S
Swain
Pamela
P
Rooney
Robert
R
Falb
Dean
D
Alaoui-Ismaili
Moulay Hicham
MH
D054392:Adipokines; D000293:Adolescent; D000328:Adult; D015415:Biomarkers / Q000378:metabolism; D055396:Bone Morphogenetic Protein 2 / Q000235:genetics / Q000378:metabolism; D055419:Bone Morphogenetic Protein 7 / Q000494:pharmacology*; D002453:Cell Cycle / Q000187:drug effects / Q000235:genetics; D002454:Cell Differentiation / Q000187:drug effects*; D002478:Cells, Cultured; D000071451:Chitinase-3-Like Protein 1; D016000:Cluster Analysis; D016207:Cytokines / Q000235:genetics / Q000378:metabolism; D015536:Down-Regulation / Q000187:drug effects; D020869:Gene Expression Profiling; D006023:Glycoproteins / Q000235:genetics / Q000378:metabolism; D055428:Growth Differentiation Factor 5 / Q000235:genetics / Q000378:metabolism; D006801:Humans; D037102:Lectins; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D008875:Middle Aged; D010006:Osteoblasts / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects
D054392:Adipokines; D015415:Biomarkers; D055396:Bone Morphogenetic Protein 2; D055419:Bone Morphogenetic Protein 7; C084533:CHI3L1 protein, human; D000071451:Chitinase-3-Like Protein 1; D016207:Cytokines; D006023:Glycoproteins; D055428:Growth Differentiation Factor 5; D037102:Lectins; D034741:RNA, Small Interfering