Regulation of BMP-induced transcription in cultured human bone marrow stromal cells.
BACKGROUND
Adherent bone marrow stromal cells are inducible osteoprogenitors, giving rise to cells expressing osteoblast markers including alkaline phosphatase, osteopontin, osteocalcin, and bone sialoprotein. However, the potency of inducers varies in a species-specific manner. Glucocorticoids such as dexamethasone induce alkaline phosphatase activity in both human and rat mesenchymal stem cells, while mouse bone marrow stromal cells are refractory to dexamethasone-induced alkaline phosphatase activity. In contrast, BMP induces alkaline phosphatase activity in both mouse and rat bone marrow stromal cells, while BMP effects on human bone marrow stromal cells are poorly characterized.


METHODS
Bone marrow samples were isolated from patients undergoing hip replacement. Mononuclear marrow cells were cultured and grown to confluence without or with 10 (-7) M dexamethasone. Cells from each isolate were passaged into medium containing 100 micro g/mL ascorbate phosphate and treated with dexamethasone, 100 ng/mL BMP, or no inducer. At day 6, alkaline phosphatase activity was assayed, and RNA was prepared for mRNA analyses by real-time polymerase chain reaction.


RESULTS
Bone marrow stromal cells from twenty-four of twenty-six patients showed no significant osteogenic response to BMP-2, 4, or 7 as determined by alkaline phosphatase induction. However, BMPs induced elevated levels of other genes associated with osteogenesis such as bone sialoprotein and osteopontin as well as BMP-2 and noggin. If primary cultures of human bone marrow stromal cells were pretreated with dexamethasone, BMP-2 treatment of first-passage cells induced alkaline phosphatase in approximately half of the isolates, and significantly greater induction was seen in cells from males. Dexamethasone treatment, like BMP treatment, also increased expression of the BMP-binding protein noggin.


CONCLUSIONS
Most human femur bone marrow stromal cell samples appear incapable of expressing elevated alkaline phosphatase levels in response to BMPs. Since BMP treatment induced expression of several other BMP-regulated genes, the defect in alkaline phosphatase induction is presumably not due to impaired BMP signaling. We hypothesize that the mechanism by which BMPs modulate alkaline phosphatase expression is indirect, involving a BMP-regulated transcription factor for alkaline phosphatase expression that is controlled differently in humans and rodents.
Diefenderfer
David L
DL
Osyczka
Anna M
AM
Garino
Jonathan P
JP
Leboy
Phoebe S
PS
D000469:Alkaline Phosphatase / Q000235:genetics / Q000378:metabolism; D001854:Bone Marrow Cells / Q000166:cytology* / Q000187:drug effects; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000235:genetics / Q000494:pharmacology*; D002352:Carrier Proteins; D002454:Cell Differentiation / Q000187:drug effects / Q000235:genetics*; D002478:Cells, Cultured; D003907:Dexamethasone / Q000494:pharmacology; D004790:Enzyme Induction / Q000187:drug effects / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D006801:Humans; D009363:Neoplasm Proteins; D010012:Osteogenesis / Q000187:drug effects / Q000235:genetics*; D016133:Polymerase Chain Reaction; D011506:Proteins / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D013234:Stem Cells / Q000166:cytology* / Q000187:drug effects; D017154:Stromal Cells / Q000166:cytology* / Q000187:drug effects; D014157:Transcription Factors / Q000235:genetics; D014158:Transcription, Genetic / Q000187:drug effects*; D016212:Transforming Growth Factor beta; D000099176:Noggin Protein
D000469:Alkaline Phosphatase; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins; D002352:Carrier Proteins; D003907:Dexamethasone; D009363:Neoplasm Proteins; D011506:Proteins; D012333:RNA, Messenger; D014157:Transcription Factors; D016212:Transforming Growth Factor beta; D000099176:Noggin Protein; C525718:BMP2 protein, human; C525719:Bmp2 protein, mouse; C525720:Bmp2 protein, rat