Dimethyloxalylglycine Promotes Bone Marrow Mesenchymal Stem Cell Osteogenesis via Rho/ROCK Signaling.
OBJECTIVE
We investigated the role of dimethyloxalylglycine (DMOG) in bone marrow mesenchymal stem cell (BMSC) osteogenesis mediated by RhoA/ROCK.


METHODS
BMSCs were cultured with and without DMOG and/or Y-27632 (ROCK1 inhibitor). Cell proliferation, alkaline phosphatase (ALP) levels, and calcium deposits were determined. The expression of Runx2, OSX, p-cofilin, RhoA, and GTP-bound RhoA was determined by real-time RT-PCR and Western blot. Rho-associated coiled-coil-containing protein kinase (ROCK) activity was determined by measuring the phosphorylation of myosin-binding subunit of myosin phosphatase using an ELISA kit. Actin morphology was observed by immunofluorescence.


RESULTS
After 24 h, DMOG (0.5 mM) increased the expression of GTP-bound RhoA (+141%, P < 0.001) and enhanced ROCK activity (315%, P < 0.001). DMOG (0.5 mM) enhanced ALP levels after 3, 7, and 21 days of osteogenic induction (all P < 0.001) and strengthened calcium deposition (P < 0.001). In addition, compared with controls, DMOG (0.5 mM) increased the mRNA levels of osteogenesis genes RUNX2 and OSX (all P < 0.001). Furthermore, compared with controls, DMOG increased the expression of p-cofilin (+57%, P < 0.001), which resulted in rearrangement of actin filaments. All these effects were abolished, at least in part, by Y-27632.


CONCLUSIONS
DMOG promotes BMSC osteogenic differentiation via activation of RhoA/ROCK, suggesting clues for future therapies using BMSCs.
Department of rehabilitation medicine, the first people's hospital of Yunnan province, Kunming, China.
Lei
L
Zhang
Guoliang
G
Jiang
Xueling
X
Zhao
Yuekun
Y
Gong
D017667:Adipocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D000577:Amides / Q000494:pharmacology; D000600:Amino Acids, Dicarboxylic / Q000494:pharmacology*; D000818:Animals; D001854:Bone Marrow Cells / Q000187:drug effects / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D019902:Chondrocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D051338:Cofilin 1 / Q000235:genetics / Q000378:metabolism; D050659:Core Binding Factor Alpha 1 Subunit / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D008297:Male; D059630:Mesenchymal Stem Cells / Q000187:drug effects* / Q000378:metabolism; D051379:Mice; D043263:Myosin-Light-Chain Phosphatase / Q000235:genetics / Q000378:metabolism; D010006:Osteoblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D010012:Osteogenesis / Q000187:drug effects* / Q000235:genetics; D061251:Primary Cell Culture; D011725:Pyridines / Q000494:pharmacology; D015398:Signal Transduction; D000076182:Sp7 Transcription Factor; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D020741:rho GTP-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D054460:rho-Associated Kinases / Q000235:genetics* / Q000378:metabolism; D020742:rhoA GTP-Binding Protein
D000577:Amides; D000600:Amino Acids, Dicarboxylic; D051338:Cofilin 1; D050659:Core Binding Factor Alpha 1 Subunit; D011725:Pyridines; C493716:Runx2 protein, mouse; D000076182:Sp7 Transcription Factor; C479466:Sp7 protein, mouse; D014157:Transcription Factors; C108830:Y 27632; C515854:Rock1 protein, mouse; D054460:rho-Associated Kinases; D043263:Myosin-Light-Chain Phosphatase; C526380:RhoA protein, mouse; D020741:rho GTP-Binding Proteins; D020742:rhoA GTP-Binding Protein; D002118:Calcium; C040947:oxalylglycine