A pterostilbene derivative suppresses osteoclastogenesis by regulating RANKL-mediated NFκB and MAPK signaling in RAW264.7 cells.
BACKGROUND
A dysfunctional osteoclast activity is often the cause of bone destructive diseases, such as osteoporosis, periodontitis, erosive arthritis, and cancer. The NFκB ligand (RANKL) has been identified as a major mediator of bone resorption. Agents that suppress RANKL signaling have the potential to inhibit bone resorption or osteoclastogenesis. The present study aimed to determine the effect of a pterostilbene derivative (PTERC-T) for suppressing RANKL or tumor cells-induced osteoclastogenesis in RAW264.7 murine macrophages.


METHODS
Cytotoxicity was measured by MTT assay and inhibitory effect on osteoclastogenesis was analyzed by counting the number of tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells and measuring the expression levels of the osteoclast-specific genes. The reactive oxygen species (ROS) generation was detected by FACS. Further, signaling pathways were analyzed by immunofluorescence and immunoblot analyses.


RESULTS
PTERC-T suppressed the differentiation of monocytes to osteoclasts in a dose and time-dependent manner. The expression of osteoclast marker genes like TRAP, cathepsin K (CTSK), matrix metalloproteinase 9 (MMP9) and transcription factors c-Fos, and nuclear factor of activated T cells cytoplasmic 1 (NFATc1) were also diminished by PTERC-T. PTERC-T scavenged intracellular ROS generation within osteoclast precursors during RANKL-stimulated osteoclastogenesis. Mechanistically, PTERC-T abrogated the phosphorylation of MAPKs (ERK and JNK) and inhibited RANKL-induced activation of NFκB by suppressing IκBα phosphorylation and preventing NFκB/p65 nuclear translocation.


CONCLUSIONS
This study thus identifies PTERC-T as an inhibitor of osteoclast formation and provides evidence for its role in preventing osteoporosis and other bone related disorders. However, further studies are needed to establish its efficacy in vivo.
Molecular Endocrinology Laboratory, Department of Biotechnology, Indian Institute of Technology Roorkee, Roorkee, India.
Kumar
K
Nikhil
Molecular Endocrinology Laboratory, Department of Biotechnology, Indian Institute of Technology Roorkee, Roorkee, India.
Shruti
S
Sharan
Molecular Endocrinology Laboratory, Department of Biotechnology, Indian Institute of Technology Roorkee, Roorkee, India. Electronic address: paroyfbs@iitr.ernet.in.
Partha
P
Roy
D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism*; D010010:Osteoclasts / Q000166:cytology* / Q000187:drug effects*; D053245:RANK Ligand / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000067996:RAW 264.7 Cells; D017382:Reactive Oxygen Species / Q000378:metabolism; D013267:Stilbenes / Q000737:chemistry* / Q000494:pharmacology; D013882:Thiosemicarbazones / Q000494:pharmacology*; D014157:Transcription Factors / Q000378:metabolism
D016328:NF-kappa B; D053245:RANK Ligand; D017382:Reactive Oxygen Species; D013267:Stilbenes; D013882:Thiosemicarbazones; D014157:Transcription Factors; C000610514:pterostilbene carboxaldehyde thiosemicarbazone; D020928:Mitogen-Activated Protein Kinases
MAPK; NFκB; Osteoclastogenesis; PTERC-T; RANKL