Mice
Cell lines
Analysis of bone phenotype
Plasmids
Antibodies
Cellular fractionation
Immunofluorescence staining
Coimmunoprecipitation to analyze association with p300
RNA extraction and real-time reverse transcription polymerase chain reaction analysis
Primer sequences used for PCR
RNA interference
Biochemical analysis of osteocytes in vivo and in vitro
Measurement of intracellular ROS level
FSS experiments
ChIP assay
Hemilateral hindlimb unloading experiments
Statistical analysis
NF-κB–responsive reporter plasmid (pNF-κB Luc) and Renilla luciferase expression plasmid (phRL-TK) were cotransfected with the indicated expression plasmids into HEK293 cells. Twenty-four hours after transfection, cells were subjected to experiments. Luciferase activity was measured using the Dual-Glo Luciferase Assay System (Promega, Madison, WI, USA). Specific NF-κB Luc activity in individual samples was determined by normalizing firefly luciferase activity to Renilla luciferase activity. For all assays, experiments were performed in triplicate.