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
Luciferase reporter assay
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
Casflox/flox and Relaflox/+ mice of C57BL/6 background (35, 36) were mated with Dmp1-Cre transgenic mice, in which the Cre recombinase gene was specifically expressed in osteocytes (37). Dmp1-Cre+/−, Casflox/flox (Cas cKO) and Dmp1-Cre−/− Casflox/flox (normal littermates) mice were generated by mating Dmp1-Cre−/−, Casflox/flox male mice with Dmp1-Cre+/−, Casflox/+ female mice. Dmp1-Cre+/−;Casflox/flox;Relaflox/+(Cas/Rela cKO) and Dmp1-Cre−/−;Casflox/flox;Relaflox/+ (normal littermates) mice were generated by mating Dmp1-Cre+/−;Casflox/flox;Rela +/+ male mice with Dmp1-Cre−/−;Casflox/flox;Relaflox/+ female mice. Dmp1-Cre+/–Relaflox/+ (Rela cKO) and Dmp1-Cre−/−;Relaflox/+ (normal littermates) mice were generated by mating Dmp1-Cre+/−;Rela +/+ male mice with Dmp1-Cre−/−;Relaflox/flox female mice. All animals were housed under specific pathogen–free conditions and treated with humane care under approval from the Animal Care and Use Committee of Tokyo Metropolitan Institute of Gerontology, National Rehabilitation Center for Persons with Disabilities, and Nadogaya Hospital. For all mouse analyses except for those at 1 day after birth (fig. S1), male mice were used.