Integrin β₁ silencing in osteoblasts alters substrate-dependent responses to 1,25-dihydroxy vitamin D₃

Surface microroughness increases osteoblast differentiation and enhances responses of osteoblasts to 1,25-dihydroxyvitamin D₃ [1α,25(OH)₂D₃]. The observations that β₁ integrin expression is increased in osteoblasts grown on Ti substrates with rough microarchitecture, and that it is regulated by 1α,25(OH)₂D₃ in a surface-dependent manner, suggest that β₁ may play a role in mediating osteoblast response. To test this hypothesis, we silenced β₁ expression in MG63 human osteoblast-like cells using small interfering RNA (siRNA) and examined the responses of the β₁-silenced osteoblasts to surface microtopography and 1α,25(OH)₂D₃. To better understand the role of β₁, MG63 cells were also treated with two different monoclonal antibodies to human β₁ to block ligand binding. β₁-silenced MG63 cells grown on a tissue culture plastic had reduced alkaline phosphatase activity and levels of osteocalcin, transforming growth factor β1, prostaglandin E₂, and osteoprotegerin in comparison with control cells. Moreover, β₁-silencing inhibited the effects of surface roughness on these parameters and partially inhibited effects of 1α,25(OH)₂D₃. Anti β₁ antibody AIIB2 had no significant effect on cell number and osteocalcin, but decreased alkaline phosphatase; MAB2253Z caused dose-dependent decreases in cell number and alkaline phosphatase and an increase in osteocalcin. Effects of 1α,25(OH)₂D₃ on cell number and alkaline phosphatase were reduced and effects on osteocalcin were increased. These findings indicate that β₁ plays a major and complex role in osteoblastic differentiation modulated by either surface microarchitecture or 1α,25(OH)₂D₃. The results also show that β₁ mediates, in part, the synergistic effects of surface roughness and 1α,25(OH)₂D₃.