The effect of substratum topography on osteoblast adhesion mediated signal transduction and phosphorylation

Substratum surface topography is a powerful modulator of cell behaviour, but how it influences intracellular signaling is largely unknown. We investigated the influence of microfabricated topographies on the activation of nonreceptor tyrosine kinases Src, FAK and ERK 1/2, as well as the transcription factor, Runx2, in rat osteoblasts, cultured on substrata that varied in their ability to promote bone-like tissue formation. Total tyrosine phosphorylation increased on grooves, tapered pits, and gap cornered boxes, relative to the levels found on smooth surfaces, with the greatest activity at 1 week. Src levels was higher on smooth than on any other surface, but FAK and ERK 1/2 phosphorylation were highest on groove and gap-cornered boxes up to 6 weeks. Inhibition of Src phosphorylation with PP2 inhibited FAK and ERK 1/2 phosphorylation on grooves, but had no detectable effect on either FAK or ERK 1/2 on smooth substratum. We suggest that osteoblast response to substrata with specific topographical features requires FAK–Y397–Src–Y416 complexes for ERK 1/2 phosphorylation, but on smooth surfaces, Src independent methods of ERK 1/2 activation are present.