Systems-based identification of temporal processing pathways during bone cell mechanotransduction.

Bone has long been established to be a highly mechanosensitive tissue. When subjected to mechanical loading, bone exhibits profoundly different anabolic responses depending on the temporal pattern in which the stimulus is applied. This phenomenon has been termed temporal processing, and involves complex signal amplification mechanisms that are largely unidentified. In this study, our goal was to characterize transcriptomic perturbations arising from the insertion of intermittent rest periods (a temporal variation with profound effects on bone anabolism) in osteoblastic cells subjected to fluid flow, and assess the utility of these perturbations to identify signaling pathways that are differentially activated by this temporal variation. At the level of the genome, we found that the common and differential alterations in gene expression arising from the two flow conditions were distributionally distinct, with the differential alterations characterized by many small changes in a large number of genes. Using bioinformatics analysis, we identified distinct up- and down-regulation transcriptomic signatures associated with the insertion of rest intervals, and found that the up-regulation signature was significantly associated with MAPK signaling. Confirming the involvement of the MAPK pathway, we found that the insertion of rest intervals significantly elevated flow-induced p-ERK1/2 levels by enabling a second spike in activity that was not observed in response to continuous flow. Collectively, these studies are the first to characterize distinct transcriptomic perturbations in bone cells subjected to continuous and intermittent stimulation, and directly demonstrate the utility of systems-based transcriptomic analysis to identify novel acute signaling pathways underlying temporal processing in bone cells.