Activation of RhoA and FAK induces ERK-mediated osteopontin expression in mechanical force-subjected periodontal ligament fibroblasts

Abstract

The precise mechanism by which Rho kinase translates the mechanical signals into OPN up-regulation in force-exposed fibroblasts has not been elucidated. Human periodontal ligament fibroblasts (hPLFs) were exposed to mechanical force by centrifuging the culture plates at a magnitude of 50 g/cm(2) for 60 min. At various times of the force application, they were processed for analyzing cell viability, trypan blue exclusion, and OPN expression at protein and RNA levels. Cellular mechanism(s) of the force-induced OPN up-regulation was also examined using various kinase inhibitors or antisense oligonucleotides specific to mechanosensitive factors. Centrifugal force up-regulated OPN expression and induced a rapid and transient increase in the phosphorylation of focal adhesion kinase (FAK), extracellular signal-regulated kinase (ERK), and Elk1. Pharmacological blockade of RhoA/Rho-associated coiled coil-containing kinase (ROCK) signaling markedly reduced force-induced FAK and ERK1/2 phosphorylation. Transfecting hPLFs with FAK antisense oligonucleotide diminished ERK1/2 activation and force-induced OPN expression. Further, ERK inhibitor inhibited significantly OPN expression, Elk1 phosphorylation, and activator protein-1 (AP-1)-DNA binding activation, but not FAK phosphorylation, in the force-applied cells. These results demonstrate that FAK signaling plays critical roles in force-induced OPN expression in hPLFs through interaction with Rho/ROCK as upstream effectors and ERK-Elk1/ERK-c-Fos as downstream effectors.

Mechanosensitive mechanisms in transcriptional regulation

“…Mechanically activated ERK1/2 and/or JNK signals are transmitted into the nucleus where they activate the transcription factor AP1 and, thereby, upregulate expression of molecules that are crucial for tissue formation and remodeling, such as type I collagen and osteopontin in bone (Hong et al, 2010;Jeon et al, 2009;Kook et al, 2009). Mechanical strain and distortion of the cell shape also activate membrane-associated phospholipases and, thereby, increase the metabolism of inositol lipids and arachidonic acid in the cytoplasm, which results in the release of Ca 2+ from intracellular stores, activation of protein kinase C (PKC) and the remodeling of cardiomyocytes through activation of mechanosensitive transcription factors, such as EGR1 and AP1 (Fig.…”

Modulation of microenvironment for controlling the fate of periodontal ligament cells: the role of Rho/ROCK signaling and cytoskeletal dynamics

“…It has been suggested that mechanical stress-mediated Rho/ROCK signaling is linked to the expression of a series of osteogenic molecules, such as activator protein-1, osteopontin, and receptor activator of nuclear factor-kappa B ligand, in PDL cells (Kletsas et al 2002;Yamashiro et al 2007;Wongkhantee et al 2008;Hong et al 2010;Pan et al 2014). A better understanding of how PDL cells adapt to the mechanical stress may contribute to the analysis of their osteogenic lineage.…”

Cyclic Tensile Stress During Physiological Occlusal Force Enhances Osteogenic Differentiation of Human Periodontal Ligament Cells via ERK1/2-Elk1 MAPK Pathway

“…The direct involvement of MAPKs on mechano transduction has been largely suggested in muscle cells (Li et al, 2000;Zampetaki et al, 2005) and in bone cells (Franceschi and Xiao, 2003;Liedert et al, 2006). In periodontium, centrifugal force during OTM up-regulated COL I expression in human periodontal fibroblasts through the activation of MAPK-AP-1signaling (Kook et al, 2009;Hong et al, 2010). However, little knowledge is available about the mechanism of physiological occlusal force on hPDLCs.…”

“…Human PDL fibroblasts stimulate osteoclastogenesis in response to compression force through TNF-alpha-mediated activation of CD4 + T cells (Kook et al, 2011b). Osteopontin expression in mechanical force-subjected PDL fibroblasts was activated by the ERK pathway (Hong et al, 2010). The inconsistent findings show that the mechanisms involved in force-applied hPDLCs also depend on the culture conditions and stress regime.…”