Mechanical stretch inhibits chondrogenesis through ERK-1/2 phosphorylation in micromass culture

Abstract

Chondrocyte differentiation has been known to be affected by shearing stress, compressive or expansive force in vivo and in vitro. In the present study, we investigated the transient phosphorylation of extracellular signal-regulated kinase (ERK)-1/2 under stretch stimulation, and this ERK-1/2 phosphorylation mediates mechanical stretch signaling during chondrogenesis. Dissociated embryonic E12 rat limb bud cells were assembled to micromass culture on a silicon bottom plate. After 4 days, micromass cultures were stretched prior to the isolation of protein samples. Stretch stimulation was also loaded with or without MEK-1 or MEK-1/2 inhibitor. Western blot analysis revealed that phosphorylation of ERK-1/2 increased and peaked at 1.0 h after stretch loaded. Alcian blue staining and semi-quantitative reverse transcription-polymerase chain reaction analysis for type II collagen gene expression revealed that inhibited chondrogenesis by mechanical stretch stimulation was rescued by inhibiting MEK-1 and MEK-2 activities. It was concluded that signaling through ERK-1/2 was activated by stretch stimulation in micromass cultures and was involved in the inhibition of chondrogenesis by stretch stimulation.