Rac1 Mediates Load-Driven Attenuation of mRNA Expression of Nerve Growth Factor Beta in Cartilage and Chondrocytes

Abstract

Objectives: To determine effect of gentle loads applied to the knee on mRNA expression of nerve growth factor, particularly, the active beta subunit (NGFβ) in cartilage and chondrocyte. Methods: Cyclic compressive loads in vivo and fluid flow in vitro were used to determine the mRNA levels. Alteration of Rac1 GTPase as well as effect of salubrinal, a specific inhibitor of eIF2α phosphatase was assessed using fluorescence resonance energy transfer (FRET)-based Rac1 biosensor. Results: Knee loading at 1 N reduced mRNA levels of NGFβ and its low affinity receptor, p75 in cartilage and subchondral bone. In cartilage, knee loading at 1 N reduced the phosphorylation level of p38 MAPK (p38-p) and activity of Rac1 GTPase. Consistent with in vivo results, fluid flow at 5 and 10 dyn/cm(2) reduced mRNA levels of NGFβ and p75 in C28/I2 human chondrocytes. SB203580, which decreases p38-p, reduced the mRNA levels of NGFβ and p75. Silencing Rac1 by siRNA decreased the levels of p38-p and NGFβ mRNA but not p75. Furthermore, administration of salubrinal reduced FRET-based activity of Rac1 as well as the mRNA levels of NGFβ and p75. Conclusions: These results provide evidence that mechanical stimulation and salubrinal may attenuate pain perception-linked NGFβ signaling through Rac1-mediated p38 MAPK.